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SUGASAWA KaoruBiosignal Research CenterProfessor
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■ Research news- 08 Dec. 2020, New findings shed light on the repair of UV-induced DNA damage -The impact of the protein degradation system on DNA damage recognition and repair-
- 27 Apr. 2016, Mechanism revealed for repairing DNA damaged by UV radiation
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■ Paper- The production of reactive oxygen species in response to RNA virus infection results in the oxidation of viral genomic RNA within infected cells. These oxidative RNA lesions undergo replication catalyzed by the viral replisome. G to U transversion mutations are frequently observed in the SARS-CoV-2 genome and may be linked to the replication process catalyzed by RNA-dependent RNA polymerase (RdRp) past the oxidative RNA lesion 7,8-dihydro-8-oxo-riboguanosine (8-oxo-rG). To better understand the mechanism of viral RNA mutagenesis, it is crucial to elucidate the role of RdRp in replicating across oxidative lesions. In this study, we investigated the RNA synthesis catalyzed by the reconstituted SARS-CoV-2 RdRp past a single 8-oxo-rG. The RdRp-mediated primer extension was significantly inhibited by 8-oxo-rG on the template RNA. A steady-state multiple-turnover reaction demonstrated that the turnover rate of RdRp was significantly slow when replication was blocked by 8-oxo-rG, reflecting low bypass efficiency even with prolonged reaction time. Once RdRp was able to bypass 8-oxo-rG, it preferentially incorporated rCMP, with a lesser amount of rAMP opposite 8-oxo-rG. In contrast, RdRp demonstrated greater activity in extending from the mutagenic rA:8-oxo-rG terminus compared to the lower efficiency of extension from the rC:8-oxo-rG pair. Based on steady-state kinetic analyses for the incorporation of rNMPs opposite 8-oxo-rG and chain extension from rC:8-oxo-rG or rA:8-oxo-rG, the relative bypass frequency for rA:8-oxo-rG was found to be seven-fold higher than that for rC:8-oxo-rG. Therefore, the properties of RdRp indicated in this study may contribute to the mechanism of mutagenesis of the SARS-CoV-2 genome.Apr. 2025, The Journal of biological chemistry, 108512 - 108512, English, International magazineScientific journal
- Summary Fanconi anemia (FA) is a rare genetic disease caused by the loss of function of one of the 22 associated genes and is characterized by bone marrow failure, cancer predisposition, and developmental defects. The proteins encoded by these genes (FA proteins) mainly function in DNA damage response and repair. Although FA deficiency has multiple effects on the regulation of lipid metabolism, the molecular function of FA proteins in the context of FA pathology remains unclear. In the present study, we demonstrated that FANCD2, a key component of FA proteins, interacts with lipid metabolism-related factors and that FANCD2 deficiency downregulates the cellular levels of fatty acids. Moreover, a portion of FANCD2 is localized to nuclear lipid droplets in response to oleic acid treatment. These subcellular dynamics are independent of FANCD2 monoubiquitination, which is essential for the DNA damage response. Collectively, these findings demonstrate that FANCD2 responds to not only DNA damage but also oleic acid exposure, providing insights into the pathogenesis of lipid dysregulation in FA.Cold Spring Harbor Laboratory, Feb. 2025
- Feb. 2025, 生化学, 97(1) (1), 61 - 65, Japaneseヌクレオチド除去修復のDNA損傷認識を制御するクロマチンダイナミクス[Refereed]Scientific journal
- How the fate (folding versus degradation) of glycoproteins is determined in the endoplasmic reticulum (ER) is an intriguing question. Monoglucosylated glycoproteins are recognized by lectin chaperones to facilitate their folding, whereas glycoproteins exposing well-trimmed mannoses are subjected to glycoprotein ER-associated degradation (gpERAD); we have elucidated how mannoses are sequentially trimmed by EDEM family members (George et al., 2020; 2021 eLife). Although reglucosylation by UGGT was previously reported to have no effect on substrate degradation, here we directly tested this notion using cells with genetically disrupted UGGT1/2. Strikingly, the results showed that UGGT1 delayed the degradation of misfolded substrates and unstable glycoproteins including ATF6α. An experiment with a point mutant of UGGT1 indicated that the glucosylation activity of UGGT1 was required for the inhibition of early glycoprotein degradation. These and overexpression-based competition experiments suggested that the fate of glycoproteins is determined by a tug-of-war between structure formation by UGGT1 and degradation by EDEMs. We further demonstrated the physiological importance of UGGT1, since ATF6α cannot function properly without UGGT1. Thus, our work strongly suggests that UGGT1 is a central factor in ER protein quality control via the regulation of both glycoprotein folding and degradation.Dec. 2024, eLife, 12, English, International magazineScientific journal
- It remains uncertain how global genome nucleotide excision repair (GG-NER) efficiently removes various helix distorting DNA lesions in the cell nucleus. Here, we present a protocol to assess the contribution of factors of interest to GG-NER using two types of fluorescence-microscopy-based techniques. First, we describe steps for analyzing the localization of the factors upon local ultraviolet (UV) irradiation. We then detail the second technique, which quantifies the removal of UV-induced photolesions combined with lesion-specific antibodies and program-based image analysis. For complete details on the use and execution of this protocol, please refer to Kusakabe et al.1.Jun. 2023, STAR protocols, 4(3) (3), 102378 - 102378, English, International magazineScientific journal
- Elsevier BV, Jun. 2023, Journal of Biological Chemistry, 105002 - 105002Scientific journal
- Springer Science and Business Media LLC, Apr. 2023, Nature, 617(7959) (7959), 170 - 175Scientific journal
- Nov. 2022, The Journal of Dermatology, 50(3) (3), 407 - 408, English, International magazine[Refereed]Scientific journal
- The XPC protein complex plays a central role in DNA lesion recognition for global genome nucleotide excision repair (GG-NER). Lesion recognition can be accomplished in either a UV-DDB-dependent or -independent manner; however, it is unclear how these sub-pathways are regulated in chromatin. Here, we show that histone deacetylases 1 and 2 facilitate UV-DDB-independent recruitment of XPC to DNA damage by inducing histone deacetylation. XPC localizes to hypoacetylated chromatin domains in a DNA damage-independent manner, mediated by its structurally disordered middle (M) region. The M region interacts directly with the N-terminal tail of histone H3, an interaction compromised by H3 acetylation. Although the M region is dispensable for in vitro NER, it promotes DNA damage removal by GG-NER in vivo, particularly in the absence of UV-DDB. We propose that histone deacetylation around DNA damage facilitates the recruitment of XPC through the M region, contributing to efficient lesion recognition and initiation of GG-NER.Elsevier BV, Apr. 2022, iScience, 25(4) (4), 104040 - 104040, English, International magazine[Refereed]Scientific journal
- SIRT2 and SIRT3 protein deacetylases maintain genome integrity and stability. However, their mechanisms for maintaining the genome remain unclear. To examine the roles of SIRT2 and SIRT3 in DSB repair, I-SceI-based GFP reporter assays for HR, single-strand annealing (SSA) and nonhomologous end joining (NHEJ) repair were performed under SIRT2- or SIRT3-depleted conditions. SIRT2 or SIRT3 depletion inhibited HR repair equally to RAD52 depletion, but did not affect SSA and NHEJ repairs. SIRT2 or SIRT3 depletion disturbed the recruitment of RAD51 to DSB sites, an essential step for RAD51-dependent HR repair, but not directly through RAD52 deacetylation. SIRT2 or SIRT3 depletion decreased the colocalization of γH2AX foci with RPA1, and thus, they might be involved in initiating DSB end resection for the recruitment of RAD51 to DSB sites at an early step in HR repair. These results show the novel underlying mechanism of the SIRT2 and SIRT3 functions in HR for genome stability.Feb. 2021, Genes to cells : devoted to molecular & cellular mechanisms, 26(5) (5), 328 - 335, English, International magazineScientific journal
- Nucleotide excision repair (NER) is a pathway involved in the repair of a variety of potentially mutagenic lesions that distort the DNA double helix. The ubiquitin E3-ligase complex UV-DDB is required for the recognition and repair of UV-induced cyclobutane pyrimidine dimers (CPDs) lesions through NER. DDB2 directly binds CPDs and subsequently undergoes ubiquitination and proteasomal degradation. DDB2 must remain on damaged chromatin, however, for sufficient time to recruit and hand-off lesions to XPC, a factor essential in the assembly of downstream repair components. Here we show that the tumor suppressor USP44 directly deubiquitinates DDB2 to prevent its premature degradation and is selectively required for CPD repair. Cells lacking USP44 have impaired DDB2 accumulation on DNA lesions with subsequent defects in XPC retention. The physiological importance of this mechanism is evident in that mice lacking Usp44 are prone to tumors induced by NER lesions introduced by DMBA or UV light. These data reveal the requirement for highly regulated ubiquitin addition and removal in the recognition and repair of helix-distorting DNA damage and identify another mechanism by which USP44 protects genomic integrity and prevents tumors.2021, Frontiers in cell and developmental biology, 9, 663411 - 663411, English, International magazineScientific journal
- Squamous cell carcinoma (SCC) occurs frequently in the human Xeroderma Pigmentosum (XP) syndrome and is characterized by deficient UV-damage repair. SCC is the most common equine ocular cancer and the only associated genetic risk factor is a UV-damage repair protein. Specifically, a missense mutation in horse DDB2 (T338M) was strongly associated with both limbal SCC and third eyelid SCC in three breeds of horses (Halflinger, Belgian, and Rocky Mountain Horses) and was hypothesized to impair binding to UV-damaged DNA. Here, we investigate DDB2-T338M mutant's capacity to recognize UV lesions in vitro and in vivo, together with human XP mutants DDB2-R273H and -K244E. We show that the recombinant DDB2-T338M assembles with DDB1, but fails to show any detectable binding to DNA substrates with or without UV lesions, due to a potential structural disruption of the rigid DNA recognition β-loop. Consistently, we demonstrate that the cellular DDB2-T338M is defective in its recruitment to focally radiated DNA damages, and in its access to chromatin. Thus, we provide direct functional evidence indicating the DDB2-T338M recapitulates molecular defects of human XP mutants, and is the causal loss-of-function allele that gives rise to equine ocular SCCs. Our findings shed new light on the mechanism of DNA recognition by UV-DDB and on the initiation of ocular malignancy.Jan. 2021, DNA repair, 97, 103022 - 103022, English, International magazineScientific journal
- The ubiquitin-proteasome system (UPS) plays crucial roles in regulation of various biological processes, including DNA repair. In mammalian global genome nucleotide excision repair (GG-NER), activation of the DDB2-associated ubiquitin ligase upon UV-induced DNA damage is necessary for efficient recognition of lesions. To date, however, the precise roles of UPS in GG-NER remain incompletely understood. Here, we show that the proteasome subunit PSMD14 and the UPS shuttle factor RAD23B can be recruited to sites with UV-induced photolesions even in the absence of XPC, suggesting that proteolysis occurs at DNA damage sites. Unexpectedly, sustained inhibition of proteasome activity results in aggregation of PSMD14 (presumably with other proteasome components) at the periphery of nucleoli, by which DDB2 is immobilized and sequestered from its lesion recognition functions. Although depletion of PSMD14 alleviates such DDB2 immobilization induced by proteasome inhibitors, recruitment of DDB2 to DNA damage sites is then severely compromised in the absence of PSMD14. Because all of these proteasome dysfunctions selectively impair removal of cyclobutane pyrimidine dimers, but not (6-4) photoproducts, our results indicate that the functional integrity of the proteasome is essential for the DDB2-mediated lesion recognition sub-pathway, but not for GG-NER initiated through direct lesion recognition by XPC.Nov. 2020, Scientific Reports, 10(1) (1), 19704 - 19704, English, International magazine[Refereed]Scientific journal
- Ribonucleoside triphosphates are often incorporated into genomic DNA during DNA replication. The accumulation of unrepaired ribonucleotides is associated with genomic instability, which is mediated by DNA topoisomerase 1 (Top1) processing of embedded ribonucleotides. The cleavage initiated by Top1 at the site of a ribonucleotide leads to the formation of a Top1-DNA cleavage complex (Top1cc), occasionally resulting in a DNA double-strand break (DSB). In humans, tyrosyl-DNA phosphodiesterases (TDPs) are essential repair enzymes that resolve the trapped Top1cc followed by downstream repair factors. However, there is limited cellular evidence of the involvement of TDPs in the processing of incorporated ribonucleotides in mammals. We assessed the role of TDPs in mutagenesis induced by a single ribonucleotide embedded into DNA. A supF shuttle vector site-specifically containing a single riboguanosine (rG) was introduced into the human lymphoblastoid TK6 cell line and its TDP1-, TDP2-, and TDP1/TDP2-deficient derivatives. TDP1 and TDP2 insufficiency remarkably decreased the mutant frequency caused by an embedded rG. The ratio of large deletion mutations induced by rG was also substantially lower in TDP1/TDP2-deficient cells than wild-type cells. Furthermore, the disruption of TDPs reduced the length of rG-mediated large deletion mutations. The recovery ratio of the propagated plasmid was also increased in TDP1/TDP2-deficient cells after the transfection of the shuttle vector containing rG. The results suggest that TDPs-mediated ribonucleotide processing cascade leads to unfavorable consequences, whereas in the absence of these repair factors, a more error-free processing pathway might function to suppress the ribonucleotide-induced mutagenesis. Furthermore, base substitution mutations at sites outside the position of rG were detected in the supF gene via a TDPs-independent mechanism. Overall, we provide new insights into the mechanism of mutagenesis induced by an embedded ribonucleotide in mammalian cells, which may lead to the fatal phenotype in the ribonucleotide excision repair deficiency.2020, PloS one, 15(12) (12), e0244790, English, International magazineScientific journal
- Centrin 2 is a small conserved calcium-binding protein that localizes to the centriolar distal lumen in human cells. It is required for efficient primary ciliogenesis and nucleotide excision repair (NER). Centrin 2 forms part of the xeroderma pigmentosum group C protein complex. To explore how centrin 2 contributes to these distinct processes, we mutated the four calcium-binding EF-hand domains of human centrin 2. Centrin 2 in which all four EF-hands had been mutated to ablate calcium binding (4DA mutant) was capable of supporting in vitro NER and was as effective as the wild-type protein in rescuing the UV sensitivity of centrin 2-null cells. However, we found that mutation of any of the EF-hand domains impaired primary ciliogenesis in human TERT-RPE1 cells to the same extent as deletion of centrin 2. Phenotypic analysis of the 4DA mutant revealed defects in centrosome localization, centriole satellite assembly, ciliary assembly and function and in interactions with POC5 and SFI1. These observations indicate that centrin 2 requires calcium-binding capacity for its primary ciliogenesis functions, but not for NER, and suggest that these functions require centrin 2 to be capable of forming complexes with partner proteins.This article has an associated First Person interview with the first author of the paper.Oct. 2019, Journal of cell science, 132(19) (19), English, International magazine[Refereed]
- DNA polymerases often incorporate non-canonical nucleotide, i.e., ribonucleoside triphosphates into the genomic DNA. Aberrant accumulation of ribonucleotides in the genome causes various cellular abnormalities. Here, we show the possible role of human nucleotide excision repair (NER) and DNA polymerase η (Pol η) in processing of a single ribonucleotide embedded into DNA. We found that the reconstituted NER system can excise the oxidized ribonucleotide on the plasmid DNA. Taken together with the evidence that Pol η accurately bypasses a ribonucleotide, i.e., riboguanosine (rG) or its oxidized derivative (8-oxo-rG) in vitro, we further assessed the mutagenic potential of the embedded ribonucleotide in human cells lacking NER or Pol η. A single rG on the supF reporter gene predominantly induced large deletion mutations. An embedded 8-oxo-rG caused base substitution mutations at the 3'-neighboring base rather than large deletions in wild-type cells. The disruption of XPA, an essential factor for NER, or Pol η leads to the increased mutant frequency of 8-oxo-rG. Furthermore, the frequency of 8-oxo-rG-mediated large deletions was increased by the loss of Pol η, but not XPA. Collectively, our results suggest that base oxidation of the embedded ribonucleotide enables processing of the ribonucleotide via alternative DNA repair and damage tolerance pathways.Sep. 2019, Scientific reports, 9(1) (1), 13910 - 13910, English, International magazine[Refereed]Scientific journal
- Jul. 2019, Nature, 571(7764) (7764), E6, EnglishPublisher Correction: DNA damage detection in nucleosomes involves DNA register shifting.[Refereed]Scientific journal
- Jul. 2019, Nature, 571(7763) (7763), 79 - 84, EnglishDNA damage detection in nucleosomes involves DNA register shifting.[Refereed]Scientific journal
- Nucleotide excision repair (NER) is a versatile DNA repair pathway that eliminates various helix-distorting base lesions such as ultraviolet (UV)-induced photolesions. Several recessive human disorders, such as xeroderma pigmentosum (XP), are caused by hereditary defects in NER, implying that the pathway plays critical roles in suppressing diverse pathogenic processes, including carcinogenesis. In general, discrimination of lesion sites from intact DNA, which is present in vast excess, is a key determinant of the overall efficiency of DNA repair. In mammalian cells, global genomic NER lesion recognition is initiated by the XPC protein complex, which achieves broad DNA-binding specificity by sensing destabilized base pairs rather than lesions per se. To avert unnecessary incisions at lesion-free sites, and thereby ensure the fidelity of the repair system, transcription factor IIH and the XPA protein then verify the presence of relevant lesions at suspicious sites bound by XPC. In the case of UV-induced photolesions, a specialized lesion sensor called UV-damaged DNA-binding protein (UV-DDB) contributes to efficient lesion recognition and the recruitment of XPC to lesion sites. The ubiquitin-proteasome system plays a crucial role in the handoff of lesions from UV-DDB to XPC and the subsequent NER process. In addition, recognition of lesions targeted by global genomic NER is intricately regulated by higher-order chromatin structures, which play distinct roles depending on the type of lesion.2019, The Enzymes, 45, 99 - 138, English, International magazine[Refereed]
- Nucleotide excision repair (NER) is a versatile DNA repair pathway, which can remove an extremely broad range of base lesions from the genome. In mammalian global genomic NER, the XPC protein complex initiates the repair reaction by recognizing sites of DNA damage, and this depends on detection of disrupted/destabilized base pairs within the DNA duplex. A model has been proposed that XPC first interacts with unpaired bases and then the XPD ATPase/helicase in concert with XPA verifies the presence of a relevant lesion by scanning a DNA strand in 5'-3' direction. Such multi-step strategy for damage recognition would contribute to achieve both versatility and accuracy of the NER system at substantially high levels. In addition, recognition of ultraviolet light (UV)-induced DNA photolesions is facilitated by the UV-damaged DNA-binding protein complex (UV-DDB), which not only promotes recruitment of XPC to the damage sites, but also may contribute to remodeling of chromatin structures such that the DNA lesions gain access to XPC and the following repair proteins. Even in the absence of UV-DDB, however, certain types of histone modifications and/or chromatin remodeling could occur, which eventually enable XPC to find sites with DNA lesions. Exploration of novel factors involved in regulation of the DNA damage recognition process is now ongoing.BMC, 2019, Genes and environment : the official journal of the Japanese Environmental Mutagen Society, 41, 2 - 2, English, International magazine[Refereed]Scientific journal
- Fanconi anemia (FA) is a rare genetic disease characterized by the deficiency of the cellular response and repair pathway for DNA interstrand crosslink (ICL) damage. Although recent studies have revealed the detailed molecular functions of FA proteins encoded by 22 genes, the mechanism of occurrence of endogenous ICLs in the human body remains poorly understood. In this short review, we summarize the potential endogenous sources of ICLs counteracted by FA proteins, and provide perspectives on the unanswered questions regarding FA.2019, Genes and environment : the official journal of the Japanese Environmental Mutagen Society, 41, 6 - 6, English, International magazine[Refereed]Scientific journal
- Blackwell Publishing Ltd, Jun. 2018, British Journal of Dermatology, 178(6) (6), 1451 - 1452, EnglishScientific journal
- The p300 and CBP histone acetyltransferases are recruited to DNA double-strand break (DSB) sites where they induce histone acetylation, thereby influencing the chromatin structure and DNA repair process. Whether p300/CBP at DSB sites also acetylate non-histone proteins, and how their acetylation affects DSB repair, remain unknown. Here we show that p300/CBP acetylate RAD52, a human homologous recombination (HR) DNA repair protein, at DSB sites. Using in vitro acetylated RAD52, we identified 13 potential acetylation sites in RAD52 by a mass spectrometry analysis. An immunofluorescence microscopy analysis revealed that RAD52 acetylation at DSBs sites is counteracted by SIRT2- and SIRT3-mediated deacetylation, and that non-acetylated RAD52 initially accumulates at DSB sites, but dissociates prematurely from them. In the absence of RAD52 acetylation, RAD51, which plays a central role in HR, also dissociates prematurely from DSB sites, and hence HR is impaired. Furthermore, inhibition of ataxia telangiectasia mutated (ATM) protein by siRNA or inhibitor treatment demonstrated that the acetylation of RAD52 at DSB sites is dependent on the ATM protein kinase activity, through the formation of RAD52, p300/CBP, SIRT2, and SIRT3 foci at DSB sites. Our findings clarify the importance of RAD52 acetylation in HR and its underlying mechanism.Mar. 2018, PLoS genetics, 14(3) (3), e1007277, English, International magazine[Refereed]Scientific journal
- Feb. 2018, Genes Cells, 23(3) (3), 200 - 213, EnglishMutations at multiple CDK phosphorylation consensus sites on Cdt2 increase the affinity of CRL4Cdt2 for PCNA and its ubiquitination activity in S phase.[Refereed]Scientific journal
- Thymine DNA glycosylase (TDG) is a base excision repair (BER) enzyme, which is implicated in correction of deamination-induced DNA mismatches, the DNA demethylation process and regulation of gene expression. Because of these pivotal roles associated, it is crucial to elucidate how the TDG functions are appropriately regulated invivo. Here, we present evidence that the TDG protein undergoes degradation upon various types of DNA damage, including ultraviolet light (UV). The UV-induced degradation of TDG was dependent on proficiency in nucleotide excision repair and on CRL4(CDT2)-mediated ubiquitination that requires a physical interaction between TDG and DNA polymerase clamp PCNA. Using the Tdg-deficient mouse embryonic fibroblasts, we found that ectopic expression of TDG compromised cellular survival after UV irradiation and repair of UV-induced DNA lesions. These negative effects on cellular UV responses were alleviated by introducing mutations in TDG that impaired its BER function. The expression of TDG induced a large-scale alteration in the gene expression profile independently of its DNA glycosylase activity, whereas a subset of genes was affected by the catalytic activity of TDG. Our results indicate the presence of BER-dependent and BER-independent functions of TDG, which are involved in regulation of cellular DNA damage responses and gene expression patterns.WILEY, Apr. 2017, GENES TO CELLS, 22(4) (4), 392 - 405, English[Refereed]Scientific journal
- Ultraviolet (UV) radiation is a carcinogen that generates DNA lesions. Here, we demonstrate an unexpected role for DGCR8, an RNA binding protein that canonically functions with Drosha to mediate microRNA processing, in the repair of UV-induced DNA lesions. Treatment with UV induced phosphorylation on serine 153 (S153) of DGCR8 in both human and murine cells. S153 phosphorylation was critical for cellular resistance to UV, the removal of UV-induced DNA lesions, and the recovery of RNA synthesis after UV exposure but not for microRNA expression. The RNA-binding and Drosha-binding activities of DGCR8 were not critical for UV resistance. DGCR8 depletion was epistatic to defects in XPA, CSA, and CSB for UV sensitivity. DGCR8 physically interacted with CSB and RNA polymerase II. JNKs were involved in the UV-induced S153 phosphorylation. These findings suggest that UV-induced S153 phosphorylation mediates transcription-coupled nucleotide excision repair of UV-induced DNA lesions in a manner independent of microRNA processing.CELL PRESS, Apr. 2017, CELL REPORTS, 19(1) (1), 162 - 174, English[Refereed]Scientific journal
- In the mammalian global genome nucleotide excision repair pathway, two damage recognition factors, XPC and UV-DDB, play pivotal roles in the initiation of the repair reaction. However, the molecular mechanisms underlying regulation of the lesion recognition process in the context of chromatin structures remain to be understood. Here, we show evidence that damage recognition factors tend to associate with chromatin regions devoid of certain types of acetylated histones. Treatment of cells with histone deacetylase inhibitors retarded recruitment of XPC to sites of UV-induced DNA damage and the subsequent repair process. Biochemical studies showed novel multifaceted interactions of XPC with histone H3, which were profoundly impaired by deletion of the N-terminal tail of histone H3. In addition, histone H1 also interacted with XPC. Importantly, acetylation of histone H3 markedly attenuated the interaction with XPC invitro, and local UV irradiation of cells decreased the level of H3K27ac in the damaged areas. Our results suggest that histone deacetylation plays a significant role in the process of DNA damage recognition for nucleotide excision repair and that the localization and functions of XPC can be regulated by acetylated states of histones.WILEY, Mar. 2017, GENES TO CELLS, 22(3) (3), 310 - 327, English[Refereed]Scientific journal
- Cdt1 is rapidly degraded by CRL4(Cdt2) E3 ubiquitin ligase after UV (UV) irradiation. Previous reports revealed that the nucleotide excision repair (NER) pathway is responsible for the rapid Cdt1-proteolysis. Here, we show that mismatch repair (MMR) proteins are also involved in the degradation of Cdt1 after UV irradiation in the G1 phase. First, compared with the rapid (within approximate to 15min) degradation of Cdt1 in normal fibroblasts, Cdt1 remained stable for approximate to 30min in NER-deficient XP-A cells, but was degraded within approximate to 60min. The delayed degradation was also dependent on PCNA and CRL4(Cdt2). The MMR proteins Msh2 and Msh6 were recruited to the UV-damaged sites of XP-A cells in the G1 phase. Depletion of these factors with small interfering RNAs prevented Cdt1 degradation in XP-A cells. Similar to the findings in XP-A cells, depletion of XPA delayed Cdt1 degradation in normal fibroblasts and U2OS cells, and co-depletion of Msh6 further prevented Cdt1 degradation. Furthermore, depletion of Msh6 alone delayed Cdt1 degradation in both cell types. When Cdt1 degradation was attenuated by high Cdt1 expression, repair synthesis at the damaged sites was inhibited. Our findings demonstrate that UV irradiation induces multiple repair pathways that activate CRL4(Cdt2) to degrade its target proteins in the G1 phase of the cell cycle, leading to efficient repair of DNA damage.TAYLOR & FRANCIS INC, 2017, CELL CYCLE, 16(7) (7), 673 - 684, English[Refereed]Scientific journal
- Huge amounts (>10,000/day) of apurinic/apyrimidinic (AP) sites are produced in genomes, but their structures in chromatin remain undetermined. We determined the crystal structure of the nucleosome containing AP-site analogs at two symmetric sites, which revealed structural polymorphism: one forms an inchworm configuration without an empty space at the AP site, and the other forms a B-form-like structure with an empty space and the orphan base. This unexpected inchworm configuration of the AP site is important to understand the AP DNA repair mechanism, because it may not be recognized by the major AP-binding protein, APE1, during the base excision repair process.NATURE PUBLISHING GROUP, Jan. 2017, SCIENTIFIC REPORTS, 7(7) (7), 41783, English[Refereed]Scientific journal
- BACKGROUND: Most patients with xeroderma pigmentosum complementation group D (XP-D) from Western countries suffer from neurological symptoms, whereas Japanese patients display only skin manifestations without neurological symptoms. We have previously suggested that these differences in clinical manifestations in XP-D patients are attributed partly to a predominant mutation in ERCC2, and the allele frequency of S541R is highest in Japan. METHODS: We diagnosed a child with mild case of XP-D by the evaluation of DNA repair activity and whole-genome sequencing, and followed her ten years. RESULTS: Skin cancer, mental retardation, and neurological symptoms were not observed. Her minimal erythema dose was 41 mJ/cm(2) , which was slightly lower than that of healthy Japanese volunteers. The patient's cells showed sixfold hypersensitivity to UV in comparison with normal cells. Post-UV unscheduled DNA synthesis was 20.4%, and post-UV recovery of RNA synthesis was 58% of non-irradiated samples, which was lower than that of normal fibroblasts. Genome sequence analysis indicated that the patient harbored a compound heterozygous mutation of c.1621A>C and c.591_594del, resulting in p.S541R and p.Y197* in ERCC2: then, patient was diagnosed with XP-D. Y197* has not been described before. CONCLUSION: Her mild skin manifestations might be attributed to the mutational site on her genome and daily strict sun protection. c.1621A>C might be a founder mutation of ERCC2 among Japanese XP-D patients, as it was identified most frequently in Japanese XP-D patients and it has not been found elsewhere outside Japan.Jul. 2016, Photodermatology, photoimmunology & photomedicine, 32(4) (4), 174 - 80, English, International magazine[Refereed][Invited]Scientific journal
- The cullin-RING ubiquitin E3 ligase (CRL) family comprises over 200 members in humans. The COP9 signalosome complex (CSN) regulates CRLs by removing their ubiquitin-like activator NEDD8. The CUL4A-RBX1-DDB1-DDB2 complex (CRL4A(DDB2)) monitors the genome for ultraviolet-light-induced DNA damage. CRL4A(DBB2) is inactive in the absence of damaged DNA and requires CSN to regulate the repair process. The structural basis of CSN binding to CRL4A(DDB2) and the principles of CSN activation are poorly understood. Here we present cryo-electron microscopy structures for CSN in complex with neddylated CRL4A ligases to 6.4 angstrom resolution. The CSN conformers defined by cryo-electron microscopy and a novel apo-CSN crystal structure indicate an induced-fit mechanism that drives CSN activation by neddylated CRLs. We find that CSN and a substrate cannot bind simultaneously to CRL4A, favouring a deneddylated, inactive state for substrate-free CRL4 complexes. These architectural and regulatory principles appear conserved across CRL families, allowing global regulation by CSN.NATURE PUBLISHING GROUP, Mar. 2016, NATURE, 531(7596) (7596), 598 - +, English[Refereed]Scientific journal
- The cullin-RING ubiquitin E3 ligase (CRL) family comprises over 200 members in humans. The COP9 signalosome complex (CSN) regulates CRLs by removing their ubiquitin-like activator NEDD8. The CUL4A-RBX1-DDB1-DDB2 complex (CRL4A(DDB2)) monitors the genome for ultraviolet-light-induced DNA damage. CRL4A(DBB2) is inactive in the absence of damaged DNA and requires CSN to regulate the repair process. The structural basis of CSN binding to CRL4A(DDB2) and the principles of CSN activation are poorly understood. Here we present cryo-electron microscopy structures for CSN in complex with neddylated CRL4A ligases to 6.4 angstrom resolution. The CSN conformers defined by cryo-electron microscopy and a novel apo-CSN crystal structure indicate an induced-fit mechanism that drives CSN activation by neddylated CRLs. We find that CSN and a substrate cannot bind simultaneously to CRL4A, favouring a deneddylated, inactive state for substrate-free CRL4 complexes. These architectural and regulatory principles appear conserved across CRL families, allowing global regulation by CSN.NATURE PUBLISHING GROUP, Mar. 2016, NATURE, 531(7596) (7596), 598 - +, EnglishScientific journal
- The cyclobutane pyrimidine dimer (CPD) is induced in genomic DNA by ultraviolet (UV) light. In mammals, this photolesion is primarily induced within nucleosomal DNA, and repaired exclusively by the nucleotide excision repair (NER) pathway. However, the mechanism by which the CPD is accommodated within the nucleosome has remained unknown. We now report the crystal structure of a nucleosome containing CPDs. In the nucleosome, the CPD induces only limited local backbone distortion, and the affected bases are accommodated within the duplex. Interestingly, one of the affected thymine bases is located within 3.0 angstrom from the undamaged complementary adenine base, suggesting the formation of complementary hydrogen bonds in the nucleosome. We also found that UV-DDB, which binds the CPD at the initial stage of the NER pathway, also efficiently binds to the nucleosomal CPD. These results provide important structural and biochemical information for understanding how the CPD is accommodated and recognized in chromatin. (C) 2016 Elsevier Inc. All rights reserved.ACADEMIC PRESS INC ELSEVIER SCIENCE, Feb. 2016, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 471(1) (1), 117 - 122, English[Refereed]Scientific journal
- Feb. 2016, Journal of pharmaceutical negative results, 7, 42 - 45, English2,2,4-Triamino-5(2H)-oxazolone is a weak substrate for nucleotide excision repair[Refereed]Scientific journal
- Objective: 2,2,4-Triamino-5(2H)-oxazolone (Oz) is a guanine lesion produced by reactive oxygen radicals and photosensitized oxidation. This nucleobase is a potentially mutagenic lesion, and is removed by several base excision repair enzymes. Our purpose is to analyze whether Oz is the substrate of nucleotide excision repair (NER). Materials and Methods: A lymphoblastoid cell line from the patient with xeroderma pigmentosum (XP) complementation group C (XP3BE) was used. Cell-free NER reactions with covalently closed circular DNAs containing the Oz lesion were performed using the XP3BE whole cell extracts with or without the XPC-RAD23B complex. In addition, DNA fragments (180 bp in length) containing the Oz lesion were used for binding reactions with the XPC-RAD23B complex. Results: We analyzed the cell-free NER activity on Oz and the binding affinity of XPC-RAD23B, which initiates NER. Human cell-free NER activity on Oz was detected, though the reactivity to Oz was lower than that on ultra violet (UV)-induced pyrimidine (6-4) pyrimidone photoproduct (6-4PP). Also, binding of XPC-RAD23B with Oz was lower than that with 6-4PP. Conclusion: Because of the low binding affinity of Oz for XPC-RAD23B, NER efficiency on Oz is very low. Therefore, general NER is not the appropriate repair system for Oz.Medknow Publications, Jan. 2016, Journal of Pharmaceutical Negative Results, 7(1) (1), 42 - 45, English[Refereed]Scientific journal
- UV-DDB, an initiation factor for the nucleotide excision repair pathway, recognizes 6-4PP lesions through a base flipping mechanism. As genomic DNA is almost entirely accommodated within nucleosomes, the flipping of the 6-4PP bases is supposed to be extremely difficult if the lesion occurs in a nucleosome, especially on the strand directly contacting the histone surface. Here we report that UV-DDB binds efficiently to nucleosomal 6-4PPs that are rotationally positioned on the solvent accessible or occluded surface. We determined the crystal structures of nucleosomes containing 6-4PPs in these rotational positions, and found that the 6-4PP DNA regions were flexibly disordered, especially in the strand exposed to the solvent. This characteristic of 6-4PP may facilitate UV-DDB binding to the damaged nucleosome. We present the first atomic-resolution pictures of the detrimental DNA cross-links of neighboring pyrimidine bases within the nucleosome, and provide the mechanistic framework for lesion recognition by UV-DDB in chromatin.Nov. 2015, Scientific reports, 5, 16330 - 16330, English, International magazine[Refereed]Scientific journal
- In global genome repair (GGR), XPC detects damaged nucleotides and recruits TFIIH complex. The small acidic region of XPC binds to the pleckstrin homology (PH) domain of TFIIH subunit p62; however, the recognition mechanism remains elusive. Here, we use nuclear magnetic resonance to present the tertiary structure of XPC bound to the PH domain. The XPC acidic region forms a long string stabilized by insertion of Trp133 and Val136 into two separate hollows of the PH domain, coupled with extensive electrostatic contacts. Analysis of several XPC mutants revealed that particularly Trp133 is essential for binding to the PH domain. In cell lines stably expressing mutant XPC, alanine substitution at Trp133 or Trp133/Val136 compromised UV resistance, recruitment of TFIIH to DNA damage, and removal of UV-induced photoproducts from genomic DNA. These findings show how TFIIH complex is recruited by XPC to damaged DNA, advancing our understanding of the early stage of GGR.CELL PRESS, Oct. 2015, STRUCTURE, 23(10) (10), 1827 - 1837, English[Refereed]Scientific journal
- Transcription factor IIH (TFIIH) is essential for both transcription and nucleotide excision repair (NER). DNA lesions are initially detected by NER factors XPC and XPE or stalled RNA polymerases, but only bulky lesions are preferentially repaired by NER. To elucidate substrate specificity in NER, we have prepared homogeneous human ten-subunit TFIIH and its seven-subunit core (Core7) without the CAK module and show that bulky lesions in DNA inhibit the ATPase and helicase activities of both XPB and XPD in Core7 to promote NER, whereas non-genuine NER substrates have no such effect. Moreover, the NER factor XPA activates unwinding of normal DNA by Core7, but inhibits the Core7 helicase activity in the presence of bulky lesions. Finally, the CAK module inhibits DNA binding by TFIIH and thereby enhances XPC-dependent specific recruitment of TFIIH. Our results support a tripartite lesion verification mechanism involving XPC, TFIIH, and XPA for efficient NER.CELL PRESS, Sep. 2015, MOLECULAR CELL, 59(6) (6), 1025 - 1034, English[Refereed]Scientific journal
- The xeroderma pigmentosum group C (XPC) protein complex is a key factor that detects DNA damage and initiates nucleotide excision repair (NER) in mammalian cells. Although biochemical and structural studies have elucidated the interaction of XPC with damaged DNA, the mechanism of its regulation in vivo remains to be understood in more details. Here, we show that the XPC protein undergoes modification by small ubiquitin-related modifier (SUMO) proteins and the lack of this modification compromises the repair of UV-induced DNA photolesions. In the absence of SUMOylation, XPC is normally recruited to the sites with photolesions, but then immobilized profoundly by the UV-damaged DNA-binding protein (UV-DDB) complex. Since the absence of UV-DDB alleviates the NER defect caused by impaired SUMOylation of XPC, we propose that this modification is critical for functional interactions of XPC with UV-DDB, which facilitate the efficient damage handover between the two damage recognition factors and subsequent initiation of NER.Jun. 2015, Scientific reports, 5(5) (5), 10984 - 10984, English, International magazine[Refereed]Scientific journal
- The xeroderma pigmentosum group C (XPC) protein complex is a key factor that detects DNA damage and initiates nucleotide excision repair (NER) in mammalian cells. Although biochemical and structural studies have elucidated the interaction of XPC with damaged DNA, the mechanism of its regulation in vivo remains to be understood in more details. Here, we show that the XPC protein undergoes modification by small ubiquitin-related modifier (SUMO) proteins and the lack of this modification compromises the repair of UV-induced DNA photolesions. In the absence of SUMOylation, XPC is normally recruited to the sites with photolesions, but then immobilized profoundly by the UV-damaged DNA-binding protein (UV-DDB) complex. Since the absence of UV-DDB alleviates the NER defect caused by impaired SUMOylation of XPC, we propose that this modification is critical for functional interactions of XPC with UV-DDB, which facilitate the efficient damage handover between the two damage recognition factors and subsequent initiation of NER.Jun. 2015, Scientific reports, 5, 10984 - 10984, English, International magazine[Refereed]Scientific journal
- Jun. 2015, Structure, 5, 10984, English[Refereed]Scientific journal
- ELSEVIER SCIENCE BV, Apr. 2015, DNA REPAIR, 28, 139 - 140, EnglishStructures of the nucleosomes containing UV-damaged DNA bases and the damaged base recognition mechanism by DNA repair proteins
- In eukaryotes, holo-Mediator consists of four modules: head, middle, tail, and CDK/Cyclin. The head module performs an essential function involved in regulation of RNA polymerase II (Pol II). We studied the human head module subunit MED17 (hMED17). Recent structural studies showed that yeast MED17 may function as a hinge connecting the neck and movable jaw regions of the head module to the fixed jaw region. Luciferase assays in hMED17-knockdown cells showed that hMED17 supports transcriptional activation, and pulldown assays showed that hMED17 interacted with Pol II and the general transcription factors TFIIB, TBP, TFIIE, and TFIIH. In addition, hMED17 bound to a DNA helicase subunit of TFIIH, XPB, which is essential for both transcription and nucleotide excision repair (NER). Because hMED17 associates with p53 upon UV-C irradiation, we treated human MCF-7 cells with either UV-C or the MDM2 inhibitor Nutlin-3. Both treatments resulted in accumulation of p53 in the nucleus, but hMED17 remained concentrated in the nucleus in response to UV-C. hMED17 colocalized with the NER factors XPB and XPG following UV-C irradiation, and XPG and XPB bound to hMED17 in vitro. These findings suggest that hMED17 may play essential roles in switching between transcription and NER.WILEY-BLACKWELL, Mar. 2015, GENES TO CELLS, 20(3) (3), 191 - 202, English[Refereed]Scientific journal
- In mammalian nucleotide excision repair, the DDB1-DDB2 complex recognizes UV-induced DNA photolesions and facilitates recruitment of the XPC complex. Upon binding to damaged DNA, the Cullin 4 ubiquitin ligase associated with DDB1-DDB2 is activated and ubiquitinates DDB2 and XPC. The structurally disordered N-terminal tail of DDB2 contains seven lysines identified as major sites for ubiquitination that target the protein for proteasomal degradation; however, the precise biological functions of these modifications remained unknown. By exogenous expression of mutant DDB2 proteins in normal human fibroblasts, here we show that the N-terminal tail of DDB2 is involved in regulation of cellular responses to UV. By striking contrast with behaviors of exogenous DDB2, the endogenous DDB2 protein was stabilized even after UV irradiation as a function of the XPC expression level. Furthermore, XPC competitively suppressed ubiquitination of DDB2 in vitro, and this effect was significantly promoted by centrin-2, which augments the DNA damage-recognition activity of XPC. Based on these findings, we propose that in cells exposed to UV, DDB2 is protected by XPC from ubiquitination and degradation in a stochastic manner; thus XPC allows DDB2 to initiate multiple rounds of repair events, thereby contributing to the persistence of cellular DNA repair capacity.Feb. 2015, Nucleic acids research, 43(3) (3), 1700 - 13, English, International magazine[Refereed]Scientific journal
- For organisms, a fundamental problem is how the huge genome can be searched appropriately for DNA damage in order to prevent genome instability and/or cell death. In mammalian nucleotide excision repair (NER), this process involves multiple protein factors containing the xeroderma pigmentosum gene products (UV-DDB, XPC, TFIIH), which sense different aspects of DNA structural abnormalities, respectively. Coordinated stepwise actions of these factors contribute to simultaneous achievement of versatility, efficiency and accuracy of the DNA quality control for NER.The Biophysical Society of Japan General Incorporated Association, 2015, Seibutsu Butsuri, 55(3) (3), 137 - 141, Japanese
- The Fanconi anemia (FA) pathway, of which the FANCD2 protein is a key component, plays crucial roles in the maintenance of hematopoietic stem cells and suppression of carcinogenesis. However, the function of FANCD2 remains unclear. Here, we report that FANCD2 is a novel and specific substrate of caspase 3. Cleavage of FANCD2 by caspase 3 did not require either the FA core complex or mono-ubiquitylation of FANCD2, and was stimulated by p53. In addition, we identified the cleavage sites and generated cell lines that stably express a caspase-resistant FANCD2 mutant. Our data suggest that FANCD2 is regulated by caspase-mediated degradation during apoptosis induced by DNA damage.Elsevier, Oct. 2014, FEBS Letters, 588(20) (20), 3778 - 3785, English[Refereed]Scientific journal
- The Fanconi anemia (FA) pathway, of which the FANCD2 protein is a key component, plays crucial roles in the maintenance of hematopoietic stem cells and suppression of carcinogenesis. However, the function of FANCD2 remains unclear. Here, we report that FANCD2 is a novel and specific substrate of caspase 3. Cleavage of FANCD2 by caspase 3 did not require either the FA core complex or mono-ubiquitylation of FANCD2, and was stimulated by p53. In addition, we identified the cleavage sites and generated cell lines that stably express a caspase-resistant FANCD2 mutant. Our data suggest that FANCD2 is regulated by caspase-mediated degradation during apoptosis induced by DNA damage. (C) 2014 The Authors. Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies.ELSEVIER SCIENCE BV, Oct. 2014, FEBS LETTERS, 588(20) (20), 3778 - 3785, English[Refereed]Scientific journal
- To maintain genetic integrity, ultraviolet light-induced photoproducts in DNA must be removed by the nucleotide excision repair (NER) pathway, which is initiated by damage recognition and dual incisions of the lesion-containing strand. We intended to detect the dual-incision step of cellular NER, by using a fluorescent probe. A 140-base pair linear duplex containing the (6-4) photoproduct and a fluorophore-quencher pair was prepared first. However, this type of DNA was found to be degraded rapidly by nucleases in cells. Next, a plasmid was used as a scaffold. In this case, the fluorophore and the quencher were attached to the same strand, and we expected that the dual-incision product containing them would be degraded in cells. At 3 h after transfection of HeLa cells with the plasmid-type probes, fluorescence emission was detected at the nuclei by fluorescence microscopy only when the probe contained the (6-4) photoproduct, and the results were confirmed by flow cytometry. Finally, XPA fibroblasts and the same cells expressing the XPA gene were transfected with the photoproduct-containing probe. Although the transfer of the probe into the cells was slow, fluorescence was detected depending on the NER ability of the cells.NATURE PUBLISHING GROUP, Jul. 2014, SCIENTIFIC REPORTS, 4, 5578, English[Refereed]Scientific journal
- To maintain genetic integrity, ultraviolet light-induced photoproducts in DNA must be removed by the nucleotide excision repair (NER) pathway, which is initiated by damage recognition and dual incisions of the lesion-containing strand. We intended to detect the dual-incision step of cellular NER, by using a fluorescent probe. A 140-base pair linear duplex containing the (6-4) photoproduct and a fluorophore-quencher pair was prepared first. However, this type of DNA was found to be degraded rapidly by nucleases in cells. Next, a plasmid was used as a scaffold. In this case, the fluorophore and the quencher were attached to the same strand, and we expected that the dual-incision product containing them would be degraded in cells. At 3 h after transfection of HeLa cells with the plasmid-type probes, fluorescence emission was detected at the nuclei by fluorescence microscopy only when the probe contained the (6-4) photoproduct, and the results were confirmed by flow cytometry. Finally, XPA fibroblasts and the same cells expressing the XPA gene were transfected with the photoproduct-containing probe. Although the transfer of the probe into the cells was slow, fluorescence was detected depending on the NER ability of the cells.NATURE PUBLISHING GROUP, Jul. 2014, SCIENTIFIC REPORTS, 4, 5578, English[Refereed]Scientific journal
- Thymine DNA glycosylase (TDG) is a base excision repair enzyme that interacts with the small ubiquitin-related modifier (SUMO)-targeted ubiquitin E3 ligase RNF4 and functions in the active DNA demethylation pathway. Here we showed that both SUMOylated and non-modified forms of endogenous TDG fluctuated during the cell cycle and in response to drugs that perturbed cell cycle progression, including hydroxyurea and nocodazole. Additionally, we detected a SUMOylation-independent association between TDG and RNF4 in vitro as well as in vivo, and observed that both forms of TDG were efficiently degraded in RNF4-depleted cells when arrested at S phase. Our findings provide insights into the in vivo dynamics of TDG SUMOylation and further clarify the TDG-RNF4 interaction. (C) 2014 Elsevier Inc. All rights reserved.ACADEMIC PRESS INC ELSEVIER SCIENCE, May 2014, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 447(3) (3), 419 - 424, English[Refereed]Scientific journal
- The Fanconi anemia (FA) pathway is critically involved in the maintenance of hematopoietic stem cells and the suppression of carcinogenesis. A key FA protein, FANCD2, is monoubiquitinated and accumulates in chromatin in response to DNA interstrand crosslinks (ICLs), where it coordinates DNA repair through mechanisms that are still poorly understood. Here, we report that CtIP protein directly interacts with FANCD2. A region spanning amino acids 166 to 273 of CtIP and monoubiquitination of FANCD2 are both essential for the FANCD2-CtIP interaction and mitomycin C (MMC)-induced CtIP foci. Remarkably, both FANCD2 and CtIP are critical for MMC-induced RPA2 hyperphosphorylation, an event that accompanies end resection of double-strand breaks. Collectively, our results reveal a role of monoubiquitinated FANCD2 in end resection that depends on its binding to CtIP during ICL repair.CELL PRESS, May 2014, CELL REPORTS, 7(4) (4), 1039 - 1047, English[Refereed]Scientific journal
- Thymine DNA glycosylase (TDG) is a base excision repair enzyme that interacts with the small ubiquitin-related modifier (SUMO)-targeted ubiquitin E3 ligase RNF4 and functions in the active DNA demethylation pathway. Here we showed that both SUMOylated and non-modified forms of endogenous TDG fluctuated during the cell cycle and in response to drugs that perturbed cell cycle progression, including hydroxyurea and nocodazole. Additionally, we detected a SUMOylation-independent association between TDG and RNF4 in vitro as well as in vivo, and observed that both forms of TDG were efficiently degraded in RNF4-depleted cells when arrested at S phase. Our findings provide insights into the in vivo dynamics of TDG SUMOylation and further clarify the TDG-RNF4 interaction. (C) 2014 Elsevier Inc. All rights reserved.ACADEMIC PRESS INC ELSEVIER SCIENCE, May 2014, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 447(3) (3), 419 - 424, English[Refereed]Scientific journal
- The Fanconi anemia (FA) pathway is critically involved in the maintenance of hematopoietic stem cells and the suppression of carcinogenesis. A key FA protein, FANCD2, is monoubiquitinated and accumulates in chromatin in response to DNA interstrand crosslinks (ICLs), where it coordinates DNA repair through mechanisms that are still poorly understood. Here, we report that CtIP protein directly interacts with FANCD2. A region spanning amino acids 166 to 273 of CtIP and monoubiquitination of FANCD2 are both essential for the FANCD2-CtIP interaction and mitomycin C (MMC)-induced CtIP foci. Remarkably, both FANCD2 and CtIP are critical for MMC-induced RPA2 hyperphosphorylation, an event that accompanies end resection of double-strand breaks. Collectively, our results reveal a role of monoubiquitinated FANCD2 in end resection that depends on its binding to CtIP during ICL repair.CELL PRESS, May 2014, CELL REPORTS, 7(4) (4), 1039 - 1047, English[Refereed]Scientific journal
- The Biophysical Society of Japan General Incorporated Association, 2014, Seibutsu Butsuri, 54(1) (1), S214, English
- Ten-eleven translocation (TET) proteins oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). 5fC and 5caC can be excised and repaired by the base excision repair (BER) pathway, implicating 5mC oxidation in active DNA demethylation. Genome-wide DNA methylation is erased in the transition from metastable states to the ground state of embryonic stem cells (ESCs) and in migrating primordial germ cells (PGCs), although some resistant regions become demethylated only in gonadal PGCs. Understanding the mechanisms underlying global hypomethylation in naive ESCs and developing PGCs will be useful for realizing cellular pluripotency and totipotency. In this study, we found that PRDM14, the PR domain-containing transcriptional regulator, accelerates the TET-BER cycle, resulting in the promotion of active DNA demethylation in ESCs. Induction of Prdm14 expression transiently elevated 5hmC, followed by the reduction of 5mC at pluripotency-associated genes, germline-specific genes and imprinted loci, but not across the entire genome, which resembles the second wave of DNA demethylation observed in gonadal PGCs. PRDM14 physically interacts with TET1 and TET2 and enhances the recruitment of TET1 and TET2 at target loci. Knockdown of TET1 and TET2 impaired transcriptional regulation and DNA demethylation by PRDM14. The repression of the BER pathway by administration of pharmacological inhibitors of APE1 and PARP1 and the knockdown of thymine DNA glycosylase (TDG) also impaired DNA demethylation by PRDM14. Furthermore, DNA demethylation induced by PRDM14 takes place normally in the presence of aphidicolin, which is an inhibitor of G1/S progression. Together, our analysis provides mechanistic insight into DNA demethylation in naive pluripotent stem cells and developing PGCs.COMPANY OF BIOLOGISTS LTD, Jan. 2014, DEVELOPMENT, 141(2) (2), 269 - 280, English[Refereed]Scientific journal
- 2014, Genes Cells., 20(3) (3), 191 - 202, EnglishHuman mediator MED17 subunit plays essential roles in gene regulation by associating with the transcription and DNA repair machineries.[Refereed]Scientific journal
- Ten-eleven translocation (TET) proteins oxidize 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). 5fC and 5caC can be excised and repaired by the base excision repair (BER) pathway, implicating 5mC oxidation in active DNA demethylation. Genome-wide DNA methylation is erased in the transition from metastable states to the ground state of embryonic stem cells (ESCs) and in migrating primordial germ cells (PGCs), although some resistant regions become demethylated only in gonadal PGCs. Understanding the mechanisms underlying global hypomethylation in naive ESCs and developing PGCs will be useful for realizing cellular pluripotency and totipotency. In this study, we found that PRDM14, the PR domain-containing transcriptional regulator, accelerates the TET-BER cycle, resulting in the promotion of active DNA demethylation in ESCs. Induction of Prdm14 expression transiently elevated 5hmC, followed by the reduction of 5mC at pluripotency-associated genes, germline-specific genes and imprinted loci, but not across the entire genome, which resembles the second wave of DNA demethylation observed in gonadal PGCs. PRDM14 physically interacts with TET1 and TET2 and enhances the recruitment of TET1 and TET2 at target loci. Knockdown of TET1 and TET2 impaired transcriptional regulation and DNA demethylation by PRDM14. The repression of the BER pathway by administration of pharmacological inhibitors of APE1 and PARP1 and the knockdown of thymine DNA glycosylase (TDG) also impaired DNA demethylation by PRDM14. Furthermore, DNA demethylation induced by PRDM14 takes place normally in the presence of aphidicolin, which is an inhibitor of G1/S progression. Together, our analysis provides mechanistic insight into DNA demethylation in naive pluripotent stem cells and developing PGCs.COMPANY OF BIOLOGISTS LTD, Jan. 2014, DEVELOPMENT, 141(2) (2), 269 - 280, English[Refereed]Scientific journal
- The human XPC-RAD23B complex and its yeast ortholog, Rad4-Rad23, are the primary initiators of global genome nucleotide excision repair. In this study, two types of DNA binding assays were used for the detailed analysis of interaction of these proteins with damaged DNA. An electrophoretic mobility shift assay revealed that human and yeast orthologs behave similarly in DNA binding. Quantitative analyses of XPC/Rad4 binding to the model DNA structures were performed using fluorescent depolarization measurements. The XPC-RAD23B and the Rad4-Rad23 proteins bind to the damaged 15 nt bubble-DNA structure mimicking in size the "transcription bubble" DNA intermediate with the highest affinity (KD values ~10-10 M or less) that is reduced in the following order: damaged bubble > undamaged bubble > damaged duplex > undamaged duplex. The affinity of XPC/Rad4 for various DNAs was shown to correlate with DNA bending angle. The results obtained show clearly that more deviation from regular DNA structure leads to higher XPC/Rad4 affinity. Copyright © 2013 John Wiley & Sons, Ltd.Dec. 2013, Journal of Molecular Recognition, 26(12) (12), 653 - 661, English[Refereed]Scientific journal
- Centrin-2 is an evolutionarily conserved, calmodulin-related protein, which is involved in multiple cellular functions including centrosome regulation and nucleotide excision repair (NER) of DNA. Particularly to exert the latter function, complex formation with the XPC protein, the pivotal NER damage recognition factor, is crucial. Here, we show that the C-terminal half of centrin-2, containing two calcium-binding EF-hand motifs, is necessary and sufficient for both its localization to the centrosome and interaction with XPC. In XPC-deficient cells, nuclear localization of overexpressed centrin-2 largely depends on co-overexpression of XPC, and mutational analyses of the C-terminal domain suggest that XPC and the major binding partner in the centrosome share a common binding surface on the centrin-2 molecule. On the other hand, the N-terminal domain of centrin-2 also contains two EF-hand motifs but shows only low-binding affinity for calcium ions. Although the N-terminal domain is dispensable for enhancement of the DNA damage recognition activity of XPC, it contributes to augmenting rather weak physical interaction between XPC and XPA, another key factor involved in NER. These results suggest that centrin-2 may have evolved to bridge two protein factors, one with high affinity and the other with low affinity, thereby allowing delicate regulation of various biological processes.Aug. 2013, Nucleic acids research, 41(14) (14), 6917 - 29, English, International magazine[Refereed]Scientific journal
- DNA double-strand breaks (DSB) occur frequently during replication in sister chromatids and are dramatically increased when cells are exposed to chemotherapeutic agents including camptothecin. Such DSBs are efficiently repaired specifically by homologous recombination (HR) with the intact sister chromatid. HR, therefore, plays pivotal roles in cellular proliferation and cellular tolerance to camptothecin. Mammalian cells carry several structure-specific endonucleases, such as Xpf-Ercc1 and Mus81-Eme1, in which Xpf and Mus81 are the essential subunits for enzymatic activity. Here, we show the functional overlap between Xpf and Mus81 by conditionally inactivating Xpf in the chicken DT40 cell line, which has no Mus81 ortholog. Although mammalian cells deficient in either Xpf or Mus81 are viable, Xpf inactivation in DT40 cells was lethal, resulting in a marked increase in the number of spontaneous chromosome breaks. Similarly, inactivation of both Xpf and Mus81 in human HeLa cells and murine embryonic stem cells caused numerous spontaneous chromosome breaks. Furthermore, the phenotype of Xpf-deficient DT40 cells was reversed by ectopic expression of human Mus81-Eme1 or human Xpf-Ercc1 heterodimers. These observations indicate the functional overlap of Xpf-Ercc1 and Mus81-Eme1 in the maintenance of genomic DNA. Both Mus81-Eme1 and Xpf-Ercc1 contribute to the completion of HR, as evidenced by the data that the expression of Mus81-Eme1 or Xpf-Ercc1 diminished the number of camptothecin-induced chromosome breaks in Xpf-deficient DT40 cells, and to preventing early steps in HR by deleting XRCC3 suppressed the nonviability of Xpf-deficient DT40 cells. In summary, Xpf and Mus81 have a substantially overlapping function in completion of HR. (C) 2013 AACR.AMER ASSOC CANCER RESEARCH, Jul. 2013, CANCER RESEARCH, 73(14) (14), 4362 - 4371, English[Refereed]Scientific journal
- May 2013, Nucleic Acids Res, EnglishStructure-function analysis of the EF-hand protein centrin-2 for its intracellular localization and nucleotide excision repair.[Refereed]Scientific journal
- DNA double-strand breaks (DSB) occur frequently during replication in sister chromatids and are dramatically increased when cells are exposed to chemotherapeutic agents including camptothecin. Such DSBs are efficiently repaired specifically by homologous recombination (HR) with the intact sister chromatid. HR, therefore, plays pivotal roles in cellular proliferation and cellular tolerance to camptothecin. Mammalian cells carry several structure-specific endonucleases, such as Xpf-Ercc1 and Mus81-Eme1, in which Xpf and Mus81 are the essential subunits for enzymatic activity. Here, we show the functional overlap between Xpf and Mus81 by conditionally inactivating Xpf in the chicken DT40 cell line, which has no Mus81 ortholog. Although mammalian cells deficient in either Xpf or Mus81 are viable, Xpf inactivation in DT40 cells was lethal, resulting in a marked increase in the number of spontaneous chromosome breaks. Similarly, inactivation of both Xpf and Mus81 in human HeLa cells and murine embryonic stem cells caused numerous spontaneous chromosome breaks. Furthermore, the phenotype of Xpf-deficient DT40 cells was reversed by ectopic expression of human Mus81-Eme1 or human Xpf-Ercc1 heterodimers. These observations indicate the functional overlap of Xpf-Ercc1 and Mus81-Eme1 in the maintenance of genomic DNA. Both Mus81-Eme1 and Xpf-Ercc1 contribute to the completion of HR, as evidenced by the data that the expression of Mus81-Eme1 or Xpf-Ercc1 diminished the number of camptothecin-induced chromosome breaks in Xpf-deficient DT40 cells, and to preventing early steps in HR by deleting XRCC3 suppressed the nonviability of Xpf-deficient DT40 cells. In summary, Xpf and Mus81 have a substantially overlapping function in completion of HR.Apr. 2013, Cancer Res, 73(14) (14), 4362 - 71, English, International magazine[Refereed]Scientific journal
- The human XPC-RAD23B complex and its yeast ortholog, Rad4-Rad23, are the primary initiators of global genome nucleotide excision repair. The interaction of these proteins with damaged DNA was analyzed using model DNA duplexes containing a single fluorescein-substituted dUMP analog as a lesion. An electrophoretic mobility shift assay revealed similarity between human and yeast proteins in DNA binding. Quantitative analyses of XPC/Rad4 binding to the model DNA structures were performed by fluorescent depolarization measurements. XPC-RAD23B and Rad4-Rad23 proteins demonstrate approximately equal binding affinity to the damaged DNA duplex (K-D similar to (0.5 +/- 0.1) and (0.6 +/- 0.3) nM, respectively). Using photoreactive DNA containing 5-iodo-dUMP in defined positions, XPC/Rad4 location on damaged DNA was shown. Under conditions of equimolar binding to DNA both proteins exhibited the highest level of cross-links to 5I-dUMP located exactly opposite the damaged nucleotide. The positioning of the XPC and Rad4 proteins on damaged DNA by photocross-linking footprinting is consistent with x-ray analysis of the Rad4-DNA crystal complex. The identity of the XPC and Rad4 location illustrates the common principles of structure organization of DNA damage-scanning proteins from different Eukarya organisms.AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, Apr. 2013, JOURNAL OF BIOLOGICAL CHEMISTRY, 288(15) (15), 10936 - 10947, English[Refereed]Scientific journal
- The human XPC-RAD23B complex and its yeast ortholog, Rad4-Rad23, are the primary initiators of global genome nucleotide excision repair. The interaction of these proteins with damaged DNA was analyzed using model DNA duplexes containing a single fluorescein-substituted dUMP analog as a lesion. An electrophoretic mobility shift assay revealed similarity between human and yeast proteins in DNA binding. Quantitative analyses of XPC/Rad4 binding to the model DNA structures were performed by fluorescent depolarization measurements. XPC-RAD23B and Rad4-Rad23 proteins demonstrate approximately equal binding affinity to the damaged DNA duplex (K-D similar to (0.5 +/- 0.1) and (0.6 +/- 0.3) nM, respectively). Using photoreactive DNA containing 5-iodo-dUMP in defined positions, XPC/Rad4 location on damaged DNA was shown. Under conditions of equimolar binding to DNA both proteins exhibited the highest level of cross-links to 5I-dUMP located exactly opposite the damaged nucleotide. The positioning of the XPC and Rad4 proteins on damaged DNA by photocross-linking footprinting is consistent with x-ray analysis of the Rad4-DNA crystal complex. The identity of the XPC and Rad4 location illustrates the common principles of structure organization of DNA damage-scanning proteins from different Eukarya organisms.AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, Apr. 2013, JOURNAL OF BIOLOGICAL CHEMISTRY, 288(15) (15), 10936 - 10947, English[Refereed]Scientific journal
- The Biophysical Society of Japan General Incorporated Association, 2013, Seibutsu Butsuri, 53(1) (1), S178, Japanese
- GENETICS SOC JAPAN, Dec. 2012, GENES & GENETIC SYSTEMS, 87(6) (6), 411 - 411, EnglishIdentification of new acetylated proteins involved in DNA damage response[Refereed]
- The WD40-repeat protein DDB2 is essential for efficient recognition and subsequent removal of ultraviolet (UV)-induced DNA lesions by nucleotide excision repair (NER). However, how DDB2 promotes NER in chromatin is poorly understood. Here, we identify poly(ADP-ribose) polymerase 1 (PARP1) as a novel DDB2-associated factor. We demonstrate that DDB2 facilitated poly(ADP-ribosyl)ation of UV-damaged chromatin through the activity of PARP1, resulting in the recruitment of the chromatin-remodeling enzyme ALC1. Depletion of ALC1 rendered cells sensitive to UV and impaired repair of UV-induced DNA lesions. Additionally, DDB2 itself was targeted by poly(ADP-ribosyl)ation, resulting in increased protein stability and a prolonged chromatin retention time. Our in vitro and in vivo data support a model in which poly(ADP-ribosyl)ation of DDB2 suppresses DDB2 ubiquitylation and outline a molecular mechanism for PARP1-mediated regulation of NER through DDB2 stabilization and recruitment of the chromatin remodeler ALC1.ROCKEFELLER UNIV PRESS, Oct. 2012, JOURNAL OF CELL BIOLOGY, 199(2) (2), 235 - 249, English[Refereed]Scientific journal
- Recent work identified the E3 ubiquitin ligase CRL4(Cdt2) as mediating the timely degradation of Cdt1 during DNA replication and following DNA damage. In both cases, proliferating cell nuclear antigen (PCNA) loaded on chromatin mediates the CRL4(Cdt2)-dependent proteolysis of Cdt1. Here, we demonstrate that while replication factor C subunit 1 (RFC1)-RFC is required for Cdt1 degradation after UV irradiation during the nucleotide excision repair process, another RFC complex, Ctf18-RFC, which is known to be involved in the establishment of cohesion, has a key role in Cdt1 degradation in S phase. Cdt1 segments having only the degron, a specific sequence element in target protein for ubiquitination, for CRL4(Cdt2) were stabilized during S phase in Ctf18-depleted cells. Additionally, endogenous Cdt1 was stabilized when both Skp2 and Ctf18 were depleted. Since a substantial amount of PCNA was detected on chromatin in Ctf18-depleted cells, Ctf18 is required in addition to loaded PCNA for Cdt1 degradation in S phase. Our data suggest that Ctf18 is involved in recruiting CRL4(Cdt2) to PCNA foci during S phase. Ctf18-mediated Cdt1 proteolysis occurs independent of cohesion establishment, and depletion of Ctf18 potentiates rereplication. Our findings indicate that individual RFC complexes differentially control CRL4(Cdt2)-dependent proteolysis of Cdt1 during DNA replication and repair.AMER SOC MICROBIOLOGY, Jun. 2012, MOLECULAR AND CELLULAR BIOLOGY, 32(12) (12), 2279 - 2288, English[Refereed]Scientific journal
- Jun. 2012, Mol Cell Biol, 32巻、12号、2279-2288頁, EnglishTwo different replication factor C proteins, Ctf18 and RFC1, separately controlScientific journal
- The DDB1-CUL4-RBX1 (CRL4) ubiquitin ligase family regulates a diverse set of cellular pathways through dedicated substrate receptors (DCAFs). The DCAF DDB2 detects UV-induced pyrimidine dimers in the genome and facilitates nucleotide excision repair. We provide the molecular basis for DDB2 receptor-mediated cyclobutane pyrimidine dimer recognition in chromatin. The structures of the fully assembled DDB1-DDB2-CUL4A/B-RBX1 (CRL4(DDB2)) ligases reveal that the mobility of the ligase arm creates a defined ubiquitination zone around the damage, which precludes direct ligase activation by DNA lesions. Instead, the COP9 signalosome (CSN) mediates the CRL4(DDB2) inhibition in a CSN5 independent, nonenzymatic, fashion. In turn, CSN inhibition is relieved upon DNA damage binding to the DDB2 module within CSN-CRL4(DDB2). The Cockayne syndrome A DCAF complex crystal structure shows that CRL4(DCAF(WD40)) ligases share common architectural features. Our data support a general mechanism of ligase activation, which is induced by CSN displacement from CRL4(DCAF) on substrate binding to the DCAF.Nov. 2011, Cell, 147(5) (5), 1024 - 39, English, International magazine[Refereed]Scientific journal
- Nov. 2011, DNA Repair (ed. Kruman, I.), 453-476, EnglishDNA damage recognition for mammalian global genome nucleotide excision repair.Scientific journal
- Translesion DNA synthesis, a process orchestrated by monoubiquitinated PCNA, is critical for DNA damage tolerance. While the ubiquitin-conjugating enzyme RAD6 and ubiquitin ligase RAD18 are known to monoubiquitinate PCNA, how they are regulated by DNA damage is not fully understood. We show that NBS1 (mutated in Nijmegen breakage syndrome) binds to RAD18 after UV irradiation and mediates the recruitment of RAD18 to sites of DNA damage. Disruption of NBS1 abolished RAD18-dependent PCNA ubiquitination and Polη focus formation, leading to elevated UV sensitivity and mutation. Unexpectedly, the RAD18-interacting domain of NBS1, which was mapped to its C terminus, shares structural and functional similarity with the RAD18-interacting domain of RAD6. These domains of NBS1 and RAD6 allow the two proteins to interact with RAD18 homodimers simultaneously and are crucial for Polη-dependent UV tolerance. Thus, in addition to chromosomal break repair, NBS1 plays a key role in translesion DNA synthesis.Sep. 2011, Molecular cell, 43(5) (5), 788 - 97, English, International magazine[Refereed]Scientific journal
- Translesion DNA synthesis, a process orchestrated by monoubiquitinated PCNA, is critical for DNA damage tolerance. While the ubiquitin-conjugating enzyme RAD6 and ubiquitin ligase RAD18 are known to monoubiquitinate PCNA, how they are regulated by DNA damage is not fully understood. We show that NBS1 (mutated in Nijmegen breakage syndrome) binds to RAD18 after UV irradiation and mediates the recruitment of RAD18 to sites of DNA damage. Disruption of NBS1 abolished RAD18-dependent PCNA ubiquitination and Polη focus formation, leading to elevated UV sensitivity and mutation. Unexpectedly, the RAD18-interacting domain of NBS1, which was mapped to its C terminus, shares structural and functional similarity with the RAD18-interacting domain of RAD6. These domains of NBS1 and RAD6 allow the two proteins to interact with RAD18 homodimers simultaneously and are crucial for Polη-dependent UV tolerance. Thus, in addition to chromosomal break repair, NBS1 plays a key role in translesion DNA synthesis.Sep. 2011, Molecular cell, 43(5) (5), 788 - 97, English, International magazine[Refereed]Scientific journal
- The nucleotide excision repair (NER) system is a fundamental cellular stress response that uses only a handful of DNA binding factors, mutated in the cancer-prone syndrome xeroderma pigmentosum (XP), to detect an astounding diversity of bulky base lesions, including those induced by ultraviolet light, electrophilic chemicals, oxygen radicals and further genetic insults. Several of these XP proteins are characterized by a mediocre preference for damaged substrates over the native double helix but, intriguingly, none of them recognizes injured bases with sufficient selectivity to account for the very high precision of bulky lesion excision. Instead, substrate versatility as well as damage specificity and strand selectivity are achieved by a multistage quality control strategy whereby different subunits of the XP pathway, in succession, interrogate the DNA double helix for a distinct abnormality in its structural or dynamic parameters. Through this step-by-step filtering procedure, the XP proteins operate like a systematic decision making tool, generally known as decision tree analysis, to sort out rare damaged bases embedded in a vast excess of native DNA. The present review is focused on the mechanisms by which multiple XP subunits of the NER pathway contribute to the proposed decision tree analysis of DNA quality in eukaryotic cells. (C) 2011 Elsevier B.V. All rights reserved.ELSEVIER SCIENCE BV, Jul. 2011, DNA REPAIR, 10(7) (7), 673 - 683, English[Refereed]Scientific journal
- To estimate the space-radiation effects separately from other space-environmental effects such as microgravity, frozen human lymphoblastoid TK6 cells were sent to the "Kibo" module of the International Space Station (ISS), preserved under frozen condition during the mission and finally recovered to Earth (after a total of 134 days flight, 72 mSv). Biological assays were performed on the cells recovered to Earth. We observed a tendency of increase (2.3-fold) in thymidine kinase deficient (TK-) mutations over the ground control. Loss of heterozygosity (LOH) analysis on the mutants also demonstrated a tendency of increase in proportion of the large deletion (beyond the TK locus) events, 6/41 in the in-flight samples and 1/17 in the ground control. Furthermore, in-flight samples exhibited 48% of the ground-control level in TK- mutation frequency upon exposure to a subsequent 2 Gy dose of X-rays, suggesting a tendency of radioadaptation when compared with the ground-control samples. The tendency of radioadaptation was also supported by the post-flight assays on DNA double-strand break repair: a 1.8- and 1.7-fold higher efficiency of in-flight samples compared to ground control via non-homologous end-joining and homologous recombination, respectively. These observations suggest that this system can be used as a biodosimeter, because DNA damage generated by space radiation is considered to be accumulated in the cells preserved frozen during the mission, Furthermore, this system is also suggested to be applicable for evaluating various cellular responses to low-dose space radiation, providing a better understanding of biological space-radiation effects as well as estimation of health influences of future space explores.SPRINGER, Mar. 2011, RADIATION AND ENVIRONMENTAL BIOPHYSICS, 50(1) (1), 125 - 134, English[Refereed]Scientific journal
- Jan. 2011, Biochemistry, 76: 16-23, EnglishMultiple DNA damage recognition factors involved in mammalian nucleotide excision repair[Refereed]Scientific journal
- A multifunctional calcium-binding protein, centrin-1, is specifically expressed in male germ cells, certain neurons and ciliated cells. We identified centrin-1 as a protein interacting with SUMO-2/3 using yeast two-hybrid screening of a mouse testicular cDNA library. In bead halo assays, the interaction between centrin-1 and SUMO-2/3 was reduced in the presence of EGTA and facilitated by the addition of CaCl(2), immunostaining of seminiferous tubules in 35-day-old mouse testes revealed that cells in the layer containing spermatogonia showed colocalization of SUMO-2/3 with centrin-1 in cytoplasmic spots. Identification of centrin-1 as the EGTA-sensitive SUMO-2/3-interacting protein indicates the possible role of calcium in modulating the centrin-1-SUMO-2/3 interaction and suggests the importance of this interaction in mouse testis. Copyright (C) 2010 John Wiley & Sons, Ltd.JOHN WILEY & SONS LTD, Oct. 2010, CELL BIOCHEMISTRY AND FUNCTION, 28(7) (7), 604 - 612, English[Refereed]Scientific journal
- We showed that XPC complex, which is a DNA damage detector for nucleotide excision repair, stimulates activity of thymine DNA glycosylase (TDG) that initiates base excision repair. XPC appeared to facilitate the enzymatic turnover of TDG by promoting displacement from its own product abasic site, although the precise mechanism underlying this stimulation has not been clarified. Here we show that XPC has only marginal effects on the activity of E. coli TDG homolog (EcMUG), which remains bound to the abasic site like human TDG but does not significantly interacts with XPC. On the contrary, XPC significantly stimulates the activities of sumoylated TDG and SMUG1, both of which exhibit quite different enzymatic kinetics from unmodified TDG but interact with XPC. These results point to importance of physical interactions for stimulation of DNA glycosylases by XPC and have implications in the molecular mechanisms underlying mutagenesis and carcinogenesis in XP-C patients.Jul. 2010, Journal of nucleic acids, 2010, English, International magazine[Refereed]Scientific journal
- We have developed fluorescent probes for the detection of strand scission in the excision repair of oxidatively damaged bases. They were hairpin-shaped oligonucleotides, each containing an isomer of thymine glycol or 5,6-dihydrothymine as a damaged base in the center, with a fluorophore and a quencher at the 5'- and 3'-ends, respectively. Fluorescence was detected when the phosphodiester linkage at the damage site was cleaved by the enzyme, because the short fragment bearing the fluorophore could not remain in a duplex form hybridized to the rest of the molecule at the incubation temperature. The substrate specificities of Escherichia coli endonuclease III and its human homolog, NTH1, determined by using these probes agreed with those determined previously by gel electrophoresis using (32)P-labeled substrates. Kinetic parameters have also been determined by this method. Since different fluorophores were attached to the oligonucleotides containing each lesion, reactions with two types of substrates were analyzed separately in a single tube, by changing the excitation and detection wavelengths. These probes were degraded during an incubation with a cell extract. Therefore, phosphorothioate linkages were incorporated to protect the probes from nonspecific nucleases, and the base excision repair activity was successfully detected in HeLa cells.OXFORD UNIV PRESS, Apr. 2010, NUCLEIC ACIDS RESEARCH, 38(7) (7), English[Refereed]Scientific journal
- We have developed fluorescent probes for the detection of strand scission in the excision repair of oxidatively damaged bases. They were hairpin-shaped oligonucleotides, each containing an isomer of thymine glycol or 5, 6-dihydrothymine as a damaged base in the center, with a fluorophore and a quencher at the 5′- and 3′-ends, respectively. Fluorescence was detected when the phosphodiester linkage at the damage site was cleaved by the enzyme, because the short fragment bearing the fluorophore could not remain in a duplex form hybridized to the rest of the molecule at the incubation temperature. The substrate specificities of Escherichia coli endonuclease III and its human homolog, NTH1, determined by using these probes agreed with those determined previously by gel electrophoresis using 32P-labeled substrates. Kinetic parameters have also been determined by this method. Since different fluorophores were attached to the oligonucleotides containing each lesion, reactions with two types of substrates were analyzed separately in a single tube, by changing the excitation and detection wavelengths. These probes were degraded during an incubation with a cell extract. Therefore, phosphorothioate linkages were incorporated to protect the probes from nonspecific nucleases, and the base excision repair activity was successfully detected in HeLa cells. © The Author(s) 2010. Published by Oxford University Press.Jan. 2010, Nucleic Acids Research, 38(7) (7), e101, EnglishScientific journal
- For mammalian nucleotide excision repair (NER), DNA lesions are recognized in at least two steps involving detection of unpaired bases by the XPC protein complex and the subsequent verification of injured bases. Although lesion verification is important to ensure high damage discrimination and the accuracy of the repair system, it has been unclear how this is accomplished. Here, we show that damage verification involves scanning of a DNA strand from the site where XPC is initially bound. Translocation by the NER machinery exhibits a 5'-to-3' directionality, strongly suggesting involvement of the XPD helicase, a component of TFIIH. Furthermore, the initial orientation of XPC binding is crucial in that only one DNA strand is selected to search for the presence of lesions. Our results dissect the intricate molecular mechanism of NER and provide insights into a strategy for mammalian cells to survey large genomes to detect DNA damage.Nov. 2009, Molecular cell, 36(4) (4), 642 - 53, English, International magazine[Refereed]Scientific journal
- Nov. 2009, Chembiochem : a European journal of chemical biology, 10(16) (16), 2613 - 6, English, International magazine[Refereed]Scientific journal
- Although the basic principle of nucleotide excision repair (NER), which can eliminate various DNA lesions, have been dissected at the genetic, biochemical and cellular levels, the important in vivo regulation of the critical damage recognition step is poorly understood. Here we analyze the in vivo dynamics of the essential NER damage recognition factor XPC fused to the green fluorescence protein (GFP). Fluorescence recovery after photobleaching analysis revealed that the UV-induced transient immobilization of XPC, reflecting its actual engagement in NER, is regulated in a biphasic manner depending on the number of (6-4) photoproducts and titrated by the number of functional UV-DDB molecules. A similar biphasic UV-induced immobilization of TFIIH was observed using XPB-GFP. Surprisingly, subsequent integration of XPA into the NER complex appears to follow only the low UV dose immobilization of XPC. Our results indicate that when only a small number of (6-4) photoproducts are generated, the UV-DDB-dependent damage recognition pathway predominates over direct recognition by XPC, and they also suggest the presence of rate-limiting regulatory steps in NER prior to the assembly of XPA.Jun. 2009, DNA repair, 8(6) (6), 767 - 76, English, International magazineScientific journal
- Although the basic principle of nucleotide excision repair (NER), which can eliminate various DNA lesions, have been dissected at the genetic, biochemical and cellular levels, the important in vivo regulation of the critical damage recognition step is poorly understood. Here we analyze the in vivo dynamics of the essential NER damage recognition factor XPC fused to the green fluorescence protein (GFP). Fluorescence recovery after photobleaching analysis revealed that the UV-induced transient immobilization of XPC, reflecting its actual engagement in NER, is regulated in a biphasic manner depending on the number of (6-4) photoproducts and titrated by the number of functional UV-DDB molecules. A similar biphasic UV-induced immobilization of TFIIH was observed using XPB-GFP. Surprisingly, subsequent integration of XPA into the NER complex appears to follow only the low UV dose immobilization of XPC. Our results indicate that when only a small number of (6-4) photoproducts are generated, the UV-DDB-dependent damage recognition pathway predominates over direct recognition by XPC, and they also suggest the presence of rate-limiting regulatory steps in NER prior to the assembly of XPA.Jun. 2009, DNA repair, 8(6) (6), 767 - 76, English, International magazine[Refereed]Scientific journal
- Mar. 2009, Mutat. Res., 685: 19-37, EnglishRegulation of damage recognition in mammalian global genomic nucleotide excision repair[Refereed]Scientific journal
- The interaction of the nucleotide excision repair (NER) protein dimeric complex XPC-hHR23B, which is implicated in the DNA damage recognition step, with three Cy3.5 labeled 90-bp double-stranded DNA substrates (unmodified, with a central unpaired region, and cholesterol modified) and a 90-mer single-strand DNA was investigated in solution by fluorescence correlation spectroscopy. Autocorrelation functions obtained in the presence of an excess of protein show larger diffusion times (tau(d)) than for free DNA, indicating the presence of DNA-protein bound complexes. The fraction of DNA bound (theta), as a way to describe the percentage of protein bound to DNA, was directly estimated from FCS data. A significantly stronger binding capability for the cholesterol modified substrate (78% DNA bound) than for other double-stranded DNA substrates was observed, while the lowest affinity was found for the single-stranded DNA (27%). This is in accordance with a damage recognition role of the XPC protein. The similar affinity of XPC for undamaged and 'bubble' DNA substrates (58% and 55%, respectively) indicates that XPC does not specifically bind to this type of DNA substrate comprising a large (30-nt) central unpaired region.SPRINGER/PLENUM PUBLISHERS, Sep. 2008, JOURNAL OF FLUORESCENCE, 18(5) (5), 987 - 995, English[Refereed]Scientific journal
- The interaction of the nucleotide excision repair (NER) protein dimeric complex XPC-hHR23B, which is implicated in the DNA damage recognition step, with three Cy3.5 labeled 90-bp double-stranded DNA substrates (unmodified, with a central unpaired region, and cholesterol modified) and a 90-mer single-strand DNA was investigated in solution by fluorescence correlation spectroscopy. Autocorrelation functions obtained in the presence of an excess of protein show larger diffusion times (τ d) than for free DNA, indicating the presence of DNA-protein bound complexes. The fraction of DNA bound (θ), as a way to describe the percentage of protein bound to DNA, was directly estimated from FCS data. A significantly stronger binding capability for the cholesterol modified substrate (78% DNA bound) than for other double-stranded DNA substrates was observed, while the lowest affinity was found for the single-stranded DNA (27%). This is in accordance with a damage recognition role of the XPC protein. The similar affinity of XPC for undamaged and 'bubble' DNA substrates (58% and 55%, respectively) indicates that XPC does not specifically bind to this type of DNA substrate comprising a large (30-nt) central unpaired region. © 2008 The Author(s).Sep. 2008, Journal of Fluorescence, 18(5) (5), 987 - 995, EnglishScientific journal
- Xeroderma pigmentosum (XP) is an autosomal recessive disease, which is characterized by susceptibility to ultraviolet light (UV)-induced skin cancer. Among eight genes so far identified as responsible for XP, XPA through XPG are involved in nucleotide excision repair of DNA damage induced by UV as well as various chemical carcinogens. Since this repair system removes a major UV photoproduct, the cyclobutane pyrimidine dimer, quite slowly from the global genome, this lesion must be accurately bypassed during replication by DNA polymerase η, encoded by the XPV gene. Recent studies have revealed that each of these XP genes possesses additional functions, some of which are concerned with other DNA repair pathways and/or cellular DNA damage responses. Such differential functions not only explain clinical heterogeneity among different genetic complementation groups but also have implications for the promotion of carcinogenic processes in XP patients. © The Author 2008. Published by Oxford University Press. All rights reserved.Mar. 2008, Carcinogenesis, 29(3) (3), 455 - 465, English[Refereed]Scientific journal
- Mutagenic radioadaptation in a human lymphoblastoid cell line.We investigated the mutagenic radioadaptive response of human lymphoblastoid TK6 cells by pretreating them with a low dose (5 cGy) of X-rays followed by a high (2 Gy) dose 6h later. Pretreatment reduced the 2-Gy-induced mutation frequency (MF) of the thymidine kinase (TK) gene (18.3 x 10(-6)) to 62% of the original level (11.4 x 10(-6)). A loss of heterozygosity (LOH) detection analysis applied to the isolated TK(-) mutants revealed the mutational events as non-LOH (resulting mostly from a point mutation in the TK gene), hemizygous LOH (resulting from a chromosomal deletion), or homozygous LOH (resulting from homologous recombination (HR) between chromosomes). For non-LOH events, pretreatment decreased the frequency to 27% of the original level (from 7.1 x 10(-6) to 1.9 x 10(-6)). cDNAs prepared from the non-LOH mutants revealed that the decrease was due mainly to the repression of base substitutions. The frequency of hemizygous LOH events, however, was not significantly altered by pretreatment. Mapping analysis of chromosome 17 demonstrated that the distribution and the extent of hemizygous LOH events were also not significantly influenced by pretreatment. For homozygous LOH events, pretreatment reduced the frequency to 61% of the original level (from 5.1 x 10(-6) to 3.1 x 10(-6)), reflecting an enhancement in HR repair of DNA double-strand breaks. Our findings suggest that the radioadaptive response in TK6 cells follows mainly from mutations at the base-sequence level, not the chromosome level.Feb. 2008, Mutation research, 638(1-2) (1-2), 48 - 55, English, International magazine[Refereed]Scientific journal
- In vivo destabilization and functional defects of the xeroderma pigmentosum C protein caused by a pathogenic missense mutation.Xeroderma pigmentosum group C (XPC) protein plays an essential role in DNA damage recognition in mammalian global genome nucleotide excision repair (NER). Here, we analyze the functional basis of NER inactivation caused by a single amino acid substitution (Trp to Ser at position 690) in XPC, previously identified in the XPC patient XP13PV. The Trp690Ser change dramatically affects the in vivo stability of the XPC protein, thereby causing a significant reduction of its steady-state level in XP13PV fibroblasts. Despite normal heterotrimeric complex formation and physical interactions with other NER factors, the mutant XPC protein lacks binding affinity for both undamaged and damaged DNA. Thus, this single amino acid substitution is sufficient to compromise XPC function through both quantitative and qualitative alterations of the protein. Although the mutant XPC fails to recognize damaged DNA, it is still capable of accumulating in a UV-damaged DNA-binding protein (UV-DDB)-dependent manner to UV-damaged subnuclear domains. However, the NER factors transcription factor IIH and XPA failed to colocalize stably with the mutant XPC. As well as highlighting the importance of UV-DDB in recruiting XPC to UV-damaged sites, these findings demonstrate the role of DNA binding by XPC in the assembly of subsequent NER intermediate complexes.Oct. 2007, Molecular and cellular biology, 27(19) (19), 6606 - 14, English, International magazine[Refereed]Scientific journal
- Jun. 2007, Tanpakushitsu kakusan koso. Protein, nucleic acid, enzyme, 52(7) (7), 760 - 7, Japanese, Domestic magazine[Roles for ubiquitylation in DNA repair].Scientific journal
- 共立出版, Nov. 2006, 蛋白質核酸酵素, 51(14) (14), 2126-2133 - 2133,1919, Japanese紫外線によるDNA損傷の修復機構
- Intracellular inclusions play a profound role in many neurodegenerative diseases. Here, we report that HR23B and HR23A, proteins that are involved in both DNA repair and shuttling proteins to the 26S proteasome for degradation, accumulate in neuronal inclusions in brain from a mouse model for FXTAS, as well as in brain material from HD, SCA3, SCA7, FTDP-17 and PD patients. Interestingly, HR23B did not significantly accumulate in tau-positive aggregates (neurofibrillary tangles) from AD patients while ubiquitin did. The sequestration of HR23 proteins in intracellular inclusions did not cause detectable accumulation of their stable binding partner in DNA repair, XPC. Surprisingly, no reduction in repair capacity was observed in primary human fibroblasts that overexpressed GFP-polyQ, a polypeptide that induces HR23B-positive inclusions in these transfected cells. This illustrates that impairment of the ubiquitin-proteasome system (UPS) by expanded glutamine repeats, including the sequestration of HR23B, is not affecting NER. (c) 2006 Elsevier Inc. All rights reserved.ACADEMIC PRESS INC ELSEVIER SCIENCE, Sep. 2006, NEUROBIOLOGY OF DISEASE, 23(3) (3), 708 - 716, English[Refereed]Scientific journal
- Jun. 2006, Seikagaku. The Journal of Japanese Biochemical Society, 78(6) (6), 516 - 21, Japanese, Domestic magazine[Repair mechanism of DNA base lesions induced by ultraviolet light irradiation].[Refereed]
- Modification of cellular proteins by the small ubiquitin-like modifier SUMO is important in regulating various cellular events. Many different nuclear proteins are targeted by SUMO, and the functional consequences of this modification are diverse. For most proteins, however, the functional and structural consequences of modification by specific SUMO isomers are unclear. Conjugation of SUMO to thymine-DNA glycosylase (TDG) induces the dissociation of TDG from its product DNA. Structure determination of the TDG central region conjugated to SUMO-1 previously suggested a mechanism in which the SUMOylation-induced conformational change in the C-terminal region of TDG releases TDG from tight binding to its product DNA. Here, we have determined the crystal structure of the central region of TDG conjugated to SUMO-3. The overall structure of SUMO-3-conjugated TDG is similar to the previously reported structure of TDG conjugated to SUMO-1, despite the relatively low level of amino acid sequence similarity between SUMO-3 and SUMO-1. The two structures revealed that the sequence of TDG that resembles the SUMO-binding motif (SBM) can form an intermolecular beta-sheet with either SUMO-1 or SUMO-3. Structural comparison with the canonical SBM shows that this SBM-like sequence of TDG retains all of the characteristic interactions of the SBM, indicating sequence diversity in the SBM.May 2006, Journal of molecular biology, 359(1) (1), 137 - 47, English, International magazine[Refereed]Scientific journal
- Centrin 2 stimulates nucleotide excision repair by interacting with xeroderma pigmentosum group C protein.Xeroderma pigmentosum group C (XPC) protein plays a key role in DNA damage recognition in global genome nucleotide excision repair (NER). The protein forms in vivo a heterotrimeric complex involving one of the two human homologs of Saccharomyces cerevisiae Rad23p and centrin 2, a centrosomal protein. Because centrin 2 is dispensable for the cell-free NER reaction, its role in NER has been unclear. Binding experiments with a series of truncated XPC proteins allowed the centrin 2 binding domain to be mapped to a presumed alpha-helical region near the C terminus, and three amino acid substitutions in this domain abrogated interaction with centrin 2. Human cell lines stably expressing the mutant XPC protein exhibited a significant reduction in global genome NER activity. Furthermore, centrin 2 enhanced the cell-free NER dual incision and damaged DNA binding activities of XPC, which likely require physical interaction between XPC and centrin 2. These results reveal a novel vital function for centrin 2 in NER, the potentiation of damage recognition by XPC.Jul. 2005, Molecular and cellular biology, 25(13) (13), 5664 - 74, English, International magazine[Refereed]
- Signal transducer and activator of transcription 3 is a key regulator of keratinocyte survival and proliferation following UV irradiationUVB irradiation of signal transducer and activator of transcription 3 (Stat3)-deficient keratinocytes resulted in a high incidence of apoptosis compared with controls. Conversely, forced expression of Stat3 desensitized keratinocytes to UVB-induced apoptosis. Upon UVB exposure, keratinocyte Stat3 was rapidly dephosphorylated, followed by decreases of both Stat3 mRNA and protein levels in a p53-independent manner. Vanadate treatment reversed the UVB-induced down-regulation of Stat3 and generation of apoptotic keratinocytes, suggesting the involvement of a tyrosine phosphatase. Furthermore, Stat3 was required for UVB-induced proliferation of follicular keratinocytes, leading to epidermal thickening. Finally, constitutive activation of Stat3 was observed in UVB-induced squamous cell carcinomas of either mice or human origin. These data suggest that Stat3 is required for survival and proliferation of keratinocytes following UVB exposure and that Stat3 is tightly regulated as part of a novel protective mechanism against UVB-induced skin cancer.AMER ASSOC CANCER RESEARCH, Jul. 2005, CANCER RESEARCH, 65(13) (13), 5720 - 5729, English[Refereed]Scientific journal
- Crystal structure of thymine DNA glycosylase conjugated to SUMO-1.Members of the small ubiquitin-like modifier (SUMO) family can be covalently attached to the lysine residue of a target protein through an enzymatic pathway similar to that used in ubiquitin conjugation, and are involved in various cellular events that do not rely on degradative signalling via the proteasome or lysosome. However, little is known about the molecular mechanisms of SUMO-modification-induced protein functional transfer. During DNA mismatch repair, SUMO conjugation of the uracil/thymine DNA glycosylase TDG promotes the release of TDG from the abasic (AP) site created after base excision, and coordinates its transfer to AP endonuclease 1, which catalyses the next step in the repair pathway. Here we report the crystal structure of the central region of human TDG conjugated to SUMO-1 at 2.1 A resolution. The structure reveals a helix protruding from the protein surface, which presumably interferes with the product DNA and thus promotes the dissociation of TDG from the DNA molecule. This helix is formed by covalent and non-covalent contacts between TDG and SUMO-1. The non-covalent contacts are also essential for release from the product DNA, as verified by mutagenesis.Jun. 2005, Nature, 435(7044) (7044), 979 - 82, English, International magazine[Refereed]Scientific journal
- UV-induced ubiquitylation of XPC protein mediated by UV-DDB-ubiquitin ligase complex.The xeroderma pigmentosum group C (XPC) protein complex plays a key role in recognizing DNA damage throughout the genome for mammalian nucleotide excision repair (NER). Ultraviolet light (UV)-damaged DNA binding protein (UV-DDB) is another complex that appears to be involved in the recognition of NER-inducing damage, although the precise role it plays and its relationship to XPC remain to be elucidated. Here we show that XPC undergoes reversible ubiquitylation upon UV irradiation of cells and that this depends on the presence of functional UV-DDB activity. XPC and UV-DDB were demonstrated to interact physically, and both are polyubiquitylated by the recombinant UV-DDB-ubiquitin ligase complex. The polyubiquitylation altered the DNA binding properties of XPC and UV-DDB and appeared to be required for cell-free NER of UV-induced (6-4) photoproducts specifically when UV-DDB was bound to the lesion. Our results strongly suggest that ubiquitylation plays a critical role in the transfer of the UV-induced lesion from UV-DDB to XPC.May 2005, Cell, 121(3) (3), 387 - 400, English, International magazine[Refereed]
- Nucleosomal structure of undamaged DNA regions suppresses the non-specific DNA binding of the XPC complex.The XPC protein complex is a DNA damage detector of human nucleotide excision repair (NER). Although the XPC complex specifically binds to certain damaged sites, it also binds to undamaged DNA in a non-specific manner. The addition of a large excess of undamaged naked DNA competitively inhibited the specific binding of the XPC complex to (6-4) photoproducts and the NER dual incision step in cell-free extracts. In contrast, the addition of undamaged nucleosomal DNA as a competitor suppressed both of these inhibitory effects. Although nucleosomes positioned on the damaged site inhibited the binding of the XPC complex, the presence of nucleosomes in undamaged DNA regions may help specific binding of the XPC complex to damaged sites by excluding its non-specific binding to undamaged DNA regions.Mar. 2005, DNA repair, 4(3) (3), 389 - 95, English, International magazine[Refereed]
- The Biophysical Society of Japan General Incorporated Association, 2005, Seibutsu Butsuri, 45, S9, English
- Relative levels of the two mammalian Rad23 homologs determine composition and stability of the xeroderma pigmentosum group C protein complex.Mammalian cells express two Rad23 homologs, HR23A and HR23B, which have been implicated in regulation of proteolysis via the ubiquitin/proteasome pathway. Recently, the proteins have been shown to stabilize xeroderma pigmentosum group C (XPC) protein that is involved in DNA damage recognition for nucleotide excision repair (NER). Because the vast majority of XPC forms a complex with HR23B rather than HR23A, we investigated possible differences between the two Rad23 homologs in terms of their effects on the XPC protein. In wild-type mouse embryonic fibroblasts (MEFs), endogenous XPC was found to be relatively stable, while its steady-state level and stability appeared significantly reduced by targeted disruption of the mHR23B gene, but not by that of mHR23A. Loss of both mHR23 genes caused a strong further reduction of the XPC protein level. Quantification of the two mHR23 proteins revealed that in normal cells mHR23B is actually approximately 10 times more abundant than mHR23A. In addition, overexpression of mHR23A in the mHR23A/B double knock out cells restored not only the steady-state level and stability of the XPC protein, but also cellular NER activity to near wild-type levels. These results indicate that the two Rad23 homologs are largely functionally equivalent in NER, and that the difference in expression levels explains for a major part the difference in complex formation with as well as stabilization effects on XPC.Oct. 2004, DNA repair, 3(10) (10), 1285 - 95, English, International magazine[Refereed]
- Aug. 2004, Analytical biochemistry, 331(1) (1), 204 - 6, English, International magazineScientific journal
- Nucleotide excision repair of 5-formyluracil in vitro is enhanced by the presence of mismatched bases.5-Formyluracil (fU) is a major thymine lesion produced by reactive oxygen radicals and photosensitized oxidation. Although this residue is a potentially mutagenic lesion and is removed by several base excision repair enzymes, it is unknown whether fU is the substrate of nucleotide excision repair (NER). Here, we analyzed the binding specificity of XPC-HR23B, which initiates NER, and cell-free NER activity on fU opposite four different bases. The result of the gel mobility shift assay showed that XPC-HR23B binds the fU-containing substrates in the following order: fU:C > fU:T > fU:G > fU:A. Furthermore, in the presence of XPC-HR23B, the dual incision activity was the same as the order of the binding affinity of XPC-HR23B to fU. Therefore, it is concluded that even fU, regarded as a shape mimic of thymine, can be recognized as a substrate of NER incision, and the efficiency depends on instability of the base pair.Mar. 2004, Biochemistry, 43(10) (10), 2682 - 7, English, International magazine[Refereed]
- Structure of the ubiquitin-interacting motif of S5a bound to the ubiquitin-like domain of HR23B.Ubiquitination, a modification in which single or multiple ubiquitin molecules are attached to a protein, serves signaling functions that control several cellular processes. The ubiquitination signal is recognized by downstream effectors, many of which carry a ubiquitin-interacting motif (UIM). Such interactions can be modulated by regulators carrying a ubiquitin-like (UbL) domain, which binds UIM by mimicking ubiquitination. Of them, HR23B regulates the proteasomal targeting of ubiquitinated substrates, DNA repair factors, and other proteins. Here we report the structure of the UIM of the proteasome subunit S5a bound to the UbL domain of HR23B. The UbL domain presents one hydrophobic and two polar contact sites for interaction with UIM. The residues in these contact sites are well conserved in ubiquitin, but ubiquitin also presents a histidine at the interface. The pH-dependent protonation of this residue interferes with the access of ubiquitin to the UIM and the ubiquitin-associated domain (UBA), and its mutation to a smaller residue increases the affinity of ubiquitin for UIM.Feb. 2004, The Journal of biological chemistry, 279(6) (6), 4760 - 7, English, International magazine[Refereed]
- Translesion synthesis by human DNA polymerase eta across oxidative products of guanine.Guanine is the most oxidizable base among natural bases. 8-Oxoguanine (8-oxoG) is the typical oxidative product, but the amount of 8-oxoG does not directly reflect the strength of oxidative stress. Imidazolone, oxazolone and guanidinohydantoin are oxidative products of guanine and 8-oxoG. Here, we investigated enzymatic reactions with human DNA polymerase eta on these lesions.2004, Nucleic acids symposium series (2004), (48) (48), 171 - 2, English, International magazine[Refereed]
- Thymine-rich single-stranded DNA activates Mcm4/6/7 helicase on Y-fork and bubble-like substrates.The presence of multiple clusters of runs of asymmetric adenine or thymine is a feature commonly found in eukaryotic replication origins. Here we report that the helicase and ATPase activities of the mammalian Mcm4/6/7 complex are activated specifically by thymine stretches. The Mcm helicase is specifically activated by a synthetic bubble structure which mimics an activated replication origin, as well as by a Y-fork structure, provided that a single-stranded DNA region of sufficient length is present in the unwound segment or 3' tail, respectively, and that it carries clusters of thymines. Sequences derived from the human lamin B2 origin can serve as a potent activator for the Mcm helicase, and substitution of its thymine clusters with guanine leads to loss of this activation. At the fork, Mcm displays marked processivity, expected for a replicative helicase. These findings lead us to propose that selective activation by stretches of thymine sequences of a fraction of Mcm helicases loaded onto chromatin may be the determinant for selection of initiation sites on mammalian genomes.Nov. 2003, The EMBO journal, 22(22) (22), 6148 - 60, English, International magazine[Refereed]
- Primary DNA damage sensing in mammalian global genome nucleotide excision repair (GG-NER) is performed by the xeroderma pigmentosum group C (XPC)/HR23B protein complex. HR23B and HR23A are human homologs of the yeast ubiquitin-domain repair factor RAD23, the function of which is unknown. Knockout mice revealed that mHR23A and mHR23B have a fully redundant role in NER, and a partially redundant function in embryonic development. Inactivation of both genes causes embryonic lethality, but appeared still compatible with cellular viability. Analysis of mHR23A/B double-mutant cells showed that HR23 proteins function in NER by governing XPC stability via partial protection against proteasomal degradation. Interestingly, NER-type DNA damage further stabilizes XPC and thereby enhances repair. These findings resolve the primary function of RAD23 in repair and reveal a novel DNA-damage-dependent regulation mechanism of DNA repair in eukaryotes, which may be part of a more global damage-response circuitry.COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT, Jul. 2003, GENES & DEVELOPMENT, 17(13) (13), 1630 - 1645, English[Refereed]Scientific journal
- Elsevier, May 2003, DNA Repair, 2(5) (5), 639 - 652, English[Refereed]International conference proceedings
- DNA bending by the human damage recognition complex XPC-HR23BGenome integrity is maintained, despite constant assault on DNA, due to the action of a variety of DNA repair pathways. Nucleotide excision repair (NER) protects the genome from the deleterious effects of UV irradiation as well as other agents that induce chemical changes in DNA bases. The mechanistic steps required for eukaryotic NER involve the concerted action of at least six proteins or protein complexes. The specificity to incise only the DNA strand including the damage at defined positions is determined by the coordinated assembly of active protein complexes onto damaged DNA. In order to understand the molecular mechanism of the NER reactions and the origin of this specificity and control we analyzed the architecture of functional NER complexes at nanometer resolution by scanning force microscopy (SFM). In the initial step of damage recognition by XPC-HR23B we observe a protein induced change in DNA conformation. XPC-HR23B induces a bend in DNA upon binding and this is stabilized at the site of damage. We discuss the importance of the XPC-HR23B-induced distortion as an architectural feature that can be exploited for subsequent assembly of an active NER complex. (C) 2002 Elsevier Science B.V. All rights reserved.ELSEVIER SCIENCE BV, Mar. 2003, DNA REPAIR, 2(3) (3), 325 - 336, English[Refereed]Scientific journal
- Xeroderma pigmentosum group C protein interacts physically and functionally with thymine DNA glycosylase.The XPC-HR23B complex recognizes various helix-distorting lesions in DNA and initiates global genome nucleotide excision repair. Here we describe a novel functional interaction between XPC-HR23B and thymine DNA glycosylase (TDG), which initiates base excision repair (BER) of G/T mismatches generated by spontaneous deamination of 5-methylcytosine. XPC-HR23B stimulated TDG activity by promoting the release of TDG from abasic sites that result from the excision of mismatched T bases. In the presence of AP endonuclease (APE), XPC-HR23B had an additive effect on the enzymatic turnover of TDG without significantly inhibiting the subsequent action of APE. Our observations suggest that XPC-HR23B may participate in BER of G/T mismatches, thereby contributing to the suppression of spontaneous mutations that may be one of the contributory factors for the promotion of carcinogenesis in xeroderma pigmentosum genetic complementation group C patients.Jan. 2003, The EMBO journal, 22(1) (1), 164 - 73, English, International magazine[Refereed]
- The carboxy-terminal domain of the XPC protein plays a crucial role in nucleotide excision repair through interactions with transcription factor IIH.The xeroderma pigmentosum group C (XPC) protein specifically involved in genome-wide damage recognition for nucleotide excision repair (NER) was purified as a tight complex with HR23B, one of the two mammalian homologs of RAD23 in budding yeast. This XPC-HR23B complex exhibits strong binding affinity for single-stranded DNA, as well as preferential binding to various types of damaged DNA. To examine the structure-function relationship of XPC, a series of truncated mutant proteins were generated and assayed for various binding activities. The two domains participating in binding to HR23B and damaged DNA, respectively, were mapped within the carboxy-terminal half of XPC, which also contains an evolutionary conserved amino acid sequence homologous to the yeast RAD4 protein. We established that the carboxy-terminal 125 amino acids are dispensable for both HR23B and damaged DNA binding, while interactions with transcription factor IIH (TFIIH) are significantly impaired by truncation of this domain. Furthermore, deletion of the extreme carboxy-terminal domain totally abolished XPC activity in the cell-free NER reaction. These results suggest that following initial damage recognition, the carboxy terminus of XPC may be essential for the recruitment of TFIIH, and that most truncation mutations identified in XP-C patients result in non-functional proteins.Jun. 2002, DNA repair, 1(6) (6), 449 - 61, English, International magazine[Refereed]Scientific journal
- Developmental defects and male sterility in mice lacking the ubiquitin-like DNA repair gene mHR23B.mHR23B encodes one of the two mammalian homologs of Saccharomyces cerevisiae RAD23, a ubiquitin-like fusion protein involved in nucleotide excision repair (NER). Part of mHR23B is complexed with the XPC protein, and this heterodimer functions as the main damage detector and initiator of global genome NER. While XPC defects exist in humans and mice, mutations for mHR23A and mHR23B are not known. Here, we present a mouse model for mHR23B. Unlike XPC-deficient cells, mHR23B(-/-) mouse embryonic fibroblasts are not UV sensitive and retain the repair characteristics of wild-type cells. In agreement with the results of in vitro repair studies, this indicates that mHR23A can functionally replace mHR23B in NER. Unexpectedly, mHR23B(-/-) mice show impaired embryonic development and a high rate (90%) of intrauterine or neonatal death. Surviving animals display a variety of abnormalities, including retarded growth, facial dysmorphology, and male sterility. Such abnormalities are not observed in XPC and other NER-deficient mouse mutants and point to a separate function of mHR23B in development. This function may involve regulation of protein stability via the ubiquitin/proteasome pathway and is not or only in part compensated for by mHR23A.Feb. 2002, Molecular and cellular biology, 22(4) (4), 1233 - 45, English, International magazine[Refereed]
- A molecular mechanism for DNA damage recognition by the xeroderma pigmentosum group C protein complex.The XPC-HR23B complex is involved in DNA damage recognition and the initiation of global genomic nucleotide excision repair (GG-NER). Our previous studies demonstrate that XPC-HR23B recognizes and binds DNA containing a helix distortion, regardless of the presence or absence of damaged bases. Here, we describe an extended analysis of the DNA binding specificity of XPC-HR23B using various defined DNA substrates. Although XPC-HR23B showed significantly higher affinity for single-stranded DNA than double-stranded DNA, specific secondary structures of DNA, involving a single- and double-strand junction, were strongly preferred by the complex. This indicates that the presence of bases, which cannot form normal Watson-Crick base pairs in double-stranded DNA, is a critical factor in determining the specificity of XPC-HR23B binding. A DNase I footprint analysis, using a looped DNA substrate, revealed that a single XPC-HR23B complex protected a distorted site in an asymmetrical manner, consistent with the preferred secondary structure. The specific binding of XPC-HR23B is undoubtedly an important molecular process, based on which NER machinery detects a wide variety of lesions that vary in terms of chemical structure during DNA repair.Jan. 2002, DNA repair, 1(1) (1), 95 - 107, English, International magazine[Refereed]
- Centrosome protein centrin 2/caltractin 1 is part of the xeroderma pigmentosum group C complex that initiates global genome nucleotide excision repairNucleotide excision repair (NER) is carried out by xeroderma pigmentosum (XP) factors. Before the excision reaction, DNA damage is recognized by a complex originally thought to contain the XP group C responsible gene product (XPC) and the human homologue of Rad23 B (HR23B). Here, we show that centrin 2/caltractin 1 (CEN2) is also a component of the XPC repair complex. We demonstrate that nearly all XPC complexes contain CEN2, that CEN2 interacts directly with XPC, and that CEN2, in cooperation with HR23B, stabilizes XPC, which stimulates XPC NER activity in vitro. CEN2 has been shown to play an important role in centrosome duplication. Thus, those findings suggest that the XPC-CEN2 interaction may reflect coupling of cell division and NER.AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, Jun. 2001, JOURNAL OF BIOLOGICAL CHEMISTRY, 276(22) (22), 18665 - 18672, English[Refereed]Scientific journal
- Diversity of the damage recognition step in the global genomic nucleotide excision repair in vitroThe XPC-HR23B complex, a mammalian factor specifically involved in global genomic nucleotide excision repair (NER) has been shown to bind various forms of damaged DNA and initiate DNA repair in cell-free reactions. To characterize the binding specificity of this factor in more detail, a method based on immunoprecipitation was developed to assess the relative affinity of XPC-HR23B for defined lesions on DNA, Here we show that XPC-HR23B preferentially binds to UV-induced (6-4) photoproducts (6-4PPs) as well as to cholesterol, but not to the cyclobutane pyrimidine dimer (CPD), 8-oxoguanine (8-oxo-G), O-6-methylguanine (O-6-Me-G), or a single mismatch. Human whole cell extracts could efficiently excise 6-4PPs and cholesterol in an XPC-HR23B-dependent manner, but not 8-oxo-G, O-6-Me-G or mismatches. Thus, there was good correlation between the binding specificity of XPC-HR23B for certain types of lesion and the ability of human cell extracts to excise these lesions, supporting the model that XPC-HR23B initiates global genomic NER. Although, XPC-HR23B does not preferentially bind to CPDs, the excision of CPDs in human whole cell extracts was found to be absolutely dependent on XPC-HR23B, in agreement with the in vivo observation that CPDs are not removed from the global genome in XP-C mutant cells. These results suggest that, in addition to the excision repair pathway initiated by XPC-HR23B, there exists another sub-pathway for the global genomic NER that still requires XPC-HR23B but is not initiated by XPC-HR23B, Possible mechanisms will be discussed, (C) 2001 Elsevier Science B.V. All rights reserved.ELSEVIER SCIENCE BV, Apr. 2001, MUTATION RESEARCH-DNA REPAIR, 485(3) (3), 219 - 227, English[Refereed]Scientific journal
- A multistep damage recognition mechanism for global genomic nucleotide excision repairA mammalian nucleotide excision repair (NER) factor, the XPC-HR23B complex, can specifically bind to certain DNA lesions and initiate the cell-free repair reaction. Here we describe a detailed analysis of its binding specificity using various DNA substrates, each containing a single defined lesion. A highly sensitive gel mobility shift assay revealed that XPC-HR23B specifically binds a small bubble structure with or without damaged bases, whereas dual incision takes place only when damage is present in the bubble. This is evidence that damage recognition for NER is accomplished through at least two steps; XPC-HR23B first binds to a site that has a DNA helix distortion, and then the presence of injured bases is verified prior to dual incision. Cyclobutane pyrimidine dimers (CPDs) were hardly recognized by XPC-HR23B, suggesting that additional factors may be required for CPD recognition. Although the presence of mismatched bases opposite a CPD potentiated XPC-HR23B binding, probably due to enhancement of the helix distortion, cell-free excision of such compound lesions was much more efficient than expected from the observed affinity for XPC-HR23B. This also suggests that additional factors and steps are required for the recognition of some types of lesions. A multistep mechanism of this sort may provide a molecular basis for ensuring the high level of damage discrimination that is required for global genomic NER.COLD SPRING HARBOR LAB PRESS, Mar. 2001, GENES & DEVELOPMENT, 15(5) (5), 507 - 521, English[Refereed]Scientific journal
- Novel functional interactions between nucleotide excision DNA repair proteins influencing the enzymatic activities of TFIIH, XPG, and ERCC1-XPFThe multisubunit basal transcription factor III-I (TFIIH) has a dual involvement in nucleotide excision repair (NER) of a variety of DNA lesions, including UV-induced photoproducts, and RNA polymerase II transcription, In both processes, TFIIH is implicated with local DNA unwinding, which is attributed to its helicase subunits XPB and XPD. To further define the role of TFIIH in NER, functional interactions between TFIIH and other DNA repair proteins were analyzed. We show that the TFIIH-associated ATPase activity is stimulated by both XPA and the XPC-HR23B complex. However, while XPA promotes the ATPase activity specifically in the presence of damaged DNA, stimulation by XPC-HR23B is lesion independent. Furthermore, we reveal that TFIIH inhibits the structure-specific endonuclease activities of both XPG and ERCC1-XPF, responsible for the 3 ' and 5 ' incision in NER, respectively. The inhibition occurs in the absence of ATP and is reversed upon addition of ATP, These results point toward additional roles for TFIIH and ATP during NER distinct from a requirement for DNA unwinding in the regulation of the endonuclease activities of XPG and ERCC1-XPF.AMER CHEMICAL SOC, Jan. 2001, BIOCHEMISTRY, 40(1) (1), 160 - 165, English[Refereed]Scientific journal
- Retinoblastoma susceptibility protein, Rb, possesses multiple BRCT-Ws, BRCA1 carboxyl-terminus-related W regions with DNA break-binding activityThe BRCT region, the carboxyl-terminus of BRCA1 (the breast cancer susceptibility gene I product), is ubiquitous in several proteins that participate in cell cycle checkpoints and DNA repair. We have previously shown that the BRCT regions pf TopBP1 (DNA topoisomerase II binding protein 1) and BRCA1 bound DNA breaks. A BRCT-related region, BRCT-W1, in the retinoblastoma susceptibility gene product (Rb) also could bind DNA fragments, independently of DNA sequences. Five BRCT-W regions were found in the Rb family. All BRCT-Ws of Rb bound DNA fragments. Electron microscopy and treatment with an exonuclease showed that BRCT-Ws bound double-strand DIVA breaks. Since some BRCTs are exceptional common relating elements in tumor suppression, our findings reveal novel aspects of the tumor suppression mechanism.STOCKTON PRESS, Apr. 2000, ONCOGENE, 19(16) (16), 1982 - 1991, English[Refereed]Scientific journal
- Mar. 2000, Mutation research, 459(2) (2), 147 - 160[Refereed]
- The xeroderma pigmentosum group C protein complex XPC-HR23B plays an important role in the recruitment of transcription factor IIH to damaged DNAThe xeroderma pigmentosum group C protein complex XPC-HR23B was first isolated as a factor that complemented nucleotide excision repair defects of XP-C cell extracts in vitro. Recent studies have revealed that this protein complex plays an important role in the early steps of global genome nucleotide excision repair, especially in damage recognition, open complex formation, and repair protein complex formation. However, the precise function of XPC-HR23B in global genome repair is still unclear. Here we demonstrate that XPC-HR23B interacts with general transcription factor IIH (TFIIH) both in vivo and in vitro. This interaction is thought to be mediated through the specific affinity of XPC for the TFIIH subunits XPB and/or p62, which are essential for both basal transcription and nucleotide excision repair, Interestingly, association of TFIIH with DNA was observed in both wild-type and XP-A cell extracts but not in XP-C cell extracts, and XPC-HR23B could restore the association of TFIIH with DNA in XP-C cell extracts. Moreover, we found that XPC-HR23B was necessary for efficient association of TFIIH with damaged DNA in cell-free extracts. We conclude that the XPC-HR23B protein complex plays a crucial role in the recruitment of TFIIH to damaged DNA in global genome repair.AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, Mar. 2000, JOURNAL OF BIOLOGICAL CHEMISTRY, 275(13) (13), 9870 - 9875, English[Refereed]Scientific journal
- Reconstitution of damage DNA excision reaction from SV40 minichromosomes with purified nucleotide excision repair proteinsWe previously constructed the cell-free nucleotide excision repair (NER) assay system with UV-irradiated SV40 minichromosomes to analyze the mechanism of NER reaction on chromatin DNA, Here we investigate the factor that acts especially on nucleosomal DNA during the damage excision reaction, and reconstitute the damage excision reaction on SV40 minichromosomes. NER-proficient HeLa whole cell extracts were fractionated, and the amounts of known NER factors involved in the column fractions were determined by immunoblot analyses. The column fractions were quantitatively and systematically replaced by highly purified NER factors. Finally, damage DNA excision reaction on SV40 minichromosomes was reconstituted with six highly purified NER factors, XPA, XPC-HR23B, XPF-ERCC1, XPG, RPA and TFIIH, as those essential for the reaction with naked DNA. Further analysis showed that the damages on chromosomal DNA were excised as the same efficiency as those on naked DNA for short incubation. At longer incubation time, however, the damage excision efficiency on nucleosomal DNA was decreased whereas naked DNA was still vigorously repaired. These observations suggest that although the six purified NER factors have a potential to eliminate the damage DNA from SV40 minichromosomes, the chromatin structure may still have some repressive effects on NER. (C) 2000 Elsevier Science B.V. All rights reserved.ELSEVIER SCIENCE BV, Mar. 2000, MUTATION RESEARCH-DNA REPAIR, 459(2) (2), 147 - 160, English[Refereed]Scientific journal
- Interaction of hHR23 with S5a - The ubiquitin-like domain of hHR23 mediates interaction with S5a subunit of 26 S proteasomehHR23B is one of two human homologs of the Saccharomyces cerevisiae nucleotide excision repair (NER) gene product RAD23 and a component of a protein complex that specifically complements the NER defect of xeroderma pigmentosum group C (XP-C) cell extracts in vitro. Although a small proportion of hHR23B is tightly complexed with the XP-C responsible gene product, XPC protein, a vast majority exists as an XPC-free form, indicating that hHR23B has additional functions other than NER in vivo. Here we demonstrate that the human NER factor hHR23B as well as another human homolog of RAD23, hHR23A, interact specifically with S5a, a subunit of the human 26 S proteasome using the yeast two-hybrid system. Furthermore, hHR23 proteins were detected with S5a at the position where 26 S proteasome sediments in glycerol gradient centrifugation of HeLa S100 extracts, intriguingly, hHR23B showed the inhibitory effect on the degradation of I-125-lysozyme in the rabbit reticulocyte lysate. hHR23 proteins thus appear to associate with 26 S proteasome in vivo. From co-precipitation experiments using several series of deletion mutants, we defined the domains in hHR23B and S5a that mediate this interaction. From these results, we propose that part of hHR23 proteins are involved in the proteolytic pathway in cells.AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, Sep. 1999, JOURNAL OF BIOLOGICAL CHEMISTRY, 274(39) (39), 28019 - 28025, English[Refereed]Scientific journal
- (公社)日本薬学会, Mar. 1999, 日本薬学会年会要旨集, 119年会(3) (3), 114 - 114, JapaneseXPC蛋白質のNER初期反応に果たす役割についての解析
- DNA-binding polarity of human replication protein A positions nucleases in nucleotide excision repairThe human single-stranded DNA-Binding replication A protein (RPA) is involved in various DNA-processing events. By comparing the affinity of hRPA for artificial DNA hairpin structures with 3'- or 5'-protruding single-stranded arms, rye found that hRPA Binds ssDNA with a defined polarity; a strong ssDNA interaction domain of hRPA is positioned at the 5' side of its Binding region, re weak ssDNA-binding domain resides at the 3' side. Polarity appears crucial for positioning of the excision repair nucleases XPG and ERCC1-XPF ore the DNA. With the 3'-oriented side of hRPA facing a duplex ssDNA junction, hRPA interacts with and stimulates ERCC1-XPF, whereas the S'-oriented side of hRPA at a DNA junction allows stable binding of XPG to hRPA. Our data pinpoint hRPA to the undamaged strand during nucleotide excision repair. polarity of hRPA on ssDNA is likely to contribute to the directionality of other hRPA-dependent processes as well.COLD SPRING HARBOR LAB PRESS, Aug. 1998, GENES & DEVELOPMENT, 12(16) (16), 2598 - 2609, English[Refereed]Scientific journal
- Xeroderma pigmentosum group C protein complex is the initiator of global genome nucleotide excision repairThe XPC-HR23B complex is specifically involved in global genome but not transcription-coupled nucleotide excision repair (NER). Its function is unknown. Using a novel DNA damage recognition-competition assay, we identified XPC-HR23B as the earliest damage detector to initiate NER: it acts before the known damage-binding protein XPA. Coimmunoprecipitation and DNase I footprinting show that XPC-HR23B binds to a variety of NER lesions. These results resolve the function of XPC-HR23B, define the first NER stages, and suggest a two-step mechanism of damage recognition involving damage detection by XPC-HR23B followed by damage verification by XPA. This provides a plausible explanation for the extreme damage specificity exhibited by global genome repair. In analogy, in the transcription-coupled NER subpathway, RNA polymerase II may take the role of XPC. After this subpathway-specific initial lesion detection, XPA may function as a common damage verifier and adaptor to the core of the NER apparatus.CELL PRESS, Aug. 1998, MOLECULAR CELL, 2(2) (2), 223 - 232, English[Refereed]Scientific journal
- Identification and characterization of XPC-binding domain of hHR23BhHR23B was originally isolated as a component of a protein complex that specifically complements nucleotide excision repair (NER) defects of xeroderma pigmentosum group C cell extracts in vitro and was identified as one of two human homologs of the Saccharomyces cerevisiae NER gene product Rad23. Recombinant hHR23B has previously been shown to significantly stimulate the NER activity of recombinant human XPC protein (rhXPC). In this study we identify and functionally characterize the XPC-binding domain of hHR23B protein. We prepared various internal as well as terminal deletion products of hHR23B protein in a His-tagged form and examined their binding with rhXPC by using nickel-chelating Sepharose. We demonstrate that a domain covering 56 amino acids of hHR23B is required for binding to rhXPC as well as for stimulation of in vitro NER reactions. Interestingly, a small polypeptide corresponding to the XPC-binding domain is sufficient to exert stimulation of XPC NER activity. Comparison with known crystal structures and analysis with secondary structure programs provided strong indications that the binding domain has a predominantly amphipathic alpha-helical character, consistent with evidence that the affinity with XPC is based on hydrophobic interactions. Our work shows that binding to XPC alone is required and sufficient for the role of hKR23B in in vitro NER but does not rule out the possibility that the protein has additional functions in vivo.AMER SOC MICROBIOLOGY, Dec. 1997, MOLECULAR AND CELLULAR BIOLOGY, 17(12) (12), 6915 - 6923, English[Refereed]Scientific journal
- Two human homologs of Rad23 are functionally interchangeable in complex formation and stimulation of XPC repair activityXPC-hHR23B protein complex is specifically involved in nucleotide excision repair (NER) of DNA lesions on transcriptionally inactive sequences as,yell as the nontranscribed strand of active genes. Here we demonstrate that not only highly purified recombinant hHR23B (rhHR23B) but also a second human homolog of the Saccharomyces cerevisiae Rad23 repair protein, hHR23A, stimulates the in vitro repair activity of recombinant human XPC (rhXPC), revealing functional redundancy between these human Rad23 homologs. Coprecipitation experiments with His-tagged rhHR23 as well as sedimentation velocity analysis showed that both rhHR23 proteins in vitro reconstitute a physical complex with rhXPC. Both complexes were more active than fi ee rhXPC, indicating that complex assembly is required for the stimulation. rhHR23B was shown to stimulate an early stage of NER at or prior to incision. Furthermore, both rhHR23 proteins function in a defined NER system reconstituted with purified proteins, indicating direct involvement of hHR23 proteins in the DNA repair reaction via interaction with XPC.AMER SOC MICROBIOLOGY, Dec. 1997, MOLECULAR AND CELLULAR BIOLOGY, 17(12) (12), 6924 - 6931, English[Refereed]Scientific journal
- PORTLAND PRESS, Feb. 1997, BIOCHEMICAL SOCIETY TRANSACTIONS, 25(1) (1), 309 - 315, EnglishMammalian nucleotide excision repair and syndromes[Refereed]Scientific journal
- HHR23B, a human Rad23 homolog, stimulates XPC protein in nucleotide excision repair in vitroA protein complex which specifically complements defects of XP-C cell extracts in vitro was previously purified to near homogeneity from HeLa cells. The complex consists of two tightly associated proteins: the XPC gene product and HHR23B, one of two human homologs of the Saccharomyces cerevisiae repair gene product Rad23 (Masutani et al., EMBO J. 13:1831-1843, 1994). To elucidate the roles of these proteins in ''genome-overall'' repair, we expressed the XPC protein in a baculovirus system and purified it to near homogeneity. The recombinant human XPC (rhXPC) protein exhibited a high level of affinity for single-stranded DNA and corrected the repair defect in XP-C whole-cell extracts without extra addition of recombinant HHR23B (rHHR23B) protein. However, Western blot (immunoblot) experiments revealed that XP-C cell extracts contained excess endogenous HHR23B protein, which might be able to form a complex upon addition of the rhXPC protein. To investigate the role of HHR23B, we fractionated the XP-C cell extracts and constructed a reconstituted system in which neither endogenous XPC nor HHR23B proteins were present. In this assay system, rhXPC alone weakly corrected the repair defect, while significant enhancement of the correcting activity was observed upon coaddition of rHHR23B protein. Stimulation of XPC by HHR23B was found,vith simian virus 40 minichromosomes as well as with naked plasmid DNA and with UV- as well as N-acetoxy-2-acetylfluorene-induced DNA lesions, indicating a general role of HHR23B in XPC functioning in the genome-overall nucleotide excision repair subpathway.AMER SOC MICROBIOLOGY, Sep. 1996, MOLECULAR AND CELLULAR BIOLOGY, 16(9) (9), 4852 - 4861, English[Refereed]Scientific journal
- XPC and human homologs of RAD23: Intracellular localization and relationship to other nucleotide excision repair complexesThe xeroderma pigmentosum syndrome complementation group C (XP-C) is due to a defect in the global genome repair subpathway of nucleotide excision repair (NER), The XPC protein is complexed with HHR23B, one of the two human homologs of the yeast NER protein, RAD23 [Masutani at al. (1994) EMBO J. 8, 1831-1843], Using heparin chromatography, gel filtration and native gel electrophoresis we demonstrate that the majority of HHR23B is in a free, non-complexed form, and that a minor fraction is tightly associated with XPC, In contrast, we cannot detect any bound HHR23A, Thus the HHR23 proteins may have an additional function independent of XPC. The fractionation behaviour suggests that the non-bound forms of the HHR23 proteins are not necessary for the core of the NER reaction, Although both HHR23 proteins share a high level of overall homology, they migrate very differently on native gels, pointing to a difference in conformation, Gel filtration suggests the XPC-HHR23B heterodimer resides in a high MW complex, However, immunodepletion studies starting from repair-competent Manley extracts fail to reveal a stable association of a significant fraction of the HHR23 proteins or the XPC-HHR23B complex with the basal transcription/repair factor TFIIH, or with the ERCC1 repair complex, Consistent with a function in repair or DNA/chromatin metabolism, immunofluorescence studies show all XPC, HHR23B and (the free) HHR23A to reside in the nucleus.OXFORD UNIV PRESS UNITED KINGDOM, Jul. 1996, NUCLEIC ACIDS RESEARCH, 24(13) (13), 2551 - 2559, English[Refereed]Scientific journal
- Xeroderma pigmentosum (XP) is a human hereditary disease characterized by a defect in DNA repair after exposure to ultraviolet Light. Among the seven groups of XP, group A (XP-A) patients show the most severe deficiency in excision repair and a wide variety of cutaneous and neurological disorders. We have cloned homologs of the human XPA gene from chicken, Xenopus, and Drosophila, and sequence analysis revealed that these genes are highly conserved throughout evolution. Here, we report characterization of the Drosophila homolog of the human XPA gene (Dxpa). The Dxpa gene product shows DNA repair activities in an in vitro repair system, and Dxpa cDNA has been shown to complement a mutant allele of human XP-A cells by transfection. Polytene chromosome in situ hybridization mapped Dxpa to 3F6-8 on the X chromosome, where no mutant defective in excision repair was reported. Northern blot analysis showed that the gene is continuously expressed in all stages of fly development. Interestingly, the Dxpa protein is strongly expressed in the central nervous system and muscles as revealed by immunohistochemical analysis using anti-Dxpa antibodies, consistent with the results obtained in transgenic flies expressing a Dxpa-beta-galactosidase fusion gene driven by the Dxpa promoter.AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, Sep. 1995, JOURNAL OF BIOLOGICAL CHEMISTRY, 270(38) (38), 22452 - 22459, English[Refereed]Scientific journal
- DNA-REPAIR PROTEIN XPA BINDS REPLICATION PROTEIN-A (RPA)XPA is a zinc finger DNA-binding protein, which is missing or altered in group A xeroderma pigmentosum cells and known to be involved in the damage-recognition step of the nucleotide excision repair (NER) processes. Using the yeast two-hybrid system to search for proteins that interact with XPA, we obtained the 34-kDa subunit of replication protein A (RPA, also known as HSSB and RFA). RPA is a stable complex of three polypeptides of 70, 34, 11 kDa and has been shown to be essential in the early steps of NER as well as in replication and recombination. We also demonstrate here that the RPA complex associates with XPA. These results suggest that RPA may cooperate with XPA in the early steps of the NER processes.AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, Feb. 1995, JOURNAL OF BIOLOGICAL CHEMISTRY, 270(8) (8), 4152 - 4157, English[Refereed]Scientific journal
- CHROMOSOMAL LOCALIZATION OF 3 REPAIR GENES - THE XERODERMA-PIGMENTOSUM GROUP-C GENE AND 2 HUMAN HOMOLOGS OF YEAST RAD23The nucleotide excision repair (NER) disorder xeroderma pigmentosum (XP) is characterized by sun (UV) sensitivity, predisposition to skin cancer, and extensive genetic heterogeneity. Recently, we reported the cloning and analysis of three human NER genes, XPC, HHR23A, and HHR23B. The previously cloned XPC gene is involved in the common XP complementation group C, which is defective in excision repair of nontranscribed sequences in the genome. The XPC protein was found to be complexed with the product of HHR23B, one of the two human homologs of the Saccharomyces cerevisiae NER gene RAD23. Here we present the chromosomal localization by in situ hybridization using haptenized probes of all three genes. The HHR23A gene was assigned to chromosome 19p13.2. Interestingly, the HHR23B and XPC genes, the product of which forms a tight complex, were found to colocalize on band 3p25.1, Pulsed-field gel electrophoresis revealed that the HHR23B and XPC genes possibly share a MluI restriction fragment of about 625 kb. Potential involvement of the HHR23 genes ih human genetic disorders is discussed. (C) 1994 Academic Press, Inc.ACADEMIC PRESS INC JNL-COMP SUBSCRIPTIONS, Oct. 1994, GENOMICS, 23(3) (3), 651 - 658, English[Refereed]Scientific journal
- PURIFICATION AND CLONING OF A NUCLEOTIDE EXCISION-REPAIR COMPLEX INVOLVING THE XERODERMA-PIGMENTOSUM GROUP-C PROTEIN AND A HUMAN HOMOLOG OF YEAST RAD23Complementation group C of xeroderma pigmentosum (XP) represents one of the most common forms of this cancer-prone DNA repair syndrome. The primary defect is located in the subpathway of the nucleotide excision repair system, dealing with the removal of lesions from the non-transcribing sequences ('genome-overall' repair). Here we report the purification to homogeneity and subsequent cDNA cloning of a repair complex by in vitro complementation of the XP-C defect in a cell-free repair system containing UV-damaged SV40 minichromosomes. The complex has a high affinity for ssDNA and consists of two tightly associated proteins of 125 and 58 kDa. The 125 kDa subunit is an N-terminally extended version of previously reported XPCC gene product which is thought to represent the human homologue of the Saccharomyces cerevisiae repair gene RAD4. The 58 kDa species turned out to be a human homologue of yeast RAD23. Unexpectedly, a second human counterpart of RAD23 was identified. All RAD23 derivatives share a ubiquitin-like N-terminus. The nature of the XP-C defect implies that the complex exerts a unique function in the genome-overall repair pathway which is important for prevention of skin cancer.NATURE PUBLISHING GROUP, Apr. 1994, EMBO JOURNAL, 13(8) (8), 1831 - 1843, English[Refereed]Scientific journal
- CELL-FREE REPAIR OF UV-DAMAGED SIMIAN VIRUS-40 CHROMOSOMES IN HUMAN CELL-EXTRACTS .1. DEVELOPMENT OF A CELL-FREE SYSTEM DETECTING EXCISION REPAIR OF UV-IRRADIATED SV40 CHROMOSOMESSimian virus 40 minichromosomes were irradiated with ultraviolet light and used to study cell-free DNA repair reactions by soluble extracts of repair-proficient human cells. Damage-specific incorporation of labeled nucleotides was measured after linearization and electrophoresis of the viral DNA. Repair synthesis with irradiated chromosomes was dependent on the dose of ultraviolet light, but naked viral DNA irradiated at the same doses showed significantly higher template activity. The reactions absolutely required the presence of cell extract, but an extract from Chinese hamster cells was as active as those from human cells. The repair activity was apparently stimulated by the addition of unirradiated plasmid DNA, which did not affect the nucleosome density of the viral chromosomes. ATP and its regenerating system were required for the reaction, but total repair synthesis was little affected by exogenous addition of substrates for RNA synthesis and poly(ADP-ribosyl)ation. Most of the repaired viral chromosomal DNA was in the supercoiled, closed circular form, suggesting that excision repair reactions proceeded to the stage of ligation in the present cell-free system.AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, Apr. 1993, JOURNAL OF BIOLOGICAL CHEMISTRY, 268(12) (12), 9098 - 9104, English[Refereed]Scientific journal
- CELL-FREE REPAIR OF UV-DAMAGED SIMIAN VIRUS-40 CHROMOSOMES IN HUMAN CELL-EXTRACTS .2. DEFECTIVE-DNA REPAIR SYNTHESIS BY XERODERMA-PIGMENTOSUM CELL-EXTRACTSWe have constructed a cell-free DNA repair system with UV-irradiated SV40 minichromosomes, as described in the accompanying paper (Sugasawa, K, Masutani, C., and Hanaoka, F. (1993) J. Biol. Chem 268, 9098-9104). In this study, we examined DNA repair synthesis by cell extracts from seven xeroderma pigmentosum (XP) complementation groups, A through G. DNA repair synthesis by XP cell extracts was lower than that with repair-proficient human 293 cell extract and did not increase to the level with the latter on increase in the amount of cell extract or the incubation time. The defects of XP cell extracts were complemented by addition of extracts from cells of different complementation groups, indicating that defective proteins in XP-A through G cells are directly involved in DNA repair. Addition of T4 endonuclease V, which is reported to complement defects of XP cells, stimulated DNA repair synthesis by the 293 cell extract, and also complemented the defects of all XP cell extracts. The XPAC gene product was shown to be involved in DNA repair synthesis using anti-xpac serum and xpac protein produced in Escherichia coli. Anti-xpac serum inhibited DNA repair synthesis by the 293 cell extract and xpac protein reversed the inhibition. Furthermore, xpac protein complemented the defects of extracts of two lines of XP-A cells (XP20SSV and XP12ROSV) but had no effect on the reactions of extracts from cells of other complementation groups. These findings are consistent with previous results obtained in experiments with cells, indicating that our system is useful for analyzing the mechanisms of DNA excision repair in mammalian cells.AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, Apr. 1993, JOURNAL OF BIOLOGICAL CHEMISTRY, 268(12) (12), 9105 - 9109, English[Refereed]Scientific journal
- ABROGATION OF P53-MEDIATED TRANSACTIVATION BY SV40 LARGE T-ANTIGENp53 is known to bind specifically to the 44-bp human DNA sequence in an immunoprecipitation assay. We show here that the transcription of the reporter CAT gene linked with the herpesvirus thymidine kinase (tk) promoter containing the 44-base sequence is enhanced by mouse wild-type but not mutant-type p53 in F9 and p53-null Saos-2 cells. The p53-mediated transactivation was dramatically abrogated by introduction of SV40 large T antigen (SVLT) in Saos-2 cells in which p53 was clearly associated with SVLT. Furthermore, the p53-SVLT complex did not bind to the 44-base sequence at all. Thus, SVLT sequesters the transactivation function of the wild-type p53 by inhibiting the binding of p53 to the 44-base sequence. This is good evidence to show 'loss of functions' in the product of a tumor-suppressor oncogene by a dominant oncogene product at a molecular level.STOCKTON PRESS, Mar. 1993, ONCOGENE, 8(3) (3), 543 - 548, English[Refereed]Scientific journal
- COMPLEMENTATION BY A CLONED HUMAN UBIQUITIN-ACTIVATING ENZYME E1 OF THE S-PHASE-ARRESTED MOUSE FM3A CELL MUTANT WITH THERMOLABILE E1A temperature-sensitive growth mutant tsFS20 isolated from mouse FM3A cells was identified as a mutant with thermolabile ubiquitin-activating enzyme E1 by transfection with a full-length cDNA encoding the human E1 enzyme and cell-cell hybridization with an authentic E1 mutant ts85 previously isolated from FM3A cells. The resulting transformants produced thermoresistant E1 activity. Upon shift-up of temperature, asynchronously growing tsFS20 cells showed multiple points of cell-cycle arrest. At the nonpermissive temperature, tsFS20 cells that had been synchronized at the G1-S-phase progressed and accumulated in the mid-S-phase, as evidenced by the absence of G2-specific cdc2 kinase activity, while ts85 mutant cells, the widely used E1 mutant, reached the G2-phase and were arrested. Thus, the E1 mutation seemed to be involved in progression in the S-phase as well as in the G2-phase in the cell cycle. Degradation of short-lived abnormal proteins in tsFS20 cells was decreased to about 50% at the nonpermissive temperature, while the block was fully restored to the wild-type level in the transformant cells. Relevance of the unusually high incidence of the temperature-sensitive E1 mutation was discussed in terms of the E1 as a determinant of heat tolerance of cells.JAPAN SOC CELL BIOLOGY, Apr. 1992, CELL STRUCTURE AND FUNCTION, 17(2) (2), 113 - 122, English[Refereed]Scientific journal
- NONCONSERVATIVE SEGREGATION OF PARENTAL NUCLEOSOMES DURING SIMIAN VIRUS-40 CHROMOSOME-REPLICATION INVITROSimian virus 40 chromosomes can be replicated in vitro with the same set of purified proteins required for the replication of naked DNA containing the viral origin. With these reconstituted systems, the fate of parental histones during replication was examined in vitro. The assembly of nucleosomes on replicating chromosomes was hardly affected by the presence of simultaneously replicating naked DNA competitor, suggesting that replication forks can traverse nucleosomes without the displacement of histones. Moreover, we demonstrate that the nascent nucleosomes were distributed almost equally between the leading and lagging strands. This distributive mode of nucleosome segregation favors the propagation of parental chromatin structures to both daughter cells, which can maintain cellular functions dictated by these structures during cell proliferation.NATL ACAD PRESS, Feb. 1992, PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 89(3) (3), 1055 - 1059, English[Refereed]Scientific journal
- TOPOISOMERASE-II PLAYS AN ESSENTIAL ROLE AS A SWIVELASE IN THE LATE STAGE OF SV40 CHROMOSOME-REPLICATION INVITROThe effects of topoisomerases I and II on the replication of SV40 DNA were examined using an in vitro replication system of purified proteins that constitutes the monopolymerase system. In the presence of the two topoisomerases, two distinct nascent DNAs were formed. One product arising from the replication of the leading template strand was approximately half the size of the template DNA, whereas the other product derived from the lagging template strand consisted of short DNAs. These products were synthesized from both SV40 naked DNA and SV40 chromosomes. For the replication of SV40 naked DNA, either topoisomerase I or II maintained replication fork movement and supported complete leading strand synthesis. When SV40 chromosomes were replicated with the same proteins, reactions containing only topoisomerase I produced shorter leading strands. However, mature size DNA products accumulated in reactions supplemented with topoisomerase II, as well as in reactions containing only topoisomerase II. In the presence of crude extracts of HeLa cells, VP-16, a specific inhibitor of topoisomerase II, blocked elongation of the nascent DNA during the replication of SV40 chromosomes. These results indicate that topoisomerase II plays a crucial role as a swivelase in the late stage of SV40 chromosome replication in vitro.AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, Jan. 1992, JOURNAL OF BIOLOGICAL CHEMISTRY, 267(1) (1), 462 - 466, English[Refereed]Scientific journal
- REPLICATION OF THE SIMIAN VIRUS-40 CHROMOSOME WITH PURIFIED PROTEINSSV40 chromosomes prepared from infected CV-1 cells were replicated with the purified proteins of SV40 T antigen, HeLa DNA polymerase alpha-primase complex, single-stranded DNA-binding protein, and topoisomerases I and II, all of which have been shown to be essential for SV40 DNA replication in vitro. Replication started near the origin and proceeded bi-directionally. The maximum speed of replication fork movement was 200-300 nucleotides/min, which was similar to the rate of SV40 DNA replication with the same set of proteins. When replication products were digested with micrococcal nuclease, DNA fragments of 160-180 base pairs, which is the typical size of mononucleosomal DNA, were protected. This result indicates that replicated DNA was reconstructed into the nucleosome structure, complexed with parental histones.AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, Aug. 1991, JOURNAL OF BIOLOGICAL CHEMISTRY, 266(24) (24), 16141 - 16148, English[Refereed]Scientific journal
- AMER SOC MICROBIOLOGY, Oct. 1990, JOURNAL OF VIROLOGY, 64(10) (10), 4820 - 4829, EnglishASSEMBLY OF NASCENT DNA INTO NUCLEOSOME STRUCTURES IN SIMIAN-VIRUS 40 CHROMOSOMES BY HELA-CELL EXTRACT[Refereed]Scientific journal
- SPRINGER VERLAG, Oct. 1989, EUROPEAN JOURNAL OF BIOCHEMISTRY, 185(1) (1), 55 - 61, EnglishHETEROGENEOUS ASSEMBLY OF NASCENT CORE HISTONES TO FORM NUCLEOSOMAL HISTONE OCTAMERS IN MOUSE FM3A CELLS[Refereed]Scientific journal
- JAPANESE BIOCHEMICAL SOC, Dec. 1986, JOURNAL OF BIOCHEMISTRY, 100(6) (6), 1543 - 1550, EnglishCHARACTERIZATION OF THE 100,000 DALTON MATERIAL PRODUCED BY CHROMATIN CROSS-LINKING REACTION WITH DITHIO-BIS(SUCCINIMIDYL PROPIONATE)[Refereed]Scientific journal
- JAPAN SOC CELL BIOLOGY, Dec. 1985, CELL STRUCTURE AND FUNCTION, 10(4) (4), 373 - 382, EnglishRAPID PURIFICATION OF NUCLEOSOME ASSEMBLY PROTEIN (AP-I) AND PRODUCTION OF MONOCLONAL-ANTIBODIES AGAINST IT[Refereed]Scientific journal
- SPRINGER VERLAG, 1984, EUROPEAN JOURNAL OF BIOCHEMISTRY, 142(3) (3), 431 - 439, EnglishPURIFICATION AND INITIAL CHARACTERIZATION OF A PROTEIN WHICH FACILITATES ASSEMBLY OF NUCLEOSOME-LIKE STRUCTURE FROM MAMMALIAN-CELLS[Refereed]Scientific journal
- 2021, 日本環境変異原ゲノム学会大会プログラム・要旨集, 50thInvestigating the TDP1-mediated error-prone repair of a ribonucleotide embedded into the genome
- 2021, 日本環境変異原ゲノム学会大会プログラム・要旨集, 50thMolecular mechanism of excessive immune response caused by aberrant DNA repair
- 2020, 日本分子生物学会年会プログラム・要旨集(Web), 43rdTyrosyl-DNA phosphodiesterasesはDNA中のリボヌクレオチドを起因とする突然変異形成に関与する
- Springer (eds. Nishigori Chikako and Kaoru Sugasawa), Jan. 2019, DNA Repair Disorders, 1 - 23, EnglishMolecular mechanism of DNA damage recognition for global genomic nucleotide excision repair: A defense system against UV-induced skin cancer[Refereed]Introduction scientific journal
- This book focuses on the clinical aspects of DNA repair disorders. Nucleotide excision repair is an important pathway for humans, as it is involved in biologically fundamental functions. This work presents clinical features together with the pathogenesis of DNA repair disorders such as Xertoderma Pigmentosum (XP). Studies on animal models are included as well. Clinical feature characteristics of each clinical subtype of XP are depicted according to the genotype, giving accurate and detailed information about the clinical features in terms of gene alterations, change of protein structure, and dysfunction in some of the repair pathways. This book is unique in that it provides detailed information on clinical features from more than 100 patients with XP-A, which is characterized by very severe manifestation of skin photosensitivity and neurological dysfunction. It will give readers important knowledge for understanding the concept and molecular mechanisms of DNA repair disorders. It also describes how to treat and care for patients with XP based on vast experience in clinical practice. DNA Repair Disorders will be a useful resource not only for physicians and basic scientists who are interested in and/or take care of patients with DNA repair disorders, but also dermatologists, neurologists, and researchers in the field of radiation biology and photobiology.Springer Singapore, 01 Jan. 2018, DNA Repair Disorders, 1 - 221, EnglishOthers
- 2018, 日本遺伝学会大会プログラム・予稿集, 90thリボヌクレオチドが誘発する奇異突然変異とその防御機構
- 2018, 日本環境変異原学会大会プログラム・要旨集, 47thDNA中のリボヌクレオチドが引き起こす特異な突然変異とその誘発機構の解析
- ELSEVIER SCIENCE INC, May 2017, JOURNAL OF INVESTIGATIVE DERMATOLOGY, 137(5) (5), S131 - S131, EnglishTruncated XPA protein could not interact with TFIIH but presented mild clinical manifestationsSummary international conference
- 2017, 日本生化学会大会(Web), 90thDNA中のリボヌクレオチドが引き起こす突然変異とその抑制機構
- 2017, 日本環境変異原学会大会プログラム・要旨集, 46thリボヌクレオチドが誘発する突然変異の抑制におけるDNA修復機構の役割
- 技術情報協会, Oct. 2016, Cosmetic stage, 11(1) (1), 9 - 15, Japanese紫外線によるDNAの損傷とがん化のメカニズム (特集 紫外線が原因で起こる皮膚トラブルと化粧品開発)
- For faithful DNA repair, it is crucial for cells to locate lesions precisely within the vast genome. In the mammalian global genomic nucleotide excision repair (NER) pathway, this difficult task is accomplished through multiple steps, in which the xeroderma pigmentosum group C (XPC) protein complex plays a central role. XPC senses the presence of oscillating ‘normal’ bases in the DNA duplex, and its binding properties contribute to the extremely broad substrate specificity of NER. Unlike XPC, which acts as a versatile sensor of DNA helical distortion, the UV-damaged DNA-binding protein (UV-DDB) is more specialized, recognizing UV-induced photolesions and facilitating recruitment of XPC. Recent single-molecule analyses and structural studies have advanced our understanding of how UV-DDB finds its targets, particularly in the context of chromatin. After XPC binds DNA, it is necessary to verify the presence of damage in order to avoid potentially deleterious incisions at damage-free sites. Accumulating evidence suggests that XPA and the helicase activity of transcription factor IIH (TFIIH) cooperate to verify abnormalities in DNA chemistry. This chapter reviews recent findings about the mechanisms underlying the efficiency, versatility, and accuracy of NER.Elsevier B.V., 01 Aug. 2016, DNA Repair, 44, 110 - 117, English[Refereed]Book review
- FEDERATION AMER SOC EXP BIOL, Apr. 2016, FASEB JOURNAL, 30, EnglishTripartite DNA Lesion Recognition and Verification by XPC, TFIIH, and XPA in Nucleotide Excision RepairSummary international conference
- FEDERATION AMER SOC EXP BIOL, Apr. 2016, FASEB JOURNAL, 30, EnglishTripartite DNA Lesion Recognition and Verification by XPC, TFIIH, and XPA in Nucleotide Excision RepairSummary international conference
- This book is a comprehensive review of the detailed molecular mechanisms of and functional crosstalk among the replication, recombination, and repair of DNA (collectively called the "3Rs") and the related processes, with special consciousness of their biological and clinical consequences. The 3Rs are fundamental molecular mechanisms for organisms to maintain and sometimes intentionally alter genetic information. DNA replication, recombination, and repair, individually, have been important subjects of molecular biology since its emergence, but we have recently become aware that the 3Rs are actually much more intimately related to one another than we used to realize. Furthermore, the 3R research fields have been growing even more interdisciplinary, with better understanding of molecular mechanisms underlying other important processes, such as chromosome structures and functions, cell cycle and checkpoints, transcriptional and epigenetic regulation, and so on. This book comprises 7 parts and 21 chapters: Part 1 (Chapters 1-3), DNA Replication Part 2 (Chapters 4-6), DNA Recombination Part 3 (Chapters 7-9), DNA Repair Part 4 (Chapters 10-13), Genome Instability and Mutagenesis Part 5 (Chapters 14-15), Chromosome Dynamics and Functions Part 6 (Chapters 16-18), Cell Cycle and Checkpoints Part 7 (Chapters 19-21), Interplay with Transcription and Epigenetic Regulation. This volume should attract the great interest of graduate students, postdoctoral fellows, and senior scientists in broad research fields of basic molecular biology, not only the core 3Rs, but also the various related fields (chromosome, cell cycle, transcription, epigenetics, and similar areas). Additionally, researchers in neurological sciences, developmental biology, immunology, evolutionary biology, and many other fields will find this book valuable.Springer Japan, 01 Jan. 2016, DNA Replication, Recombination, and Repair: Molecular Mechanisms and Pathology, 1 - 555, EnglishOthers
- Springer Japan, 01 Jan. 2016, DNA Replication, Recombination, and Repair: Molecular Mechanisms and Pathology, v - vi, EnglishOthers
- The intrinsic instability of genomic DNA renders it susceptible to damage by spontaneously produced by-products such as reactive oxygen species and aldehyde metabolites or environmental genotoxins such as radiation and chemicals. Unrepaired DNA damage interferes with essential DNA transactions, such as replication and transcription, and eventually gives rise to the alteration of genetic information. Because genomic instability can cause cell death or carcinogenesis, DNA damage must be recognized and repaired as quickly as possible. Several mechanisms of DNA repair that function according to the type of damage underlie a highly sensitive system capable of detecting a few sites of damage among the large amount of normal DNA in the genome. Nucleotide excision repair (NER) is a major DNA repair pathway that can eliminate a wide spectrum of damage. In mammals, NER is executed by two subpathways: global genome repair (GGR) and transcription-coupled repair. Both subpathways share common core NER factors but possess unique systems for recognizing DNA damage. Transcription-coupled repair is initiated by the stalling of RNA polymerase II and is responsible for the accelerated repair of DNA damage in the transcribed strand of active genes. By contrast, GGR is initiated by xeroderma pigmentosum group C (XPC) and/or the UV-damaged DNA-binding protein (UV-DDB) complex, followed by the sequential actions of other NER-associated factors. This chapter reviews the molecular mechanisms underlying the recognition and repair of DNA damage by mammalian GGR.Springer Japan, 01 Jan. 2016, DNA Replication, Recombination, and Repair: Molecular Mechanisms and Pathology, 155 - 174, English[Refereed][Invited]Introduction other
- GENETICS SOC JAPAN, Dec. 2015, GENES & GENETIC SYSTEMS, 90(6) (6), 397 - 397, EnglishStudies on the de-ubiquitination mechanism for the xeroderma pigmentosum group C protein in UV damage responseSummary international conference
- (公社)日本生化学会, Dec. 2015, 日本生化学会大会・日本分子生物学会年会合同大会講演要旨集, 88回・38回, [3P1218] - [3P1218], English酸化的DNA/RNA脱メチル化酵素ALKBH3の機能解析
- 日本癌学会, Oct. 2015, 日本癌学会総会記事, 74回, P - 3148, English酸化的DNA/RNA脱メチル化酵素ALKBH3の発現変化は細胞増殖や代謝に影響を与える
- ELSEVIER SCIENCE BV, Apr. 2015, DNA REPAIR, 28, 144 - 144, EnglishPost-translational modifications coordinating recognition and repair of UV-induced DNA damageSummary international conference
- 2015, 日本生化学会大会(Web), 88thヌクレオチド除去修復を制御するタンパク質リン酸化の解析
- 2015, 日本生化学会大会(Web), 88th塩基除去修復因子チミンDNAグリコシラーゼの構造活性相関と機能制御
- GENETICS SOC JAPAN, Dec. 2014, GENES & GENETIC SYSTEMS, 89(6) (6), 312 - 312, EnglishStudies on the de-ubiquitylation mechanism of the xeroderma pigmentosum group C proteinSummary international conference
- WILEY-BLACKWELL, Oct. 2014, JOURNAL OF DERMATOLOGY, 41, 14 - 14, EnglishGENOTYPE -PHENOTYPE CORRELATION AMONG XERODERMA PIGMENTOSUM COMPLEMENTATION GROUP DSummary international conference
- 31 Aug. 2014, 日本遺伝学会大会プログラム・予稿集, 86th, 99, Japanese紫外線損傷修復におけるXPCタンパク質の脱ユビキチン化機構の解析
- 2014, 日本分子生物学会年会プログラム・要旨集(Web), 37thヌクレオチド除去修復におけるDNA損傷認識機構の解析
- GENETICS SOC JAPAN, Dec. 2013, GENES & GENETIC SYSTEMS, 88(6) (6), 361 - 361, EnglishA novel function of FANCD2 protein in apoptosis signalingSummary international conference
- (公社)日本生化学会, Sep. 2013, 日本生化学会大会プログラム・講演要旨集, 86回, 2T14a - 07, Japanese紫外線損傷DNAを含むヌクレオソームの生化学的および構造生物学的解析
- GENETICS SOC JAPAN, Dec. 2012, GENES & GENETIC SYSTEMS, 87(6) (6), 386 - 386, EnglishStudies on the de-ubiquitylation mechanism of the xeroderma pigmentosum group C proteinSummary international conference
- The WD40-repeat protein DDB2 is essential for efficient recognition and subsequent removal of ultraviolet (UV)-induced DNA lesions by nucleotide excision repair (NER). However, how DDB2 promotes NER in chromatin is poorly understood. Here, we identify poly(ADP-ribose) polymerase 1 (PARP1) as a novel DDB2-associated factor. We demonstrate that DDB2 facilitated poly(ADP-ribosyl)ation of UV-damaged chromatin through the activity of PARP1, resulting in the recruitment of the chromatin-remodeling enzyme ALC1. Depletion of ALC1 rendered cells sensitive to UV and impaired repair of UV-induced DNA lesions. Additionally, DDB2 itself was targeted by poly(ADP-ribosyl)ation, resulting in increased protein stability and a prolonged chromatin retention time. Our in vitro and in vivo data support a model in which poly(ADP-ribosyl)ation of DDB2 suppresses DDB2 ubiquitylation and outline a molecular mechanism for PARP1-mediated regulation of NER through DDB2 stabilization and recruitment of the chromatin remodeler ALC1.ROCKEFELLER UNIV PRESS, Oct. 2012, JOURNAL OF CELL BIOLOGY, 199(2) (2), 235 - 249, English[Refereed]Introduction scientific journal
- (一社)日本放射線影響学会, Sep. 2012, 日本放射線影響学会大会講演要旨集, 55回, 126 - 126, Japanese紫外線によるDNA損傷の修復を促進するDDB2の構造と機能制御
- 31 Aug. 2012, 日本遺伝学会大会プログラム・予稿集, 84th, 100, JapaneseDNA損傷応答に関わる新規アセチル化標的タンパク質の同定
- 31 Aug. 2012, 日本遺伝学会大会プログラム・予稿集, 84th, 72, Japanese紫外線損傷修復におけるXPCタンパク質の脱ユビキチン化機構の解析
- 放射線生物研究会, Jun. 2012, 放射線生物研究, 47(2) (2), 95 - 111, JapaneseMolecular Basis of DNA Damage Recognition in Nucleotide Excision Repair
- 2012, 日本分子生物学会年会プログラム・要旨集(Web), 35th無細胞再構成系によるヌクレオチド除去修復活性制御因子の探索
- 2012, 日本分子生物学会年会プログラム・要旨集(Web), 35thゲノム安定維持に関わるcentrin-2の新たな機能
- 2012, 日本分子生物学会年会プログラム・要旨集(Web), 35th, 2W5II-1 (WEB ONLY), JapaneseDNA損傷によるヒトRad52タンパク質のアセチル化は,DNA二重鎖切断部位への集積に必要である
- 2012, 日本分子生物学会年会プログラム・要旨集(Web), 35th, 1P-0201 (WEB ONLY), Japaneseヒト間葉系幹細胞におけるヒトRad52タンパク質のDNA損傷によるアセチル化誘導
- The nucleotide excision repair (NER) system is a fundamental cellular stress response that uses only a handful of DNA binding factors, mutated in the cancer-prone syndrome xeroderma pigmentosum (XP), to detect an astounding diversity of bulky base lesions, including those induced by ultraviolet light, electrophilic chemicals, oxygen radicals and further genetic insults. Several of these XP proteins are characterized by a mediocre preference for damaged substrates over the native double helix but, intriguingly, none of them recognizes injured bases with sufficient selectivity to account for the very high precision of bulky lesion excision. Instead, substrate versatility as well as damage specificity and strand selectivity are achieved by a multistage quality control strategy whereby different subunits of the XP pathway, in succession, interrogate the DNA double helix for a distinct abnormality in its structural or dynamic parameters. Through this step-by-step filtering procedure, the XP proteins operate like a systematic decision making tool, generally known as decision tree analysis, to sort out rare damaged bases embedded in a vast excess of native DNA. The present review is focused on the mechanisms by which multiple XP subunits of the NER pathway contribute to the proposed decision tree analysis of DNA quality in eukaryotic cells. (C) 2011 Elsevier B.V. All rights reserved.ELSEVIER SCIENCE BV, Jul. 2011, DNA REPAIR, 10(7) (7), 673 - 683, EnglishBook review
- 2011, DNA repair, Vol. , No. , pp. 453-476, EnglishDNA damage recognition for mammalian global genome nucleotide excision repair[Refereed]Introduction scientific journal
- The nucleotide excision repair (NER) subpathway operating throughout the mammalian genome is a versatile DNA repair system that can remove a wide variety of helix-distorting base lesions. This system contributes to prevention of blockage of DNA replication by the lesions, thereby suppressing mutagenesis and carcinogenesis. Therefore, it is of fundamental significance to understand how the huge genome can be surveyed for occurrence of a small number of lesions. Recent studies have revealed that this difficult task seems to be accomplished through sequential actions of multiple DNA damage recognition factors, including UV-DDB, XPC, and TFIIH. Notably, these factors adopt completely different strategies to recognize DNA damage. XPC detects disruption and/or destabilization of the base pairing, which ensures a broad spectrum of substrate specificity for global genome NER. In contrast, UV-DDB directly recognizes particular types of lesions, such as UV-induced photoproducts, thereby vitally recruiting XPC as well as further extending the substrate specificity. After DNA binding by XPC, moreover, the helicase activity associated with TFIIH scans a DNA strand to make a final search for the presence of aberrant chemical modifications of DNA. The combination of these different strategies makes a crucial contribution to simultaneously achieving efficiency, accuracy, and versatility of the entire repair system.MAIK NAUKA/INTERPERIODICA/SPRINGER, Jan. 2011, BIOCHEMISTRY-MOSCOW, 76(1) (1), 16 - 23, English[Refereed]Book review
- (公社)日本生化学会, Dec. 2010, 日本生化学会大会・日本分子生物学会年会合同大会講演要旨集, 83回・33回, 1T12 - 9, EnglishDNA損傷認識蛋白質DDB2のN末端領域の機能解析(Functional analysis of the N-terminal domain of the DNA damage recognition protein DDB2)
- GENETICS SOC JAPAN, Feb. 2010, GENES & GENETIC SYSTEMS, 85(1) (1), 86 - 86, EnglishGenotoxic stress targets Fanconi anemia group D2 protein for fragmentation by the caspase-mediated pathwaySummary international conference
- UV-damaged DNA-binding protein (UV-DDB) is characterized by its very high affinity and specificity for UV-damaged DNA. Although precise roles for UV-DDB have been quite enigmatic since its discovery, accumulating evidence indicates that it promotes recognition of and protein assembly on UV photolesions in the global genome nucleotide excision repair pathway. The recently solved crystal structure of UV-DDB bound to DNA containing a (6-4) photoproduct has revealed that the DDB2/XPE subunit is responsible for the interaction, which induces flipping out of the two affected bases into a binding pocket, indicating that UV-DDB has evolved especially to recognize dinucleotide lesions, like UV photolesions. Taken together with the previously solved structure of the DDB1-CUL4A E3 ligase, this study has also novel insights into how this factor coordinates ubiquitination of various protein substrates around the site of DNA damage. (C) 2009 Elsevier B.V. All rights reserved.ELSEVIER SCIENCE BV, Aug. 2009, DNA REPAIR, 8(8) (8), 969 - 972, English[Refereed]Introduction scientific journal
- Aug. 2009, Mutat Res., 685(1-2):29-37, 29 - 37Regulation of damage recognition in mammalian global genomic nucleotide excision repair.[Refereed]Introduction scientific journal
- Although CUL4-containing ubiquitin ligases regulate DNA repair and DNA damage checkpoints, Liu et al. (2009) report in this issue of Molecular Cell that Cul4a-deficient mice exhibit surprising resistance to ultraviolet-induced skin tumors, providing insight into how ubiquitination regulates genome integrity.CELL PRESS, May 2009, MOLECULAR CELL, 34(4) (4), 403 - 404, English[Refereed]Others
- 医歯薬出版, Jan. 2009, 医学のあゆみ, 228: 137-142(2) (2), 137 - 142, Japanese色素性乾皮症遺伝子産物によるDNA損傷認識機構[Refereed]Introduction scientific journal
- In this review, we would like to introduce a unique approach for the estimation of radioadaptation. Recently, we proposed a new methodology for evaluating the repair efficiency of DNA double-strand breaks (DSB) using a model system. The model system can trace the fate of a single DSB, which is introduced within intron 4 of the TK gene on chromosome 17 in human lymphoblastoid TK6 cells by the expression of restriction enzyme I-SceI. This methodology was first applied to examine whether repair of the DSB (at the I-SceI site) can be influenced by low-dose, low-dose rate gamma-ray irradiation. We found that such low-dose IR exposure could enhance the activity of DSB repair through homologous recombination (HR). HR activity was also enhanced due to the pre-IR irradiation under the established conditions for radioadaptation (50 mGy X-ray-6 h-I-SceI treatment). Therefore, radioadaptation might account for the reduced frequency of homozygous loss of heterozygosity (LOH) events observed in our previous experiment (50 mGy X-ray-6 h-2 Gy X-ray). We suggest that the present evaluation of DSB repair using this I-SceI system, may contribute to our overall understanding of radioadaptation.2009, Journal of Radiation Research, 50(5) (5), 407 - 413, English[Refereed]Book review
- Xeroderma pigmentosum (XP) is an autosomal recessive disease, which is characterized by susceptibility to ultraviolet light (UV)-induced skin cancer. Among eight genes so far identified as responsible for XP, XPA through XPG are involved in nucleotide excision repair of DNA damage induced by UV as well as various chemical carcinogens. Since this repair system removes a major UV photoproduct, the cyclobutane pyrimidine dimer, quite slowly from the global genome, this lesion must be accurately bypassed during replication by DNA polymerase eta, encoded by the XPV gene. Recent studies have revealed that each of these XP genes possesses additional functions, some of which are concerned with other DNA repair pathways and/or cellular DNA damage responses. Such differential functions not only explain clinical heterogeneity among different genetic complementation groups but also have implications for the promotion of carcinogenic processes in XP patients.OXFORD UNIV PRESS, Mar. 2008, CARCINOGENESIS, 29(3) (3), 455 - 465, EnglishBook review
- Through its crucial role in DNA damage recognition and initiation of nucleotide excision repair in mammals, XPC protein represents a first line of defense against carcinogenesis. The recent elucidation of the molecular structure of the yeast XPC homolog Rad4 provides new insights into how eukaryotic cells recognize structurally diverse DNA lesions.NATURE PUBLISHING GROUP, Oct. 2007, NATURE STRUCTURAL & MOLECULAR BIOLOGY, 14(10) (10), 887 - 888, EnglishOthers
- Oct. 2007, Nature structural & molecular biology, 14(10) (10), 887 - 8, English, International magazineSensing of DNA damage by XPC/Rad4: one protein for many lesions.[Refereed]Introduction scientific journal
- 2007, 生化学, 3W23-3, Japaneseヌクレオチド除去修復における損傷認識とユビキチン化
- The DNA nucleotide excision repair (NER) system is our major defense against carcinogenesis. Defects in NER are associated with several human genetic disorders including xeroderma pigmentosum (XP), which is characterized by a marked predisposition to skin cancer. For initiation of the repair reaction at the genome-wide level, a complex containing one of the gene products involved in XP, the XPC protein, must bind to the damaged DNA site. The UV-damaged DNA-binding protein (UV-DDB), which is impaired in XP group E patients, has also been implicated in damage recognition in global genomic NER, but its precise functions and its relationship to the XPC complex have not been elucidated. However, the recent discovery of the association of UV-DDB with a cullin-based ubiquitin ligase has functionally linked the two damage recognition factors and shed light on novel mechanistic and regulatory aspects of global genomic NER. This article summarizes our current knowledge of the properties of the XPC complex and UV-DDB and discusses possible roles for ubiquitylation in the molecular mechanisms that underlie the efficient recognition and repair of DNA damage, particularly that induced by ultraviolet light irradiation, in preventing damage-induced mutagenesis as well as carcinogenesis.SPRINGER, Sep. 2006, JOURNAL OF MOLECULAR HISTOLOGY, 37(5-7) (5-7), 189 - 202, EnglishBook review
- 放射線生物研究会, 20 Dec. 2005, 放射線生物研究, 40(4) (4), 345 - 359, Japaneseクロマチン構造を介したヌクレオチド除去修復
- 日本環境変異原学会, 2005, 日本環境変異原学会大会プログラム・要旨集, (34) (34), 78 - 78, EnglishS2-2 Recognition and repair of DNA base lesions for protection against cancer(Symposium(2)"Challenge against carcinogenesis, a big Black Box")
- 25 Nov. 2004, 日本分子生物学会年会プログラム・講演要旨集, 27th, 637, JapaneseXPC複合体およびUV‐DDBの損傷DNA結合活性に対するヌクレオソーム構造の影響
- JAPAN SOC CELL BIOLOGY, May 2004, CELL STRUCTURE AND FUNCTION, 29, 63 - 63, EnglishFunctional analysis of centrin2 in XPC protein complexSummary international conference
- JAPAN SOC CELL BIOLOGY, May 2004, CELL STRUCTURE AND FUNCTION, 29, 11 - 11, EnglishUbiquitylation of XPC protein by DDB-E3 complexSummary international conference
- 25 Nov. 2003, 日本分子生物学会年会プログラム・講演要旨集, 26th, 497, Japaneseヌクレオソーム構造によるXPC蛋白質複合体の非特異的DNA結合の抑制はヌクレオチド除去修復の特異性を高める
- 06 Oct. 2003, 日本放射線影響学会大会講演要旨集, 46th, 126, JapaneseXPC‐HR23Bの非特異的DNA結合はヌクレオソーム構造によって抑制される
- 2003, NMR討論会講演要旨集, 42ndHR23Bユビキチン相同ドメイン-プロテアソームS5aサブユニットユビキチン結合モチーフ複合体の構造解析
- 2003, 日本分子生物学会年会プログラム・講演要旨集, 26thプロテアソームS5aサブユニットユビキチン結合モチーフとhHR23Bユビキチン相同ドメイン複合体の構造解析
- the Japan Radiation Research Society, 2003, Journal of radiation research, 44(4) (4), 405 - 405, English74 Nucleosome structure suppresses non-specific DNA binding of XPC-HR23B(Radiation sensitivities in cells & tissues, Abstracts of the 46th Annual Meeting of the Japan Radiation Research Society) :
- The XPC-HR23B complex is a prime candidate for the initial damage recognition step during global genome nucleotide excision repair. A specific interaction between the XPC-HR23B complex and various types of damaged DNA substrates has been demonstrated in recent work by electrophoretic mobility shift assays or immunoprecipitation. Although these studies allowed the estimation of relative binding affinities for the different types of lesions, the presence of large amounts of competitor DNA or the need for glutaraldehyde fixation prevented the quantification of equilibrium constants. We have performed a quantitative study on the binding of XPC to damaged DNA using fluorescence anisotropy measurements. The XPC-HR23B complex binds with high affinity (K(D) approximately 1-3 nM) to fluorescent 36 bp DNA fragments containing a single cisplatin 1,3-intrastrand adduct or a six-nucleotide mispaired region. From stoichiometric titration experiments, it is concluded that approximately 70% of the XPC-HR23B preparation is active in DNA binding. Binding experiments employing fluorescent probes with a single defined photoproduct reveal a 30-fold preference of XPC for 6,4-photoproducts as compared to a cyclobutane dimer. Competition experiments with undamaged and damaged plasmid DNA indicate that the XPC-HR23B complex discriminates between damaged and undamaged sites with high specificity. The specificity factor is between 100 and 3000, depending on the number of nonspecific sites considered in the calculations. Upon addition of XPA to the XPC binding reaction mixtures, it was not possible to detect cooperative ternary complex formation on the platinated 36 bp probe.28 May 2002, Biochemistry, 41(21) (21), 6583 - 7, English, International magazine[Refereed]
- the Japan Radiation Research Society, 2002, Journal of radiation research, 43(4) (4), 416 - 416, EnglishFunctions of the DNA Damage Recognition Factor for Mammalian Nucleotide Excision Repair :
- 31 Jul. 2001, Tissue culture research communications : the journal of experimental & applied cell culture research = 組織培養研究, 20(2) (2), 105 - 105, JapaneseC群色素性乾皮症(XPC)タンパク質の生細胞核内でのタンパク質間の相互作用と局在
- Oct. 2000, 宇宙生物科学 = Biological sciences in space, 14(3) (3), 264 - 265, JapaneseEffects of Gravity Change on Protein : protein Interaction
- 日本組織培養学会, 30 Jun. 2000, Tissue culture research communications : the journal of experimental & applied cell culture research = 組織培養研究, 19(2) (2), 68 - 68, JapaneseCell Cycle Regulation after DNA Damage in Nucleotide Excision Repair Deficient Cells
- 01 Dec. 1998, 日本分子生物学会年会プログラム・講演要旨集, 21, 371 - 371, JapaneseDamaged DNA binding activities of the XPC-hHR23B complex
- 01 Dec. 1998, 日本分子生物学会年会プログラム・講演要旨集, 21, 371 - 371, JapaneseFunction of the XPC protein complex in nucleotide excision repair
- 01 Dec. 1998, 日本分子生物学会年会プログラム・講演要旨集, 21, 370 - 370, JapaneseStudies on XPC functional domains
- 01 Dec. 1998, 日本分子生物学会年会プログラム・講演要旨集, 21, 371 - 371, JapaneseAnalysis of the interaction between hJR23A/B nad S5a
- Aug. 1996, 日本分子生物学会年会プログラム・講演要旨集, 19, 75 - 75, Japanese真核細胞ヌクレオチド除去修復の分子機構
- 01 Aug. 1996, 日本分子生物学会年会プログラム・講演要旨集, 19, 778 - 778, JapaneseXPC-HHR23B複合体の機能の解析
- 01 Aug. 1996, 日本分子生物学会年会プログラム・講演要旨集, 19, 778 - 778, JapaneseXPC/HHR23B蛋白質合体の相互作用
- ACADEMIC PRESS INC JNL-COMP SUBSCRIPTIONS, Jan. 1991, EXPERIMENTAL CELL RESEARCH, 192(1) (1), 110 - 117, English
- JAPAN SOC CELL BIOLOGY, Dec. 1987, CELL STRUCTURE AND FUNCTION, 12(6) (6), 649 - 649, EnglishCORRELATION BETWEEN CELL-PROLIFERATION AND REGULARITY OF NUCLEOSOME STRUCTURESummary international conference
- Joint work, 技術情報協会, Oct. 2016, Japanese紫外線によるDNAの損傷とがん化のメカニズムGeneral book
- Joint editor, Springer, 2016, EnglishDNA Replication, Recombination and Repair: Molecular Mechanisms and PathologyScholarly book
- Joint work, シーエムシー出版, Apr. 2015, Japanese光老化科学の最前線Scholarly book
- Joint work, InTech, Oct. 2007, EnglishDNA Repair (Kruman, I. ed.)Scholarly book
- Springer, English, ISBN: 9789811067211DNA Repair disorders
- 第42回染色体ワークショップ・第23回核ダイナミクス研究会, Jan. 2025, JapaneseDNA の構造的要因から探るヌクレオチド除去修復制御機構Poster presentation
- 日本環境変異原ゲノム学会第53回大会, Dec. 2024, Japanese脂肪族アルデヒドの代謝異常がゲノム安定性に及ぼす影響[Invited]Invited oral presentation
- 第47回日本分子生物学会年会, Nov. 2024, JapaneseCRL4Cdt2のDNA合成部位への適切な集積と解離が細胞周期進行を制御するPoster presentation
- 第47回日本分子生物学会年会, Nov. 2024, Japaneseクライオ電子顕微鏡によるクロマチン上の紫外線損傷修復メカニズムの解明Oral presentation
- 第47回日本分子生物学会年会, Nov. 2024, Japaneseヌクレオチド除去修復のDNA損傷認識制御におけるヒストン修飾の役割Poster presentation
- 第47回日本分子生物学会年会, Nov. 2024, Japanese酸化的RNA損傷を鋳型としたSARS-CoV-2由来RNA依存性RNAポリメラーゼのRNA合成機構の解明Poster presentation
- 第47回日本分子生物学会年会, Nov. 2024, Japaneseアルデヒド脱水素酵素ALDH3A2の欠損がゲノム安定性に及ぼす影響Poster presentation
- 第47回日本分子生物学会年会, Nov. 2024, Japaneseヌクレオチド除去修復のDNA損傷認識を制御するクロマチン構造変換機構Poster presentation
- The12th 3R+3C International Symposium, Nov. 2024, EnglishImpacts of heavy water on DNA double-strand break repairs and cellular transcription, potentially via quantum-level mechanisms underlying kinetic isotope effectsPoster presentation
- The12th 3R+3C International Symposium, Nov. 2024, EnglishThe involvement of chromatin remodeling factor SMACRAD1 in response to DNA double strand breaks[Invited]Poster presentation
- The12th 3R+3C International Symposium, Nov. 2024, EnglishImpact of histone modifications on damage recognition process of global genome nucleotide excision repairPoster presentation
- The12th 3R+3C International Symposium, Nov. 2024, EnglishCryo-EM analyses of UV-damaged recognition protein UV-DDB in nucleosomes during nucleotide excision repairPoster presentation
- The12th 3R+3C International Symposium, Nov. 2024, EnglishCooperation of Cdt2 C-terminal motifs in regulating CRL4Cdt2 dynamics at the DNA replication sitePoster presentation
- The12th 3R+3C International Symposium, Nov. 2024, EnglishContribution of translesion synthesis for mutagenesis via a novel food-induced formamidopyrimidine-derivativePoster presentation
- The12th 3R+3C International Symposium, Nov. 2024, EnglishEffects of DNA substrate structures on lesion excision by nucleotide excision repair in vitro[Invited]Poster presentation
- The12th 3R+3C International Symposium, Nov. 2024, EnglishThe agile dance of XP proteins in nucleotide excision repair[Invited]Invited oral presentation
- The12th 3R+3C International Symposium, Nov. 2024, EnglishChromatin dynamics regulating DNA damage recognition for nucleotide excision repair[Invited]Invited oral presentation
- 日本放射線影響学会第67回大会, Sep. 2024, Japaneseヌクレオチド除去修復の損傷認識制御におけるヒストン修飾の役割Oral presentation
- 日本放射線影響学会第67回大会, Sep. 2024, Japaneseヌクレオチド除去修復におけるDNA損傷認識の制御機構Oral presentation
- 日本放射線影響学会第67回大会, Sep. 2024, Japaneseクライオ電子顕微鏡による細胞内クロマチン上のDNA損傷修復機構の理解Oral presentation
- 日本放射線影響学会第67回大会, Sep. 2024, Japanese重水によるDNA二重鎖切断修復阻害は、加水分解酵素反応に対する量子レベルの速度論的同位体効果によると推測されるOral presentation
- 第83回日本癌学会学術総会, Sep. 2024, Japaneseアクリルアミドの活性代謝物であるグリシドアミドのホルムアミドピリミジン誘導体による突然変異誘発機序Oral presentation
- 第31回日本免疫毒性学会学術年会, Sep. 2024, JapaneseDNA修復の機能不全によるDNA鎖切断を伴わない自然免疫応答メカニズムの解明Poster presentation
- 第31回日本免疫毒性学会学術年会, Sep. 2024, JapaneseDNA鎖切断に起因するインターフェロン応答における核酸センサーcGAS,IFI16の役割Oral presentation
- Gutenberg Workshop RNase H 2024: Structures, Functions and Disorders, Sep. 2024, EnglishUncovering distinct DNA damage responses associated with RNase H2 deficiency and its pathogenic mutation in human TK6 cells
- 55th Annual Meeting of the EMGS, Sep. 2024, EnglishIn vitro RNA synthesis past oxidative base damage by SARS-CoV-2 RNA-dependent RNA polymeraseOral presentation
- 7th DNA Polymerases Meeting, Aug. 2024, EnglishContribution of translesion synthesis for mutagenesis via a novel food-induced formamidopyrimidine-derivativePoster presentation
- 第51回日本毒性学会学術年会, Jul. 2024, Japaneseグリシドアミド付加体のホルムアミドピリミジン誘導体はDNA複製阻害と突然変異を誘発するPoster presentation
- 日本薬学会年第144年会, Mar. 2024, Japaneseグリシドアミド付加体のホルムアミドピリミジン誘導体はDNA複製阻害と突然変異を誘発するPoster presentation
- 第41回染色体ワークショップ・第22回核ダイナミクス研究会, Jan. 2024, Japaneseヌクレオチド除去修復における損傷クロマチン認識機構の構造基盤と理解Poster presentation
- 第41回染色体ワークショップ・第22回核ダイナミクス研究会, Jan. 2024, Japanese損傷クロマチン基質を用いたヌクレオチド除去修復制御機構の生化学的解析Poster presentation
- 第41回染色体ワークショップ・第22回核ダイナミクス研究会, Jan. 2024, Japaneseヌクレオチド除去修復の損傷監視制御におけるヒストン修飾の役割Oral presentation
- 第46回日本分子生物学会年会, Dec. 2023, Japanese細胞の⽣存は量⼦効果に⽀配されているPoster presentation
- 第46回日本分子生物学会年会, Dec. 2023, Japaneseヌクレオチド除去修復のDNA損傷認識制御におけるヒストンH3K9メチル化修飾の役割Poster presentation
- 第46回日本分子生物学会年会, Dec. 2023, Japanese紫外線誘発DNA損傷に対する細胞応答におけるクロマチンリモデリング因⼦SMARCAD1の関与Poster presentation
- 第46回日本分子生物学会年会, Dec. 2023, JapaneseDNAポリメラーゼ‧イータの発現調節における脱ユビキチン化酵素USP11の関与Poster presentation
- 第46回日本分子生物学会年会, Dec. 2023, Japaneseゲノム全体を対象としたヌクレオチド除去修復の損傷認識を制御するクロマチン動態Oral presentation
- 第46回日本分子生物学会年会, Dec. 2023, Japanese⾷品汚染物質アクリルアミドの活性代謝物により⽣じるホルムアミドピリミジン誘導体の突然変異誘発機構Oral presentation
- 第46回日本分子生物学会年会, Dec. 2023, Japaneseヒストン翻訳後修飾を介したXPCタンパク質の核内局在制御Poster presentation
- 第46回日本分子生物学会年会, Dec. 2023, Japaneseヌクレオチド除去修復のDNA損傷認識を制御するクロマチン構造変換機構Poster presentation
- 日本環境変異原ゲノム学会第52回大会, Nov. 2023, JapaneseDNA損傷を起因とした過剰なインターフェロン応答の分子経路の同定Poster presentation
- 日本環境変異原ゲノム学会第52回大会, Nov. 2023, JapaneseゲノムDNAに蓄積したリボヌクレオチドが誘発するゲノム不安定化の分子機構Poster presentation
- 赤川真崇、Petr Grúz、菅澤 薫、浦 聖恵、佐々 彰, Nov. 2023, JapaneseSARS-CoV-2におけるRNA依存性RNAポリメラーゼの損傷乗り越えRNA合成機構の解析Poster presentation
- 日本環境変異原ゲノム学会第52回大会, Nov. 2023, Japanese脂肪族アルデヒド脱水素酵素の欠損により生じる細胞毒性の解析Poster presentation
- 日本環境変異原ゲノム学会第52回大会, Nov. 2023, Japanese脂肪族アルデヒド脱水素酵素のゲノム安定性維持における影響Oral presentation
- 日本放射線影響学会第66回大会, Nov. 2023, Japaneseクライオ電子顕微鏡による紫外線損傷修復機構の構造基盤の理解Oral presentation
- 日本放射線影響学会第66回大会, Nov. 2023, Japaneseヌクレオチド除去修復の正確性を保証するDNA損傷認識の分子構造基盤Oral presentation
- 日本放射線影響学会第66回大会, Nov. 2023, Japaneseゲノム全体を対象としたヌクレオチド除去修復の損傷認識制御におけるヒストン修飾の役割Oral presentation
- 日本遺伝学会第95回大会, Sep. 2023, Japanese損傷クロマチン基質を用いたヌクレオチド除去修復制御機構の生化学的解析Poster presentation
- 日本遺伝学会第95回大会, Sep. 2023, Japaneseヌクレオチド除去修復の正確性を保証するDNA損傷認識の分子構造基盤Oral presentation
- 第27回DNA複製・組換え・修復ワークショップ, Jun. 2023, Japaneseヌクレオチド除去修復の損傷認識制御におけるヒストン修復の役割Oral presentation
- 量子生命科学会第5回大会, May 2023, Japanese量子効果の想定外の大きな影響：細胞の生存は量子効果に支配されているPoster presentation
- The 19th Ataxia-Telangiectasia Workshop (ATW2023), Mar. 2023, EnglishHistone modifications regulating the initiation of global genome nucleotide excision repairPoster presentation
- 若手フロンティア研究会2022, Dec. 2022, Japanese損傷クロマチン基質におけるヌクレオチド除去修復開始制御機構の解析Poster presentation
- 第45回日本分子生物学会年会, Dec. 2022, Japaneseヒストン修飾を介したヌクレオチド除去修復の損傷認識促進機構Poster presentation
- 第45回日本分子生物学会年会, Dec. 2022, Japaneseヒストン修飾を介したヌクレオチド除去修復の損傷認識促進機構Oral presentation
- 第45回日本分子生物学会年会, Nov. 2022, Japanese脂質代謝制御におけるファンコニ貧血タンパク質の機能解析Poster presentation
- 第45回日本分子生物学会年会, Nov. 2022, Japaneseヌクレオチド除去修復におけるDNA損傷認識を制御するクロマチン構造変換因子複合体Poster presentation
- 第45回日本分子生物学会年会, Nov. 2022, Japanese脂質代謝制御におけるファンコニ貧血タンパク質の機能解析Oral presentation
- 日本環境変異原ゲノム学会第51回大会, Nov. 2022, JapaneseUnderstanding the Molecular Mechanism of Innate Immune Response Caused by DNA Repair DeficiencyPoster presentation
- 日本環境変異原ゲノム学会第51回大会, Nov. 2022, JapaneseγH2AXの多角的解析による内因性DNA二本鎖切断の定量評価Poster presentation
- 日本環境変異原ゲノム学会第51回大会, Nov. 2022, Japaneseゲノム中リボヌクレオチドの蓄積が誘発するDNA二本鎖切断の修復機構Poster presentation
- 第49回国際核酸化学シンポジウム, Nov. 2022, EnglishRing-Opened N7-deoxyguanosine adduct of glycidamide induces DNA replication inhibition and mutagenesisPoster presentation
- 2022 International IBS Conference for Genomic Integrity, Oct. 2022, EnglishRoles of histone modifications in DNA damage recognition initiating global genome nucleotide excision repairPoster presentation
- 2022 International IBS Conference for Genomic Integrity, Oct. 2022, EnglishChromatin dynamics regulating initiation of nucleotide excision repair[Invited]Invited oral presentation
- International Conference of the Genetics Society of Korea 2022 (ICGSK2022), Oct. 2022, EnglishChromatin dynamics regulating initiation of nucleotide excision repair[Invited]Invited oral presentation
- International Conference of the Genetics Society of Korea 2022 (ICGSK2022), Oct. 2022, EnglishRoles of histone modifications in DNA damage recognition initiating global genome nucleotide excision repairPoster presentation
- 日本放射線影響学会第65回大会, Sep. 2022, Japaneseヌクレオチド除去修復における紫外線誘発DNA損傷の効率的な認識に寄与するヒストン修飾Oral presentation
- 日本薬学会第142年会, Mar. 2022, Japanese損傷乗り越え型DNAポリメラーゼPolκとREV1はグリシドアミドN7位デオキシグアノシン付加体による点突然変異に寄与するPoster presentation
- 第39回染色体ワークショップ・第20回核ダイナミクス研究会, Dec. 2021, Japanese色素性乾皮症C群タンパク質によるDNA損傷認識を制御するクロマチン動態Poster presentation
- 第39回染色体ワークショップ・第20回核ダイナミクス研究会, Dec. 2021, Japaneseヒストン脱アセチル化はヌクレオチド除去修復におけるDNA損傷認識を制御するPoster presentation
- 第44回日本分子生物学会年会, Dec. 2021, Japaneseヒストン脱アセチル化はヌクレオチド除去修復におけるDNA損傷認識を制御するPoster presentation
- 第44回日本分子生物学会年会, Dec. 2021, Japanese脱ユビキチン化酵素による損傷乗り越え合成の制御機構の解析Poster presentation
- 第44回日本分子生物学会年会, Dec. 2021, Japaneseファンコニ貧血タンパク質の脂質代謝における機能解析Poster presentation
- 第44回日本分子生物学会年会, Dec. 2021, Japanese色素性乾皮症C群タンパク質によるDNA損傷認識を制御するクロマチン動態Poster presentation
- 第44回日本分子生物学会年会, Dec. 2021, JapaneseグリシドアミドN7位デオキシグアノシン付加体による点突然変異に寄与する損傷乗り越え型DNAポリメラーゼの解析Oral presentation
- 第44回日本分子生物学会年会, Dec. 2021, JapaneseDNA損傷修復において機能するCRL4ユビキチンリガーゼの損傷部位集積機構のライブイメージ解析Poster presentation
- 第94回日本生化学会大会, Nov. 2021, Japaneseヌクレオチド除去修復におけるDNA損傷認識を制御するクロマチンダイナミクスOral presentation
- 第26回DNA複製・組換え・修復ワークショップ, Oct. 2021, Japaneseゲノム安定性維持に関わるCRL4Cdt2ユビキチンリガーゼの損傷部位集積機構の解析Oral presentation
- 日本放射線影響学会第 64 回大会, Sep. 2021, Japanese紫外線誘発DNA損傷の効率的な認識と修復を保障するヒストン修飾の役割Oral presentation
- 日本遺伝学会第93回大会, Sep. 2021, Japaneseヌクレオチド除去修復の損傷認識を補助するクロマチン構造変換機構の解明Oral presentation
- 第1回広帯域極限電磁波生命理工連携研究会, Sep. 2021, Japanese光刺激を用いたゲノムDNA損傷応答の研究[Invited]Invited oral presentation
- 第43回日本分子生物学会年会, Dec. 2020, JapaneseA new regulatory mechanism of chromatin dynamics triggered by DNA damage recognition protein DDB2Poster presentation
- 第43回日本分子生物学会年会, Dec. 2020, JapaneseTyrosyl-DNA phosphodiesterasesはDNA中のリボヌクレオチドを起因とする突然変異形成に関与するPoster presentation
- 日本環境変異原学会第49回大会, Nov. 2020, Japanese脂質代謝におけるファンコニ貧血タンパク質の機能解析Oral presentation
- 日本遺伝学会第92回大会, Sep. 2020, Japanese色素性乾皮症タンパク質による紫外線誘発DNA損傷認識の生体内制御機構Oral presentation
- 第47回日本毒性学会学術年会, Jun. 2020, EnglishThe N7-glycidamide adduct of 2’-deoxyguanosine on the template strand induces DNA replication inhibition and mutagenesis in human cellsPoster presentation
- 4th DNA Repair/Replication Structures and Cancer Conference, EnglishFunctional analysis of histone modifications regulating recognition of DNA lesions in nucleotide excision repair.Poster presentation
- 第42回日本分子生物学会年会, Japaneseヌクレオソーム上における DNAスライドを介した新たな紫外線損傷認識メカニズムの解明Poster presentation
- 第42回日本分子生物学会年会, JapaneseN7-グリシドアミドdG付加体により誘発されるDNA複製阻害と変異原性の解析Poster presentation
- 第42回日本分子生物学会年会, JapaneseRNAを介したヌクレオチド除去修復制御機構の解析Poster presentation
- 第42回日本分子生物学会年会, Japanese脂質代謝に応答したファンコニ貧血タンパク質のダイナミクスPoster presentation
- 第42回日本分子生物学会年会, Japaneseファンコニ貧血タンパク質FANCD2と脂質代謝関連因子の機能的連関の解析Poster presentation
- 第42回日本分子生物学会年会, JapaneseDNAポリメラーゼ・イータと脱ユビキチン化酵素の相互作用の解析Poster presentation
- 第42回日本分子生物学会年会, JapaneseTLSポリメラーゼの損傷部位への導入過程を生細胞イメージングにより解 析する試みPoster presentation
- 第42回日本分子生物学会年会, JapaneseXPCタンパク質によるDNA損傷認識の制御とヒストン修飾の役割Poster presentation
- 第42回日本分子生物学会年会, Japaneseヌクレオチド除去修復の損傷認識を制御するクロマチン構造変換因子の探 索Poster presentation
- 金沢大学薬学シンポジウム2019「多様な生体防御システム研究のイノベーション〜アンメット・メディカル・ニーズから創薬へ〜」, Nov. 2019, Japaneseゲノムの安定維持を保証するDNA損傷認識の高次制御機構[Invited]Invited oral presentation
- 6th Asian Congress on Environmental Mutagens (ACEM) and 48th Annual Meeting of the Japanese Environmental Mutagen Society (JEMS), EnglishMechanistic insight of unique mutations caused by a ribonucleotide embedded into DNA.Poster presentation
- 6th Asian Congress on Environmental Mutagens (ACEM) and 48th Annual Meeting of the Japanese Environmental Mutagen Society (JEMS), EnglishAlternate processing pathways of a single ribonucleotide incorporated into DNA and its consequences in human cells.Oral presentation
- 6th Asian Congress on Environmental Mutagens (ACEM) and 48th Annual Meeting of the Japanese Environmental Mutagen Society (JEMS), EnglishChromatin dynamics regulating DNA lesion recognition in nucleotide excision repairOral presentation
- 日本放射線影響学会第62回大会, EnglishFunctional analysis of histone modifications regulating recognition of UV-induced DNA lesions in nucleotide excision repair.Oral presentation
- Symposium of the Biosignal Research Center and the KAITAKU Project "Genome Function in Higher-Order Life Phenomena", Kobe University), Nov. 2019, EnglishRoles for ATPase activities of transcription factor IIH in nucleotide excision repair.Oral presentation
- International Symposium on "Maintenance of Genome Integrity" (Joint Symposium of the Biosignal Research Center and the KAITAKU Project "Genome Function in Higher-Order Life Phenomena", Kobe University), Nov. 2019, EnglishFunctional analysis of chromatin remodeling machinery regulating recognition of DNA lesions in nucleotide excision repair.Oral presentation
- 第25回DNA複製・組換え・修復ワークショップ, Japaneseヌクレオチド除去修復の損傷認識を制御するヒストン修飾の解析Oral presentation
- サイエンスフロンティア研究発表会, Oct. 2019, JapaneseTLSポリメラーゼの生細胞イメージングとその定量解析への取り組みPoster presentation
- 9th Asia and Oceania Conference on Photobiology, EnglishMolecular mechanism of recognition and repair of UV-induced DNA damage.[Invited]Keynote oral presentation
- 2019 Fanconi Anemia Scientific Symposium, EnglishDynamics of FANCD2 in response to lipid metabolism.Poster presentation
- 17th International Congress on Photobiology and 18th Congress of the European Society for Photobiology, EnglishChromatin dynamics regulating recognition of UV-induced DNA photolesions.Oral presentation
- 第135回日本薬理学会近畿部会, Jun. 2019, Japanese皮膚老化メカニズムの解明とその応用Poster presentation
- International Symposium on XP and other Nucleotide Excision Repair Disorders, Mar. 2019, English, Cambridge, U.K., International conferenceIn vivo regulation of DNA damage recognition for nucleotide excision repair[Invited]Invited oral presentation
- 日本環境変異原学会第47回大会, Nov. 2018, Japanese, 京都, Domestic conference脂肪族アルデヒド代謝におけるファンコニ貧血タンパク質の機能解析Oral presentation
- 日本放射線影響学会第61回大会, Nov. 2018, Japanese, 長崎, Domestic conference紫外線誘発DNA損傷修復の高次制御機構Oral presentation
- 第41回日本分子生物学会年会, Nov. 2018, Japanese, 横浜, Domestic conferenceファンコニ貧血タンパク質FANCD2と脂質代謝関連因子の相互作用解析Poster presentation
- 第41回日本分子生物学会年会, Nov. 2018, Japanese, 横浜, Domestic conferenceXPCタンパク質によるDNA損傷認識の制御におけるヒストン修飾の役割Poster presentation
- The 11th 3R&3C Symposium, Nov. 2018, English, Kanazawa Japan, International conferenceTranslesion synthesis-associated chromatin remodeling factors contribute the function of human DNA polymerase η in cisplatin-treated cells.Oral presentation
- The 11th 3R&3C Symposium, Nov. 2018, English, Kanazawa Japan, International conferenceFunctional analysis of histone modification in DNA damage recognition process of nucleotide excision repair.Oral presentation
- 日本環境変異原学会第47回大会, Nov. 2018, Japanese, 京都, Domestic conferenceDNA中のリボヌクレオチドが引き起こす特異な突然変異とその誘発機構の 解析Poster presentation
- 第41回日本分子生物学会年会, Nov. 2018, Japanese, 横浜, Domestic conferenceDNAポリメラーゼ・イータと脱ユビキチン化酵素の相互作用の解析Poster presentation
- 日本遺伝学会第90回大会, Sep. 2018, Japanese, 奈良, Domestic conferenceリボヌクレオチドが誘発する奇異突然変異とその防御機構Oral presentation
- 第91回日本生化学会大会, Sep. 2018, Japanese, 京都, Domestic conferenceヌクレオチド除去修復の損傷認識を制御するクロマチン構造変換機構の解明Oral presentation
- 第91回日本生化学会大会, Sep. 2018, Japanese, 京都, Domestic conferenceヌクレオチド除去修復のDNA損傷認識を制御するクロマチン構造動態Oral presentation
- 第91回生化学会大会, Sep. 2018, Japanese, 京都, Domestic conferenceクロマチン構造を介したヌクレオチド除去修復の高次制御機構Oral presentation
- 日本遺伝学会第90回大会, Sep. 2018, Japanese, 奈良, Domestic conferenceFunctional impact of ubiquitin-proteasome system on UV-induced DNA damage response and repairOral presentation
- 日本薬学会第138年会, Mar. 2018, Japanese, 石川県立音楽堂他全9会場, Domestic conferenceヌクレオチド除去修復におけるDNA損傷認識の高次制御機構Oral presentation
- International Meeting on RECQ Helicases and Related Diseases 2018, Feb. 2018, English, かずさアカデミアパーク, International conferenceCoordinated DNA damage recognition by xeroderma pigmentosum gene products[Invited]Invited oral presentation
- 2017年度生命科学系学会合同年次大会, Dec. 2017, Japanese, 神戸国際会議場, Domestic conferenceヌクレオチド除去修復の開始を制御するクロマチン構造の役割Oral presentation
- 2017年度生命科学系学会合同年次大会, Dec. 2017, Japanese, 神戸国際会議場, Domestic conferenceヌクレオチド除去修復における損傷認識機構の解析Poster presentation
- 日本環境変異原学会第46回大会, Nov. 2017, Japanese, 一橋大学 一橋講堂, Domestic conference脂質アルデヒド代謝におけるファンコニ貧血タンパク質の機能解析Poster presentation
- 第24回DNA複製・組換え・修復ワークショップ, Nov. 2017, Japanese, 長良川国際会議場, Domestic conferenceユビキチン-プロテアソーム系による紫外線誘発DNA損傷応答制御Oral presentation
- 第24回DNA複製・組換え・修復ワークショップ, Nov. 2017, Japanese, 長良川国際会議場, Domestic conferenceヌクレオチド除去修復の開始を制御するクロマチン構造の役割Oral presentation
- 第24回DNA複製・組換え・修復ワークショップ, Nov. 2017, Japanese, 長良川国際会議場, Domestic conferenceヌクレオチド除去修復におけるDNA損傷認識機構の解析Oral presentation
- 第24回 DNA複製・組換え・修復ワークショップ, Nov. 2017, Japanese, 長良川国際会議場, Domestic conferenceシスプラチン誘発DNA鎖内架橋の乗り越え合成に関わるヒストン修飾とクロマチン構造変換因子の探索Poster presentation
- The 2nd Biosignal Research Center International Symposium, Nov. 2017, English, Kobe University, International conferenceMechanism and regulation of DNA damage recognition in nucleotide excision repairOral presentation
- ICEM-ACEM 2017, Nov. 2017, English, Songdo Convensia/Seoul South Korea, International conferenceChromatin dynamics regulating DNA damage recognition of nucleotide excision repair[Invited]Invited oral presentation
- 2017 FA Scientific Symposium, Sep. 2017, English, Grand Hyatt Atlanta in Buckhead/Atlanta USA, International conferenceFatty aldehyde dehydrogenase as a novel binding partner of FANCD2[Invited]Invited oral presentation
- 第76回 日本癌学会, Sep. 2017, Japanese, パシフィコ横浜, Domestic conferenceChromatin remodeling factors contribute to the function of human DNA polymerase η in the cells treated with cisplatinPoster presentation
- 6th US-Japan DNA Repair Meeting, May 2017, English, Berkeley University of California/ Berkeley, USA, International conferenceInteraction of DNA damage recognition factors with chromatin.[Invited]Invited oral presentation
- 第27回太陽紫外線防御研究委員会シンポジウム, Mar. 2017, Japanese, 太陽紫外線防御研究委員会, 大阪, Domestic conference紫外線DNA損傷修復の分子機構[Invited]Invited oral presentation
- Japan-Swiss Symposium on Chromatin Structure and Dynamics, Jan. 2017, English, Basel, Switzerland, International conferenceInteraction of DNA damage recognition factors with the chromatin structureOral presentation
- 第39回日本分子生物学会年会（MBSJ2016）, Nov. 2016, Japanese, 日本分子生物学会, 横浜, Domestic conferenceタンパク質分解系による紫外線誘発DNA損傷応答制御Poster presentation
- The 10th 3R Symposium, Nov. 2016, English, The 10th 3R Symposium Organizing Committee, Matsue, Shimane, Japan, International conferenceRoles of DNA topology and TFIIH ATPases in mammalian nucleotide excision repairOral presentation
- 第39回日本分子生物学会年会（MBSJ2016), Nov. 2016, Japanese, 日本分子生物学会, 横浜, Domestic conferenceMolecular mechanism ensuring accuracy of the nucleotide excision repair systemOral presentation
- The 10th 3R Symposium, Nov. 2016, English, The 10th 3R Symposium Organizing Committee, Matsue, Shimane, Japan, International conferenceFunctional impact of ubiquitin-proteasome system on UV-induced DNA damage response and repairPoster presentation
- 第39回日本分子生物学会年会（MBSJ2016）, Nov. 2016, Japanese, 日本分子生物学会, 横浜, Domestic conferenceFANCD2新規相互作用因子の探索と機能解析Oral presentation
- 第39回日本分子生物学会年会（MBSJ2016）, Nov. 2016, Japanese, 日本分子生物学会, 横浜, Domestic conferenceDNA損傷認識を制御するクロマチン構造動態Poster presentation
- 第39回日本分子生物学会年会（MBSJ2016）, Nov. 2016, Japanese, 日本分子生物学会, 横浜, Domestic conferenceDNAのトポロジー状態がヌクレオチド除去修復に及ぼす影響Poster presentation
- 第89回日本生化学会大会, Sep. 2016, Japanese, 日本分子生物学会, 仙台, Domestic conferenceヌクレオチド除去修復のDNA損傷認識とその制御の分子基盤Oral presentation
- Symposium: Celebration Jan Hoeijmakers, Sep. 2016, English, Rotterdam, The Netherlands, International conferenceStudies on nucleotide excision repair：beginning with XPC and beyond[Invited]Invited oral presentation
- KSBMB (Korean Society for Biochemistry and Molecular Biology) International Conference 2016, May 2016, English, KSBMB, Seoul, Korea, International conferenceDNA damage recognition mechanism for mammalian nucleotide excision repair[Invited]Invited oral presentation
- Conference on Responses to DNA Damage: from Molecule to Disease, Apr. 2016, English, Egmond aan Zee, The Netherlands, International conferenceDynamics of chromatin structure regulating nucleotide excision repairPoster presentation
- Conference on Responses to DNA Damage: from Molecule to Disease, Apr. 2016, English, Egmond aan Zee, The Netherlands, International conferenceDissection of the DNA damage recognition machinery in mammalian nucleotide excision repair[Invited]Invited oral presentation
- 第13回SUMO研究会, Jan. 2016, Japanese, SUMO研究会, 大阪, Domestic conference3R研究の今後の動向についてOral presentation
- 第13回SUMO研究会, Jan. 2016, Japanese, SUMO研究会, 大阪, Domestic conference3R研究の今後の動向についてOral presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会（BMB2015）, Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conference酸化的DNA/RNA脱メチル化酵素ALKBH3の機能解析Poster presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会(BMB2015), Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conference酸化的DNA/RNA脱メチル化酵素ALKBH3の機能解析Poster presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会（BMB2015）, Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conference塩基除去修復因子チミンDNAグリコシラーゼの構造活性相関と機能制御Poster presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会(BMB2015), Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conference塩基除去修復因子チミンDNAグリコシラーゼの構造活性相関と機能制御Poster presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会（BMB2015）, Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceヒトメディエーター複合体hMED17サブユニットによる転写制御とDNA損傷修復機能の解明Poster presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会(BMB2015), Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceヒトメディエーター複合体hMED17サブユニットによる転写制御とDNA損傷修復機能の解明Poster presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会（BMB2015）, Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceヒトRAD52のアセチル化を介した相同組換えにおけるアセチル化および脱アセチル化酵素の新規機能の解明Oral presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会(BMB2015), Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceヒトRAD52のアセチル化を介した相同組換えにおけるアセチル化および脱アセチル化酵素の新規機能の解明Oral presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会（BMB2015）, Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceヌクレオチド除去修復を制御するタンパク質リン酸化の解析Poster presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会(BMB2015), Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceヌクレオチド除去修復を制御するタンパク質リン酸化の解析Poster presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会（BMB2015）, Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceヌクレオチド除去修復を制御するクロマチン構造動態の解析Poster presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会(BMB2015), Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceヌクレオチド除去修復を制御するクロマチン構造動態の解析Poster presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会（BMB2015）, Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceヌクレオチド除去修復の細胞内制御機構とその多様性Oral presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会(BMB2015), Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceヌクレオチド除去修復の細胞内制御機構とその多様性Oral presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会（BMB2015）, Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceタンパク質分解系による紫外線損傷応答制御Oral presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会(BMB2015), Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceタンパク質分解系による紫外線損傷応答制御Oral presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会（BMB2015）, Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceゲノム損傷応答におけるp53とDDB2の機能連関Poster presentation
- 第38回日本分子生物学会年会・第88回日本生化学会大会合同大会(BMB2015), Dec. 2015, Japanese, 日本分子生物学会, 神戸, Domestic conferenceゲノム損傷応答におけるp53とDDB2の機能連関Poster presentation
- 第74回日本癌学会学術総会, Oct. 2015, Japanese, 日本癌学会, 名古屋, Domestic conference紫外線誘発DNA損傷の修復促進と皮膚癌の抑制に関わる分子機構Oral presentation
- 第74回日本癌学会学術総会, Oct. 2015, Japanese, 日本癌学会, 名古屋, Domestic conference紫外線誘発DNA損傷の修復促進と皮膚癌の抑制に関わる分子機構Oral presentation
- 第74回日本癌学会学術総会, Oct. 2015, Japanese, 日本癌学会, 名古屋, Domestic conference酸化的DNA/RNA脱メチル化酵素ALKBH3の発現変化は細胞増殖や代謝に影響を与えるPoster presentation
- 第74回日本癌学会学術総会, Oct. 2015, Japanese, 日本癌学会, 名古屋, Domestic conference酸化的DNA/RNA脱メチル化酵素ALKBH3の発現変化は細胞増殖や代謝に影響を与えるPoster presentation
- 第23回DNA複製・組換え・修復ワークショップ, Oct. 2015, Japanese, 焼津, Domestic conferenceユビキチンープロテアソーム系による紫外線誘発DNA損傷応答制御Oral presentation
- 第23回DNA複製・組換え・修復ワークショップ, Oct. 2015, Japanese, DNA複製・組換え・修復ワークショップ研究会, 焼津, Domestic conferenceユビキチンープロテアソーム系による紫外線誘発DNA損傷応答制御Oral presentation
- 日本遺伝学会第87回大会, Sep. 2015, Japanese, 日本遺伝学会, 仙台, Domestic conference紫外線損傷修復におけるXPCタンパク質の脱ユビキチン化制御機構の解析Oral presentation
- 日本遺伝学会第87回大会, Sep. 2015, Japanese, 日本遺伝学会, 仙台, Domestic conference紫外線損傷修復におけるXPCタンパク質の脱ユビキチン化制御機構の解析Oral presentation
- 15th International Congress of Radiation Research, May 2015, English, 京都, International conferenceMolecular mechanism of DNA damage recognition in mammalian nucleotide excision repair.Oral presentation
- 15th International Congress of Radiation Research, May 2015, English, ICRR, 京都, International conferenceMolecular mechanism of DNA damage recognition in mammalian nucleotide excision repair.Oral presentation
- 日本薬学会第135年会, Mar. 2015, Japanese, 神戸学院大学, Domestic conferenceヌクレオチド除去修復の制御に関わるクロマチン構造動態Oral presentation
- The Mammalian DNA Repair Gordon Research Conference 2015, Feb. 2015, English, Four Points Sheraton / Holiday Inn Express, International conferenceIn Vivo Regulation of Mammalian Nucleotide Excision Repair[Invited]Invited oral presentation
- 第37回日本分子生物学会年会, Nov. 2014, Japanese, パシフィコ横浜, Domestic conference哺乳類ヌクレオチド除去修復タンパク質XPCのDNA結合モードの1分子イメージングOral presentation
- 第37回日本分子生物学会年会, Nov. 2014, Japanese, パシフィコ横浜, Domestic conference翻訳後修飾を介した色素性乾皮症遺伝子産物の機能制御Oral presentation
- 第37回日本分子生物学会年会, Nov. 2014, Japanese, パシフィコ横浜, Domestic conference遺伝性疾患の分子病態：分子機能から高次生命機能へOral presentation
- 第37回日本分子生物学会年会, Nov. 2014, Japanese, パシフィコ横浜, Domestic conferenceヒトRAD52タンパク質のアセチル化制御Poster presentation
- 第37回日本分子生物学会年会, Nov. 2014, Japanese, パシフィコ横浜, Domestic conferenceヌクレオチド除去修復におけるDNA損傷認識機構の解析Poster presentation
- 第37回日本分子生物学会年会, Nov. 2014, Japanese, パシフィコ横浜, Domestic conferenceヌクレオソームにおける紫外線損傷塩基の収納およびその認識機構の解析Poster presentation
- 第37回日本分子生物学会年会, Nov. 2014, Japanese, パシフィコ横浜, Domestic conferenceゲノム損傷応答におけるGadd45αの機能解析Poster presentation
- The 9th 3R Symposium Together with National Instituite of Genetics 2014 International Symposium, Nov. 2014, English, 御殿場高原ホテル, International conferenceSUMO regulates DNA damage recognition in nucleotide excision repairPoster presentation
- The 9th 3R Symposium Together with National Instituite of Genetics 2014 International Symposium, Nov. 2014, English, 御殿場高原ホテル, International conferenceStudies on the de-ubiquitination mechanism of the xeroderma pigmentosum group C proteinPoster presentation
- Symposium Together with National Instituite of Genetics 2014 International Symposium, Nov. 2014, English, 御殿場高原ホテル, International conferencePost-translational modifications coordinating recognition and repair of UV-induced DNA damageOral presentation
- The 9th 3R Symposium Together with National Instituite of Genetics 2014, Nov. 2014, English, 御殿場高原ホテル, International conferenceFunctional regulation of thymine DNA glycosylase involved in DNA repair and epigenetic reprogrammingPoster presentation
- The 9th 3R Symposium 2014, Nov. 2014, English, 3R organizing committee, 御殿場, International conferenceFunctional interactions between XPC and DDB2 in nucleotide excision repairPoster presentation
- 第37回日本分子生物学会年会, Nov. 2014, Japanese, パシフィコ横浜, Domestic conferenceDNA損傷応答におけるDDB2の翻訳後修飾の機能解析Poster presentation
- 第37回日本分子生物学会年会, Nov. 2014, Japanese, パシフィコ横浜, Domestic conferenceDNA修復・損傷応答におけるクロマチン構造ダイナミクスの解析Poster presentation
- 第37回日本分子生物学会年会, Nov. 2014, Japanese, パシフィコ横浜, Domestic conferenceDNA修復とエピゲノム制御に関わるチミンDNAグリコシラーゼの機能制御Poster presentation
- 日本放射線影響学会第57回大会, Oct. 2014, Japanese, かごしま県民交流センター, Domestic conference紫外線誘発DNA損傷修復の細胞内制御機構Oral presentation
- 第87回日本生化学会大会, Oct. 2014, Japanese, 国立京都国際会館, Domestic conferenceヌクレオチド除去修復におけるXPCとDDB2の機能的相互作用Oral presentation
- 第5回日米修復会議, Oct. 2014, English, グランドエクシブ鳴門, International conferencePost-translational modifications coordinating recognition and repair of UV-induced DNA damage.Oral presentation
- 日本遺伝学会第86回大会, Sep. 2014, Japanese, 長浜バイオ大学, Domestic conference紫外線損傷修復におけるXPCタンパク質の脱ユビキチン化機構の解析Poster presentation
- 3rd Eastern Asia Dermatology Congress, Sep. 2014, English, Eastern Asia Dermatology Congress, Jeju, Korea, 韓国, International conferenceGenotype -Phenotype Correlation Among Xeroderma Pigmentosum Complementation Group D.Public symposium
- 第36回日本光医学・光生物学会, Jul. 2014, Japanese, 日本光医学・光生物学会, 大阪, Domestic conference細胞の紫外線損傷DNA修復能の蛍光検出Oral presentation
- International Symposium on Xeroderma Pigmentosum and Related Diseases, Mar. 2014, English, Kobe, Japan, International conferenceSUMO regulates DNA damage recognition in nucleotide excision repairPoster presentation
- International Symposium on Xeroderma Pigmentosum and Related Diseases, Mar. 2014, English, Kobe, Japan, International conferenceRoles for post-translational protein modifications in regulation of DNA damage recognition for mammalian nucleotide excision repairOral presentation
- International Symposium on Xeroderma Pigmentosum and Related Diseases, Mar. 2014, English, Kobe, Japan, International conferenceMolecular analysis of DNA repair defects in cells from Japanese patients with xeroderma pigmentosum Group DPoster presentation
- International Symposium on Xeroderma Pigmentosum and Related Diseases, Mar. 2014, English, Kobe, Japan, International conferenceFunctional interactions between XPC and DDB2 through ubiquitylation in DNA damage responsesPoster presentation
- International Symposium on Xeroderma Pigmentosum and related diseases, Mar. 2014, English, Kobe, International conferenceA child case of xeroderma pigmentosum group D.Poster presentation
- 第36回日本分子生物学会年会, Dec. 2013, Japanese, 日本分子生物学会, 神戸, Domestic conference哺乳類ヌクレオチド除去修復におけるDNA損傷認識課程の制御Oral presentation
- 第36回日本分子生物学会年会, Dec. 2013, Japanese, 神戸, Domestic conference哺乳類ヌクレオチド除去修復タンパク質XPCの１分子イメージング：DNA上での１次元自由拡散運動の観察Poster presentation
- 第36回日本分子生物学会年会, Dec. 2013, Japanese, 日本分子生物学会, 神戸, Domestic conference酸化的脱メチル化酵素ALKBH2/3の発現変化が細胞増殖・代謝に及ぼす影響Poster presentation
- 第36回日本分子生物学会年会, Dec. 2013, Japanese, 神戸, Domestic conferenceヌクレオチド除去修復の損傷認識に関わるクロマチン構造制御Poster presentation
- 第36回日本分子生物学会年会, Dec. 2013, Japanese, 神戸, Domestic conferenceFANCD2タンパク質はアポトーシスを抑制的に制御するOral presentation
- 第36回日本分子生物学会年会, Dec. 2013, Japanese, 神戸, Domestic conferenceDNA脱メチル化酵素TDGのSUMO化修飾による機能制御Poster presentation
- 第36回日本分子生物学会年会, Dec. 2013, Japanese, 神戸, Domestic conferenceDNA損傷応答におけるXPCとDDB2のユビキチン化を介した機能連関Poster presentation
- 厚生労働省科学研究費「神経皮膚症候群に関する調査研究班」平成25年度総会, Dec. 2013, Japanese, 神経皮膚症候群に関する調査研究班, 東京, Domestic conferenceDNA修復異常を伴う光線過敏症患者の分子細胞生物学的解析Others
- 第36回日本分子生物学会年会, Dec. 2013, Japanese, 神戸, Domestic conferenceDNA修復とエピゲノム制御に関わるチミンDNAグリコシラーゼの機能制御Poster presentation
- 第31回染色体ワークショップ・第12回核ダイナミクス研究会, Nov. 2013, Japanese, 箱根, Domestic conferenceファンコニ貧血症タンパク質FANCD2によるアポトーシス制御機構Oral presentation
- 第22回DNA複製・組換え・修復ワークショップ, Nov. 2013, Japanese, 仙台, Domestic conferenceファンコニ貧血症タンパク質FANCD2によるアポトーシス制御機構Poster presentation
- 第22回 DNA 複製・組換え・修復ワークショップ, Nov. 2013, Japanese, Domestic conferenceヌクレオチド除去修復における損傷認識機構とSUMO化修飾の意義Oral presentation
- International Symposium on Transcription and Metabolism, Nov. 2013, English, Awaji, Japan, International conferenceManipulated expression of the oxidative DNA/RNA demethylation enzymes ALKBH2/3 alters cellular proliferation and metabolism.Oral presentation
- International Symposium on Transcription and Metabolism, Nov. 2013, English, Awaji, Japan, International conferenceFunctional regulation of thymine DNA glycosylase involved in DNA repair and epigenetic reprogrammingOral presentation
- International Symposium on Transcription and Metabolism, Nov. 2013, English, Awaji, Japan, International conferenceFunctional interactions of DNA damage repair with transcriptional and epigenetic regulationOral presentation
- 第22回DNA複製・組換え・修復ワークショップ, Nov. 2013, Japanese, 仙台, Domestic conferenceDNA損傷応答におけるXPCとDDB2のユビキチン化を介した機能連関Oral presentation
- International Symposium on Transcription and Metabolism, Nov. 2013, English, Awaji, Japan, International conferenceChromatin structure regulating DNA damage recognition in nucleotide excision repairOral presentation
- 第51回日本生物物理学会年会, Oct. 2013, Japanese, 京都, Domestic conference哺乳類ヌクレオチド除去修復タンパク質XPCの1分子イメージングPoster presentation
- 日本放射線影響学会第56回大会, Oct. 2013, Japanese, 青森, Domestic conference紫外線誘発DNA損傷の修復機構におけるSUMO化修飾の役割Oral presentation
- 第36回日本分子生物学会年会, Oct. 2013, Japanese, 神戸, Domestic conference酸化的脱メチル化酵素ALKBH2/3の発現変化が細胞増殖・代謝に及ぼす影響Poster presentation
- 第1回がんと代謝研究会, Oct. 2013, Japanese, 鶴岡, Domestic conferenceメチル化塩基修飾酵素ALKBH2/3の発現変化が代謝に与える影響Poster presentation
- 第86回日本生化学会大会, Sep. 2013, Japanese, 横浜, Domestic conferenceゲノムDNA損傷の修復と細胞応答を制御するタンパク質翻訳後修飾の役割Oral presentation
- 第22回日本Cell Death学会学術集会, Jul. 2013, Japanese, 京都, Domestic conferenceFANCD2はDNA修復に関わる機能とは独立してTNFα/TNFR1を介したアポトーシス機構を負に制御しているOral presentation
- 第36回日本分子生物学会年会, 2013, English, 神戸, Domestic conferenceRegulation of the DNA damage recognition machinery in mammalian nucleotide excision repairOral presentation
- 日本遺伝学会第85回大会, 2013, Japanese, 横浜, Domestic conferenceFANCD2遺伝子産物はDNA修復とは独立したアポトーシス抑制機能を有するOral presentation
- 第35回日本分子生物学会年会, Dec. 2012, Japanese, 福岡, Domestic conferenceThe Functional Analysis of the Modifications of the DNA Damage Recognition Protein DDB2(2W5II-6)Keynote oral presentation
- 第35回 日本分子生物学会年会 ポスター発表, Dec. 2012, Japanese, Domestic conferenceRoles of sumoylation in regulation of the xeroderma pigmantosum group C proteinOral presentation
- 第35回日本分子生物学会年会, Dec. 2012, Japanese, 福岡, Domestic conferenceNovel function of RFC complexes in genome stability through PCNA-CRL4Cdt2 mediated (3W5III-5) Cdt1Degradation During DNA Replication.Keynote oral presentation
- 第30回染色体ワークショップ, Dec. 2012, Japanese, 淡路, Domestic conferenceFANCD2 タンパク質はアポトーシスを抑制的に制御するPublic symposium
- 第35回日本分子生物学会年会, Dec. 2012, Japanese, 福岡, Domestic conferenceExploration of Novel Regulatory Factors in Mammalian Nucleotide Excision Repair (1ST3-018)Poster presentation
- 第85回日本生化学会, Dec. 2012, Japanese, 福岡, Domestic conferenceElucidation of a Linkage Between Ubiquitin E3 Ligase RNF4 and a Component of the (1ST3-022) Thymine DNA Glycosylase TDG-containing DNA Demethylation Complex.Poster presentation
- 第35回日本分子生物学会年会, Dec. 2012, Japanese, 日本分子生物学会, 福岡, Domestic conferenceDNA damage induced-acetylation of human Rad52 protein is required for appropriate accumulation of the protein at DNA double-strand break sites.Keynote oral presentation
- 第35回日本分子生物学会年会, Dec. 2012, Japanese, 日本分子生物学会, 福岡, Domestic conferenceDNA damage-induced acetylation of human Rad52 protein in human mesenchymal stem cellsPoster presentation
- 第35回日本分子生物学会年会, Dec. 2012, Japanese, 日本分子生物学会, 福岡, Domestic conferenceA novel function of FANCD2 protein in apoptosis signaling (2ST2-026)Poster presentation
- 第35回日本分子生物学会, Dec. 2012, Japanese, 福岡, Domestic conferenceA novel function of centrin-2 involved in maintenance of genomic stability.Oral presentation
- 第35回日本分子生物学会年会, Dec. 2012, Japanese, 日本分子生物学会, 福岡, Domestic conferenceA novel function of centrin-2 involved in maintenance of genomic stabilityKeynote oral presentation
- The 8th 3R Symposium (International Symposium on DNA Replication, Recombination and Repair), Nov. 2012, English, Awaji, International conferenceSUMO regulates DNA damage recognition in nucleotide excision repair.Public symposium
- The 8th 3R Symposium (International Symposium on DNA Replication, Recombination and Repair), Nov. 2012, English, Awaji, International conferenceGenotoxic stress induces caspase-mediated cleavage of the FANCD2 protein.Public symposium
- The 8th 3R Symposium (International Symposium on DNA Replication, Recombination and Repair), Nov. 2012, English, Awaji, International conferenceFunctional studies on ubiquitylation of the DNA damage recognition protein DDB2Public symposium
- 第10回SUMO 研究会, Nov. 2012, Japanese, SUMO 研究会, 熊本, Domestic conferenceDNA損傷応答とSUMO／ubiquitinOral presentation
- The 8th 3R Symposium (International Symposium on DNA Replication, Recombination and Repair), Nov. 2012, English, Awaji, International conferenceDNA repair synthesis defect in xeroderma pigmentosum group D cells.Public symposium
- 日本遺伝学会第84回大会, Sep. 2012, Japanese, 福岡, Domestic conference紫外線損傷修復におけるXPCタンパク質の脱ユビキチン化機構の解析Oral presentation
- 日本放射線影響学会第55回大会, Sep. 2012, Japanese, 仙台, Domestic conference紫外線によるDNA損傷の修復を促進するDDB2の構造と機能制御Oral presentation
- 第71回日本癌学会学術総会, Sep. 2012, Japanese, 札幌, Domestic conferenceFunctional regulation of the DNA damage recognition factor DDB2 via post-translational modifications.Oral presentation
- 第34回日本光医学・光生物学会, Jul. 2012, Japanese, 神戸大学大学院医学研究科, 神戸, Domestic conference紫外線誘発DNA損傷の認識と修復の分子基盤[Invited]Invited oral presentation
- US-JAPAN DNA Repair Meeting, Apr. 2012, English, Virginia, USA, International conferencePost-Translational Protein Modification Regulating Mammalian Nucleotide Excision Repair.[Invited]Invited oral presentation
- DNA Repair Mini-Symposium, Apr. 2012, English, Maryland, USA, International conferenceMolecular mechanism of DNA damage recognition in mammalian nucleotide excision repair.[Invited]Invited oral presentation
- 国立遺伝学研究所研究集会「ユビキチン・SUMOによるDNA複製およびDNA修復系の制御」, Feb. 2012, Japanese, 国立遺伝学研究所, 三島, Domestic conferenceタンパク質翻訳後修飾によるDNA損傷認識の制御Oral presentation
- 第29回染色体ワークショップ, Jan. 2012, Japanese, 仙台, Domestic conferenceDNA損傷ストレスに伴うFANCD2タンパク質のカスパーゼ依存的切断Oral presentation
- Kobe University-University of Washington Joint Symposium on Integrative Membrane Biology and Signal Transduction Medeicine, Dec. 2011, English, 神戸, International conferenceRoles for sumoylation in regulation of the xeroderma pigmentosum group C proteinPoster presentation
- 第34回日本分子生物学会年会, Dec. 2011, Japanese, 日本分子生物学会, 横浜, Domestic conferenceRoles for sumoylation in regulation of the xeroderma pigmentosum group C proteinOral presentation
- 第34回日本分子生物学会年会, Dec. 2011, Japanese, 日本分子生物学会, 横浜, Domestic conferenceRegulation of active DNA demethylation enzyme, the thymine DNA glycosylase TDG, by SUMOylation and ubiquitinylationOral presentation
- 第34回日本分子生物学会年会, Dec. 2011, Japanese, 日本分子生物学会, 横浜, Domestic conferenceReconstitution of mammalian nucleotide excision repair:molecular basis for DNA damage recognitionOral presentation
- 第34回日本分子生物学会年会, Dec. 2011, Japanese, 日本分子生物学会, 横浜, Domestic conferenceNew function of PCNA loaders, RFC1-RFC and Ctf18-RFC, in genome stability through PCNA-CRL4-mediated Cdt1 degradationOral presentation
- 第34回日本分子生物学会年会, Dec. 2011, Japanese, 日本分子生物学会, 横浜, Domestic conferenceGenotoxic stress induces caspase-mediated cleavage of the FANCD2 proteinOral presentation
- 第34回日本分子生物学会年会, Dec. 2011, Japanese, 日本分子生物学会, 横浜, Domestic conferenceFunctional analysis of the CRL4 DDB2 E3 ligase in mammalian nucleotide excision repairOral presentation
- The 27th RBC-NIRS International Symposium on "Chromatin Dynamics and Epigenetic Memory in DNA Damage Response", Dec. 2011, English, Kyoto University, コープイン京都, International conferenceDNA damage recognition in nucleotide excision repair: molecular mechanism and involvement of chromatin structure.[Invited]Invited oral presentation
- 日本放射線影響学会第54回大会, Nov. 2011, Japanese, 日本放射線影響学会, 神戸, Domestic conferenceヌクレオチド除去修復におけるDNA損傷認識の分子基盤[Invited]Invited oral presentation
- Taiwan-Japan Joint Symposium on Cell Signaling and Gene Regulation, Nov. 2011, English, National Cheng Kung University, Tainan, Taiwan, International conferenceRegulationn of DNA repair through post-translational protein modifications.[Invited]Invited oral presentation
- 第12回Wakoつくばフォーラム, Nov. 2011, Japanese, 和光純薬工業, 筑波和光ホール, Domestic conferenceDNA損傷修復とエピゲノム制御のクロストーク[Invited]Invited oral presentation
- 第10回核ダイナミクス研究会, Oct. 2011, Japanese, 核ダイナミクス研究会, 北海道 北広島, Domestic conferenceヌクレオチド除去修復の損傷認識機構におけるSUMO化修飾の役割Oral presentation
- 平成23年度遺伝研研究会｢クロマチンダイナミクスの分子機構」, Oct. 2011, Japanese, 国立遺伝学研究所, 三島, Domestic conferenceヌクレオチド除去修復におけるDNA損傷認識機構とUV-DDBの役割[Invited]Invited oral presentation
- 第84回日本生化学会大会, Sep. 2011, Japanese, 日本生化学会, 国立京都国際会館, Domestic conferenceSUMO認識型ユビキチンリガーゼRNF4によるTDGグリコシラーゼを含むDNA脱メチル化酵素複合体の機能制御Oral presentation
- 構造エピゲノム研究会第４回ワークショップ「DNA損傷とエピゲノム」, Jul. 2011, Japanese, 構造エピゲノム研究会, 理化学研究所横浜研究所交流棟ホール, Domestic conferenceUV-DDB:ヌクレオチド除去修復とエピゲノム制御のクロスロード[Invited]Invited oral presentation
- Conference on Responses to DNA Damage: from Molecular Mechanism to Human Disease, Apr. 2011, English, Organizing Committee, Conference on Responses to DNA damage, Egmond aan Zee, The Netherlands, International conferenceRegulation of DNA damage recognition in mammalian NER[Invited]Invited oral presentation
- 第33回日本分子生物学会年会, Dec. 2010, Japanese, 日本分子生物学会, 神戸ポートアイランド, Domestic conference複製と損傷に応答して機能するユビキチンリガーゼ、CRL4Cdt2の制御機構Oral presentation
- 第33回日本分子生物学会年会・第83回日本生化学会大会合同大会, Dec. 2010, Japanese, 日本分子生物学会, 神戸ポートアイランド, Domestic conferenceFunctional analysis of the N-terminal domain of the DNA damage recognition protein DDB2Oral presentation
- 第33回日本分子生物学会年会・第83回日本生化学会大会合同大会, Dec. 2010, Japanese, 日本分子生物学会, 神戸ポートアイランド, Domestic conferenceDNA損傷ストレスに伴うFANCD2タンパク質のカスパーゼ依存的切断Oral presentation
- 第33回日本分子生物学会年会・第83回日本生化学会大会合同大会, Dec. 2010, Japanese, 日本分子生物学会, 神戸ポートアイランド, Domestic conferenceA 19S proteasome subunit PSMD14 is involved in deubiquitination of XPCPoster presentation
- The 7th 3R Symposium (International Symposium on DNA Replication, Recombination and Repair), Oct. 2010, English, 第7回3Rシンポジウム実行委員会, 富山国際会議場, Domestic conferenceRoles for centrin-2 in DNA damage recognition for global genome nucleotide excision repairOral presentation
- The 7th 3R Symposium (International Symposium on DNA Replication, Recombination and Repair), Oct. 2010, English, 第7回3Rシンポジウム実行委員会, 富山, Domestic conferenceGenotoxic stress induces caspase-mediated cleavage of the FANCD2 proteinPoster presentation
- 第69回日本癌学会学術総会, Sep. 2010, Japanese, 日本癌学会, 大阪, Domestic conferenceMolecular basis for DNA damage recognition by xeroderma pigmentosum gene productsOral presentation
- 国立遺伝学研究所研究集会「ユビキチン・SUMOによるDNA複製およびDNA修復系の制御」, Sep. 2010, Japanese, 国立遺伝学研究所, 三島, Domestic conferenceヌクレオチド除去修復におけるcentrin2の機能解析Oral presentation
- 第9回核ダイナミクス研究会, May 2010, Japanese, 核ダイナミクス研究会, 伊豆, Domestic conferenceゲノム全体を監視するDNA損傷認識の分子基盤Oral presentation
- International Conference on Radiation and Cancer Biology at Nagasaki 2010, Feb. 2010, English, Nagasaki University, Nagasaki, Japan, International conferenceDNA damage recognition mechanism for mammalian nucleotide excision repairInvited oral presentation
- 第32回日本分子生物学会年会, Dec. 2009, English, 日本分子生物学会, パシフィコ横浜, Domestic conferencePost-translational modification of XPC protein controls global genome nucleotide excision repairPoster presentation
- 第32回日本分子生物学会年会, Dec. 2009, Japanese, 日本分子生物学会, パシフィコ横浜, Domestic conferenceGenotoxic stress targets Fanconi anemia group D2 protein for fragmentation by the caspase-mediated pathwayPoster presentation
- 第32回日本分子生物学会年会, Dec. 2009, Japanese, 日本分子生物学会, パシフィコ横浜, Domestic conferenceFluorescence detection of cellular ability of base excision repairPoster presentation
- 第32回日本分子生物学会年会, Dec. 2009, Japanese, 日本分子生物学会, パシフィコ横浜, Domestic conferenceDNA damage recognition coordinated by the multiple xeroderma pigmentosum gene productsOral presentation
- 第32回日本分子生物学会年会, Dec. 2009, Japanese, 日本分子生物学会, パシフィコ横浜, Domestic conferenceCentrin 2 mediates interaction between XPC and DDB2Poster presentation
- 2009 Japan-Taiwan Joint Symposium on Cell Signaling and Gene Regulation, Nov. 2009, Japanese, Kobe University, National Cheng Kung University, Kobe, International conferencePost-translational modifications involved in cellular DNA damage responseOral presentation
- 第20回 DNA複製・組換え・修復ワークショップ, Nov. 2009, Japanese, 3Rワークショップ2009事務局, 琵琶湖コンファレンスセンター, Domestic conferenceCentrin 2 は損傷認識因子XPCとDDB2の相互作用を促進するOral presentation
- 国立遺伝学研究所研究集会「ユビキチン・SUMOによるDNA複製およびDNA修復系の制御」, Oct. 2009, Japanese, 国立遺伝学研究所, 三島, Domestic conferenceヌクレオチド除去修復に関わるXPCの翻訳後修飾による機能制御Oral presentation
- 日本遺伝学会第81回大会, Sep. 2009, Japanese, 日本遺伝学会, 松本, Domestic conferenceDNA損傷に伴うFANCD2(FA-D2)タンパク質のcaspase依存的切断Oral presentation
- National Conference on Medical Genomics and Proteomics, Sep. 2009, English, Novosibirsk, Russia, International conferenceCoordinated actions of multiple DNA damage detectors in nucleotide excision repair[Invited]Invited oral presentation
- The 3rd ASM Conference on DNA Repair and Mutagenesis, Jun. 2009, English, American Society for Microbiology, Whistler, BC, Canada, International conferencePost-translational modification of XPC protein controls global genome nucleotide excision repairPoster presentation
- International Symposium on DNA Damage Response and Repair Mechanisms, Apr. 2009, English, The Integrated Project DNA Damage and Response Mechanisms, Crete, Greece, International conferenceMolecular basis for DNA damage recognition in nucleotide excision repairInvited oral presentation
- 第3回放射線防護研究センターシンポジウム, Dec. 2008, Japanese, 放射線医学総合研究所, 千葉, Domestic conferenceヒトリンパ芽球細胞における放射線適応応答Invited oral presentation
- 第3回放射線防護研究センターシンポジウム, Dec. 2008, Japanese, 放射線医学総合研究所, 千葉, Domestic conferenceヌクレオチド除去修復におけるゲノム損傷認識機構Invited oral presentation
- BMB2008 第31回日本分子生物学会年会・第81回日本生化学会大会, Dec. 2008, Japanese, 日本分子生物学会・日本生化学会, 神戸, Domestic conferenceヌクレオチド除去修復におけるTFIHへリカーゼ活性調節機構の解析Poster presentation
- BMB2008 第31回日本分子生物学会年会・第81回日本生化学会大会, Dec. 2008, Japanese, 日本分子生物学会・日本生化学会, 神戸, Domestic conferenceヌクレオチド除去修復におけるDNA損傷認識の分子機序Oral presentation
- BMB2008 第31回日本分子生物学会年会・第81回日本生化学会大会, Dec. 2008, Japanese, 日本分子生物学会・日本生化学会, 神戸, Domestic conferenceCentrin 2 enhances in vitro nucleotide excision repair via complex formation with xeroderma pigmentosum group C pretein through its C-terminal domain.Poster presentation
- 日本放射線影響学会第51回大会, Nov. 2008, Japanese, 日本放射線影響学会, 北九州, Domestic conferenceDSB修復効率からの適応応答の追及Oral presentation
- New Nuclear Research Symposium "Biological Response to Low Dose Radiation", Nov. 2008, English, 新原子力基盤クロスオーバー研究「低線量域放射線に特有な生体反応の多面的解析」, Kitakyushu, International conferenceAnalysis of mutagenic radioadaptation in human lymphoblastoid cellsInvited oral presentation
- 第67回日本癌学会学術総会, Oct. 2008, Japanese, 日本癌学会, 名古屋, Domestic conferenceXeroderma pigmentosum proteins and their functions in DNA damage recognitionOral presentation
- The 6th 3R Symposium (International Symposium on DNA Replication, Recombination and Repair), Oct. 2008, English, 第6回3Rシンポジウム実行委員会, Kakegawa, Japan, International conferenceMolecular mechanisms underlying efficient DNA damage recognition for nucleotide excision repairOral presentation
- The 6th3R Symposium (International Symposium on DNA Replication, Recombination and Repair), Oct. 2008, English, 第6回3Rシンポジウム実行委員会, Kakegawa, International conferenceAnalyses of regulatory mechanisms of TFIIH helicase activity in nucleotide exicion repairPoster presentation
- Russian-European Workshop on DNA Repair and Epigenetic Regulation of Genome Stability, Jun. 2008, English, Institute of Chemical Biology and Fundermental Medicine, Russia St. Petersburg, International conferenceDamage recognition mechanism for mammalian nucleotide excision repairOral presentation
- 2007年度組換え・染色体再編ワークショップ・第19回DNA複製・分配ワークショップ・合同ワークショップ, Mar. 2008, Japanese, 伊豆, Domestic conferenceヌクレオチド除去修復因子の細胞内動態とその制御機構Oral presentation
- 第405回（総会）日本皮膚科学会大阪地方会, Feb. 2008, Japanese, 大阪, Domestic conference色素性乾皮症C群の小児例Oral presentation
- シンポジウム「DNA修復研究の最前線」．第30回日本分子生物学会年会・第80回日本生化学会大会 (BMB2007), Dec. 2007, Japanese, 日本分子生物学会・日本生化学会, 横浜, Domestic conferenceヌクレオチド除去修復因子の細胞内動態とその制御機構の解析Poster presentation
- ワークショップ「ユビキチン化を介したゲノムダイナミクスの制御」．第30回日本分子生物学会年会・第80回日本生化学会大会 (BMB2007), Dec. 2007, Japanese, 日本分子生物学会・日本生化学会, 横浜, Domestic conferenceヌクレオチド除去修復における損傷認識とユビキチン化Oral presentation
- 第30回日本分子生物学会年会・第80回日本生化学会大会 (BMB2007), Dec. 2007, Japanese, 日本分子生物学会・日本生化学会, 横浜, Domestic conferenceDNA修復因子と転写調節因子のSUMO化による活性調節の構造的基盤Oral presentation
- 第5回SUMO研究集会, Oct. 2007, Japanese, SUMO研究会, 京都, Domestic conferenceヌクレオチド除去修復におけるSUMO化の役割Oral presentation
- Symposium on "Genomic Instability" in The 66th Annual Meeting of the Japanese Cancer Association., Oct. 2007, English, 日本癌学会, Yokohama, Japan, International conferenceRepair of UV-induced DNA damage: the molecular mechanism preventing skin cancerOral presentation
- 国立遺伝学研究所研究集会「ユビキチン・SUMOによるDNA複製およびDNA修復系の制御」, Sep. 2007, Japanese, 三島, Domestic conferenceヌクレオチド除去修復に関わるユビキチン化とSUMO化[Invited]Invited oral presentation
- バイオアーキテクトシンポジウム, Sep. 2007, Japanese, 理化学研究所, 和光, Domestic conferenceゲノム損傷修復因子のin vivoダイナミクスとその制御機構Oral presentation
- 3rd Japan-US DNA Repair Meeting, May 2007, English, Akira Yasui, Fumio Hanaoka, Kiyoji Tanaka, Errol Friedberg, Samuel Wilson, Hotel Crescent, Akiu, Sendai, International conferenceMolecular Mechanisms promoting repair of UV-induced DNA damageInvited oral presentation
- International Conference Cellular Responses to DNA Damage and Technical Workshop on DNA Repair, Apr. 2007, English, Institute of Molecular Medicine, National Tsing-Hua University, Hsin-Chu, Taiwan, International conferenceMolecular mechanism of nucleotide excision repair as a defense against cancerInvited oral presentation
- THE JAPANESE ENVIRONMENTAL MUTAGEN SOCIETY
- THE JAPANESE RADIATION RESEARCH SOCIETY
- 日本遺伝学会
- 日本癌学会
- 日本細胞生物学会
- 日本薬学会
- 日本生化学会
- 日本分子生物学会
- Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B), Grant-in-Aid for Scientific Research (B), Kobe University, Apr. 2021 - Mar. 2025, Principal investigatorStudies on the principles of chromatin functions ensuring the repair of DNA damage induced by environmental stresses
- 科学研究費補助金／基盤研究(S), Apr. 2016 - Mar. 2021, Principal investigatorCompetitive research funding
- 科学研究費補助金／新学術領域研究, Apr. 2016 - Mar. 2018, Principal investigatorCompetitive research funding
- 国立研究開発法人日本医療研究開発機構,国立大学法人大阪大学, 創薬支援推進事業・創薬総合支援事業, 2017色素性乾皮症治療薬の探索Competitive research funding
- Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research on Innovative Areas (Research in a proposed research area), University of Tsukuba, 01 Apr. 2011 - 31 Mar. 2016Crosstalk of transcriptional control and energy pathways by hub metabolitesOur research project aimed to explore the crosstalk between the transcriptional status and energy metabolism. As the organizing group of this project, we held the annual meetings of the research groups and promoted the interaction and collaboration among the members, for example, by setting up “Collaboration Proposal Meeting”. We also supported the workshops organized by graduate students and young scientists, and “Transcription-Metabolism Seminars” held 58 times all over the country. Furthermore, we spread information on a website, held public lectures and announced our achievements to the people through mass media.
- Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), 01 Apr. 2012 - 31 Mar. 2015Single-molecule direct visualization of DNA repair proteins on chromatinTo elucidate how mammalian (human) DNA repair proteins recognize base lesions, we performed single-molecule direct visualization of the proteins on DNA. We found that XPC-RAD23B protein complex, which is known to be responsible for damage recognition in mammalian nucleotide excision repair, has high affinity for a lesion and performed one-dimensional bidirectional diffusion on undamaged DNA. The diffusion coefficients of the movement indicate that the protein complex does not always perform rotational tracking of the helical pitch of the DNA while moving along DNA and thus diffuses at much higher rates. The obtained results indicate that the protein complex uses these binding modes for efficient search of DNA lesions scattered throughout the genome from a vast excess of normal bases.
- Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (B), Osaka University, 01 Apr. 2012 - 31 Mar. 2015Detection of structural change of DNA using disulfide bond formation and its application to elucidation of molecular recognition mechanismsTo detect the DNA helix bending by a chemical approach, duplexes containing 2-O-mercaptoalkyl-β-D-arabinofuranose on both sides of the major groove were synthesized, and the effects of the cisplatin adduct, the abasic site analog, and the (6-4) photoproduct on DNA structures were investigated. A disulfide bond was formed depending on the cisplatin adduct, and dynamic DNA bending was shown for the abasic site and the (6-4) photoproduct. Using the bent structure obtained by this method, the model proposed for the handover of damaged DNA from the UV-DDB protein to the XPC protein in the nucleotide excision repair was validated.
- 科学研究費補助金／基盤研究(A), Apr. 2012 - Mar. 2015, Principal investigatorCompetitive research funding
- Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), Kumamoto University, 2011 - 2013Regulaion of epigenome by the SUMO and ubiquitin modification systemsThymine DNA glycosylase (TDG) is a base excision repair (BER) enzyme that interacts with the small ubiquitin-related modifier (SUMO)-targeted ubiquitin E3 ligase RNF4 and functions in the active DNA demethylation pathway. Here we found that the majority of TDG was SUMOylated in cells undergoing mitosis, while both SUMOylated and non-modified TDG were eliminated in cells arrested at S phase. In vitro and in vivo binding analyses revealed that TDG interacted with RNF4 in a SUMOylation-independent manner. In RNF4-depleted cells, both forms of TDG were efficiently degraded, suggesting that RNF4 was not the primary E3 ligase for controlling TDG stability. Our findings would not only be important for understanding posttranslational regulation of TDG but also provide implications for the TDG-RNF4 interaction in BER-driven active DNA demethylation during cell cycle progression.
- 科学研究費補助金／新学術領域研究, 2011, Principal investigatorCompetitive research funding
- 科学研究費補助金／新学術領域研究, 2011, Principal investigatorCompetitive research funding
- 2010, Principal investigator厚生科研「神経皮膚症候群に関する調査研究」Competitive research funding
- 2009, Principal investigator厚生科研「神経皮膚症候群に関する調査研究」Competitive research funding
- 2009, Principal investigatorヌクレオチド除去修復におけるＤＮＡ損傷認識初期過程の化学生物学的研究Competitive research funding
- 科学研究費補助金／特定領域研究, 2008, Principal investigatorCompetitive research funding
- 2008, Principal investigator厚生科研「神経皮膚症候群に関する研究」Competitive research funding
- 科学研究費補助金／基盤研究(A), 2008, Principal investigatorCompetitive research funding
- 2008, Principal investigatorヌクレオチド除去修復におけるＤＮＡ損傷認識初期過程の化学生物学的研究Competitive research funding
- 特別研究員奨励費, 2008, Principal investigatorCompetitive research funding
- 2007, Principal investigator戦略的：ゲノムの修復機構を基盤とした癌化・老化の制御Competitive research funding
- 科学研究費補助金／特定領域研究, 2007, Principal investigatorCompetitive research funding
- 科学研究費補助金／基盤研究(B), 2007, Principal investigatorCompetitive research funding
- 2007厚生科研：神経皮膚症候群に関する研究Competitive research funding
- 2000 - 2004, Principal investigatorDNA修復機構のネットワークと核構造Competitive research funding
- 2000 - 2004, Principal investigatorヌクレオチド除去修復による遺伝情報維持と細胞機能制御機構Competitive research funding
- Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), RIKEN (The Institute of Physical and Chemical Reseach), 2000 - 2001Studies on functions of mammalian RAD23 homologs that link DNA repair and protein degradation systemsTo examine the possibility of ubiquitylation of the XPC protein that is specifically induced upon DNA damage, FLAG-tagged XPC (FLAG-XPC) and HA-tagged ubiquitin were simultaneously overexpressed in XP4PASV cells, which are derived from a XP group C patient and do not express endogenous XPC protein. By immunoprecipitation of FLAG-XPC from the cell extract, it was demonstrated that XPC can be indeed ubiquitylated in vivo. From similar experiments using various truncated XPC proteins, a domain in XPC was found, which is relatively more susceptible to the ubiquitylation. To investigate possible roles of the XPC ubiquitylation, we established a cell line that stably expresses a physiological amount of FLAG-XPC. When the cells were treated with DNA damaging agents such as UV and 4-NQO, a fraction of XPC became more tightly bound to chromatin, while the majority of ubiquitylated XPC could be solubilized under the same conditions. Moreover, we found that the XPC ubiquitylation occurred in a cell-cycle-independent manner, at least from the Gl/S boundary to the M phase. To explore novel factors that interact with XPC, the yeast two-hybrid screening was carried out, which resulted in the ubiquitin-like protein SUM0-1 as well as the SUMO-1 conjugating enzyme Ubc9.As done for ubiquitylation, we demonstrated that XPC can be also sumoylated in cells by simultaneous overexpression of FLAG-XPC and HA-SUMO-1. Using mouse embryonic fibroblast cells that lack both HR23A and HR23B, we established cell lines that stably express various mutant HR23B proteins. It was shown that the N-terminal ubiquitin-like domain of HR23B is essential neither for intracellular stabilization of XPC nor removal of UV-damage from the global genome.
- 日本学術振興会, 科学研究費助成事業, 特定領域研究(C), 理化学研究所, 2000 - 2000突然変異に対する防御機構としてのDNA損傷認識機構の解明哺乳類のゲノム全体で働くヌクレオチド除去修復(NER)副経路において、DNA損傷認識因子として機能するXPC-HR23B複合体のDNAに対する結合特異性の詳細な解析を行なった。特定のDNA損傷や構造を部位特異的に含むDNA断片を用いてゲルシフト法やフットプリント法を行なうことにより、数個の非対合塩基(いわゆるバブル構造)があれば特に損傷が存在しなくてもXPC-HR23Bがこれを認識して結合できることが示された。すなわち、XPC-HR23BはDNA損傷そのものではなく、損傷によって誘起されるDNA構造の歪みを認識して結合すると考えられる。一方、これらのDNA基質を用いて無細胞NER反応を行なったところ、NER機構によるオリゴヌクレオチドの切り出しにはXPC-HR23Bの結合のみでは不十分で、バブル構造中に実際に損傷塩基が存在することが必要であった。以上の結果から、NERにおける損傷認識は(1)XPC-HR23BによるDNAの構造異常の認識、および(2)それに続く損傷塩基の確認、という少なくとも2段階の機構で行われていることが示された。さらに、XPC-HR23Bと相互作用する因子を探索する目的で、XPC蛋白質をbaitとして酵母2ハイブリッド法によるスクリーニングを行ない、複数のポジティブクローンを得た。現在、これらの因子とXPC-HR23Bとの相互作用の意義について、解析を進めている。さらに、XPCや別の損傷結合因子として知られるDDB因子のサブユニットにFLAGタグを融合したものを過剰発現する細胞株の単離を進めており、これを用いて相互作用する因子の解析を今後行なう予定である。
- 日本学術振興会, 科学研究費助成事業, 特定領域研究(A), 理化学研究所, 1999 - 1999ヌクレオチド除去修復機構におけるユビキチン蛋白分解系の役割RAD23ホモローグと26SプロテアソームのサブユニットS5a蛋白質との相互作用の意義を探る目的で、無細胞蛋白質分解反応系を用いた解析を行った。その結果、ユビキチン-プロテアソームを介した蛋白分解反応に対して、RAD23ホモローグが阻害的に作用することを見いだした。これは、RAD23ホモローグがS5aと結合することによって、ユビキチン化された分解基質蛋白質のプロテアソームとの相互作用を拮抗的に阻害した結果であると考えられた。即ち、RAD23ホモローグはプロテアソームによる蛋白質分解反応に対して、新しいタイプの制御因子として機能する可能性を持つことが示唆された。一方、HR23A、およびHR23Bそれぞれのノックアウトマウスを交配して二重欠損マウスの作成を試みた。二重欠損マウスは個体レベルでは致死的であると考えられたが、8.5日胚より二重欠損線維芽細胞を単離することに成功した。この二重欠損細胞はXPC欠損細胞と同等の紫外線感受性、および不定期DNA合成活性を示したが、紫外線照射後のRNA合成の回復能は正常であったことから、XPC変異細胞と同様に転写と共役したヌクレオチド除去修復機構は正常で、ゲノム全体で働く副経路に特異的な欠損を持つことがわかった。さらに、この二重欠損細胞ではXPC蛋白質の発現量が著しく低下していたことから、RAD23ホモローグがXPC蛋白質の安定な発現に重要な役割を果たしていることが示された。以上の結果から、RAD23ホモローグはゲノム全体で機能するヌクレオチド除去修復機構に必須であると同時に、おそらく蛋白質分解系の制御を介して個体の発生、分化、成長においても重要な機能を果たしていることが強く示唆された。
- 日本学術振興会, 科学研究費助成事業, 奨励研究(A), 理化学研究所, 1998 - 1999C群色素性乾皮症(XPC)蛋白質の構造と機能前年度までの組換え蛋白質を用いた無細胞系での実験から、C末端の125個のアミノ酸を欠失した変異XPC蛋白質は基本転写因子TFIIHとの相互作用に欠損が見られ、また無細胞ヌクレオチド除去修復反応においても不活性であることが明らかになった。そこで、この変異XPC蛋白質の細胞内での機能を解析する目的で、XP-C群細胞に変異遺伝子を導入して形質転換細胞株を樹立した。間接蛍光抗体染色により、上記のC末端欠失体、およびN末端の117個のアミノ酸を欠いた変異XPCはいずれも正常に核に移行できることを確認した。しかしながら、N末端欠失XPCを発現する細胞で紫外線抵抗性の回復が見られたのと対照的に、C末端欠失XPCを発現する細胞は親株のXPC欠損細胞と同等の紫外線感受性を示した。すなわち、無細胞系の結果と一致して、C末端部分はXPC蛋白質のヌクレオチド除去修復活性の発揮に必須であることが明らかになった。現在、C末端部分についてさらに細かい欠失変異体を作成しており、今後その構造機能相関を解析していく予定である。また、種々の変異XPC蛋白質を一過性に過剰発現して間接蛍光抗体染色を行なうことにより、XPC蛋白質の核移行に関わる配列をN末端付近に2ケ所同定した。 XPC蛋白質の活性が翻訳後修飾によって制御されている可能性を調べるため、まずXP-C群細胞にHA、またはFLAGタグを付加したXPC蛋白質のcDNAを導入して形質転換細胞株を単離した。これらの細胞株が正常細胞と同レベルのXPC蛋白質を発現していること、また紫外線抵抗性の回復からタグをつけたXPCが正常な機能を発揮できることを確認した。さらに、FLAGタグが特異的な免疫沈降に非常に有効であることを確かめたので、今後紫外線照射前後でのリン酸化の変動等を解析していく予定である。
- 日本学術振興会, 科学研究費助成事業, 特定領域研究(A), 理化学研究所, 1998 - 1998ヌクレオチド除去修復機構におけるユビキチン蛋白分解系の役割出芽酵母RAD23のヒト・ホモローグであるhHR23B蛋白質は、ヌクレオチド除去修復(NER)においてDNA損傷の認識に関わるXPC蛋白質と強固な複合体を形成する。我々は、酵母two-hybrid screeningにより、26Sプロテアソーム・サブユニットの一つ、S5aが.hHR23B、およびもう一つRAD23ホモローグであるhHR23Aと相互作用することを見出している。組換え蛋白質の欠失変異体を用いたbinding assayにより、hHR23BではN末端のユビキチン相同配列、S5aの側ではマルチュビキチン鎖結合部位として同定されたC末端に近い領域が、この相互作用に必要であることがわかった。さらに、細胞抽出液を用いたグリセロール密度勾配遠心、およびプロテアソーム・サブユニットに対する抗体を用いた免疫共沈降実験から、細胞内においてもhHR23Bの少なくとも一部はプロテアソームに結合していることが示唆された。一方、二つのRAD23ホモローグのそれぞれについて、ノックアウトマウスを作成レた。mHR23A欠損マウスは外見上まったく正常であったが、mHR23B欠損マウスはメンデル則からの期待値の1/10程度の頻度でしか得られず、また個体サイズも野生型と比較して明らかに小さい。しかしながら、このマウスから単離したmHR23B欠損細胞のNER能は野生型と同様であった。このことは、NERに関しては二つのRAD23ホモローグ間に機能的互換性があること、また823Bは個体の発生、成長過程において、NER獣外の何らかの重要な機能を果たしていることを示している。
- 日本学術振興会, 科学研究費助成事業, 重点領域研究, 1995 - 1995DNA複製、修復系を標的とした治療法の開発研究代表者と研究分担者は、DNA複製系及び修復系をターゲットとした新規抗がん剤の開発を目指して、以下に述べる研究を行なった。(1)前年度に引き続いて、既に確立されているSV40DNAの無細胞複製系、及び紫外線照射したSV40ミニ染色体を鋳型とする無細胞DNA修復系を用いて、100種ほどの放線菌培養上清から新たなDNA複製、及び修復阻害剤のスクリーニングを行なった。その結果、有意な阻害効果を示すサンプルを複数個、得ることが出来た。その中でDNA修復に阻害を示す4サンプルについて、DNAの損傷部位の上流および下流側に一本鎖切断を入れる反応を調べたところ、いずれもこの切断反応を阻害し、これらのサンプルがヌクレオチド除去修復反応の初期段階を阻害する活性を有していることが判明した。このステップの阻害剤は、これまで全く報告がなく、新規阻害剤の分離が期待される。(2)前年度までにDNA複製を強く阻害するものとして精製を行なっていたRK606については、構造決定の結果、大豆サポニンの一種、soyasaponin IIであることが判明した。類縁化合物のDNA複製阻害活性を比較したところ、アグリコン部分の他に、糖鎖の部分の構造も重要であること、また糖が3つ、分岐しない形で存在していると阻害活性が高いことが示された。DNA複製を阻害する化合物と、ワクシニアウイルスのdual-specific phosphatase,VH1フォスファターゼのヒトホモローグであるVHRフォスファターゼの阻害を示す化合物とが類似のスペクトルを示すことから、RK606の標的分子がフォスファターゼである可能性が示唆された。
- 日本学術振興会, 科学研究費助成事業, 奨励研究(A), 理化学研究所, 1994 - 1994無細胞系を用いたp53による染色体複製制御機構の研究バキュロウイルス・ベクター系を利用して昆虫細胞中で大量発現させたヒト野生型p53蛋白質をモノクローナルカラム抗体を用いて精製し、HeLa細胞の粗抽出液を用いたSV40DNAの無細胞複製系に添加して複製阻害を起こすことを確認した。この際、複製産物の解析から、p53蛋白質による複製阻害の作用点が主に複製開始段階にあることが示唆された。同様に4種類の変異型p53蛋白質(156Pro、248Trp、273His、285Lys)を昆虫細胞内で発現、精製したが、いずれも複製阻害活性を示さなかった。以上の結果は、精製蛋白質を用いた再構成複製系においても基本的に変わらなかった。モノクローナル抗体を用いた共沈降実験により、野生型p53蛋白質はSV40T抗原と試験管内で複合体を形成することが確かめられたが、4種類の変異型p53蛋白質は無細胞複製系の条件下では、いずれもSV40T抗原に結合しなかった。さらに、野生型p53蛋白質はSV40抗原の複製開始点への結合を阻害し、その結果としてT抗原による複製開始点を含む二重鎖DNA断片の一本鎖への巻き戻しを阻害するが、T抗原によって一本鎖環状M13DNA上の短いオリゴヌクレオチドが遊離される反応に対しては非常に弱い阻害効果しか示さないことがわかった。また、野生型p53蛋白質がSV40の複製開始点近傍に塩基配列特異的に結合することが報告されているが、この部分のDNA配列を欠失してもp53蛋白質による複製阻害が見られたことから、p53蛋白質の複製開始点近傍への結合は複製阻害にあまり重要でないと考えられた。以上の結果から、p53蛋白質によるSV40DNA複製阻害の機構は、主にSV40抗原との複合体形成によりT抗原の複製開始点への結合を阻害することにあると考えられる。
- 日本学術振興会, 科学研究費助成事業, 奨励研究(A), 理化学研究所, 1993 - 1993細胞同期における染色体複製の制御機構に関する研究SV40DNAの無細胞複製系に対する種々のプロテインキナーゼの影響を調べる目的で、まず精製された複製蛋白質に対して試験管内でリン酸化反応を行なってみた。その結果、MAPキナーゼやカゼインキナーゼIIがいくつかの複製蛋白質をリン酸化することが明らかとなった。しかしながら、これらのリン酸化反応によって各複製蛋白質の活性、および全体の複製反応に影響が見られるという結果は得られなかった。一方、ガン抑制遺伝子産物がG1期からS期への移行、染色体複製の開始を制御するという観点から、ヒトp53蛋白質の昆虫細胞における大量発現、ならびにモノクローナル抗体を用いた精製系を確立した。この系を用いて野生型ヒトp53蛋白質、およびヒトがん細胞から単離された数種の変異型p53蛋白質を精製し、SV40DNAの無細胞複製系に対する影響を解析した。その結果、野生型p53蛋白質SV40DNA複製を強く阻害したのに対して、変異型p53蛋白質はいずれも阻害を示さなかった。このSV40DNA複製の阻害活性と、それぞれのp53蛋白質のSV40T抗原との結合能の間には良い相関が見られ、実際に複製産物の解析から野生型p53による複製の阻害が開始段階の阻害であることが示唆された。現在、この組換えp53蛋白質を試験管内でリン酸化、および脱リン酸化した時に複製阻害活性に影響があるかどうか検討中である。
- 日本学術振興会, 科学研究費助成事業, 重点領域研究, 理化学研究所, 1993 - 1993哺乳類細胞のDNA修復機構と細胞周期制御の連関細胞周期におけるヌクレオチド除去修復活性の変動を解析する目的で、マウスFM3A細胞をnocodazoleでM期に同調後、解除して適当な時間培養したものをエルトリエーターで分画し、G1期の細胞を集めた。また、同様にFM3A細胞をaphidicolinでG1/S期境界に同調、及び解除することにより、エルトリエーター分画でS期とG2/M期の細胞を得た。これらの細胞より粗抽出液を調整し、無細胞修復系における活性を比較したところ、G2/M期の細胞抽出液がG1、及びS期の抽出液よりも2倍程度高い修復活性を示すことがわかった。さらに、マウスのcdc2キナーゼに変異を有するFM3A細胞の温度感受性突然変異株、tsFT210細胞を非許容温度でG2期に停止させたもの、及びこれをnocodazole存在下に許容温度に戻してM期に進めたものについて同様に修復活性を調べた。その結果、M期の細胞抽出液の方がG2期に停止した細胞抽出液よりも若干修復活性が高いという結果が得られた。現在、除去修復反応におけるcdc2キナーゼの関与、及び無細胞修復反応に伴う抽出液中のcdc2キナーゼの活性変動について検討中である。一方、哺乳類細胞のヌクレオチド除去修復自体に関与する因子を明らかにする目的で、C群色素性乾皮症の修復欠損を無細胞系で相補する蛋白質因子を同定し、その解析を進めているが、本年度においてはこの因子を構成する二種類のポリペプチドについてcDNAクローニングを行なった。
- Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for General Scientific Research (B), The Institute of Phisical and Chemical Research (RIKEN), 1992 - 1993Study on Structure and Function of Mammalian DNA Replication EntymeWe isolated cDNA of the catalytic subunit of DNA polymerase alpha from tsFT20 cells, a temperature-sensitive mutant cell line derived from mouse FM3A cells. DNA sequence analysis revealed that the cDNA has a single mutation, a cytosine to thymine substitution that changes amino acid 1180 from serine to phenylalanine. We have also shown that tsFT20 cells could be rescued by transfection with the wild-type cDNA.In addition, we detected mutation sites in one spontaneous and six N-methyl-N'-nitro-N-nitrosoguanidine-induced growth revertants of tsFT20 cells. All revertant cell lines had a second point mutation adjacent to the first mutation site in tsFT20 cells. During activation of quiescent Swiss mouse 3T3 cells to proliferate, the levels of mRNA of the four subunits of the DNA polymerase alpha-primase complex increased before DNA synthesis. In order to analyze how the expression of these genes are controlled, we have isolated upstream region of these genes and determined their nucleotide sequence. There are 1-2 E2F-binding sequence and 10-14 dp palindromic sequence, and one AP1-binding motif in each upstream region. We overexpressed cDNAs of the DNA polymerase alpha-primase complex in E.coli or insect cells, purified the gene products and raised the specific antibodies against them. Co-precipitation experiments showed that 54K subunit has affinity to all other three subunits. We have also shown that 180K subunit has DNA polymerase activity and 46K subunit has primase activity.
- Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for General Scientific Research (B), RIKEN (The Institute of Physical and Chemical Research), 1990 - 1991Involvement of Chromosome in Regulation of DNA Replication and RepairWe have developed a call-free system for replication of SV40 minichromosomes with purified proteins. Since this reconstituted system lacked activity of de novo nucleosome assembly, we could specifically examine the behavior of parental histones during chromosome replication. Solution hybridization analyses suggested that the histones were not completely detached from but somehow associated with template DNA. Using strand-specific probes and MNase digested nascent mononucleosomal DNA, we obtained the results indicating that the parental histone octamers were segregated distributively between leading and lagging strands during the chromosome replication with purified proteifis. When SV40 chromosomes were replicated with topoI alone as a swivelase, the reaction produced shorter leading strands but those of mature size were accumulated in the reaction supplemented with topo II. These results indicate that topo II has a crucial role as a swivelase in the late stage of SV40 chromosome replication in vitro. On the other hand, we have developed a cell-free system supporting repair reactions on DNA organized into hucleosomes, that is, SV40 chromosomes. Standard reaction mixtures contained UV-irradiated SV40 chromosomes, unirradiated plasmid DNA, whole cell extracts of HeLa cells(Manley's extracts), ATP-regenerating system and several low molecular weight materials including[alpha-^<32>P]dCTP. After incubated at 30ﾟC, DNAs were isolated, digested with EcoRI and separated by agarose gel electrophoresis. Repair synthesis as well as nick-translation of DNAs were detected by autoradiography. DNA synthesis on SV40 DNA was dependent on doses of irradiated DNA. When we used cell extracts made from XP-A or XP-C cells in place of HeLa cell extracts, the UV-dependent DNA synthesis decreased remarkably. Mixing of XP-A and XP- C cell extracts restored the DNA synthesis to similar level as HeLa cell extracts. Thus our in vitro system appears to be useful for reconstituting at least some features of chromosomal DNA excision repair.