SEARCH

Search Details

UEYAMA Takehiko
Biosignal Research Center
Professor

Researcher basic information

■ Research news
■ Research Keyword
  • 1103
  • 6905
  • 6904
  • 5805
■ Research Areas
  • Life sciences / Medical biochemistry
  • Life sciences / Neuroscience - general
  • Life sciences / Pharmacology
■ Committee History
  • Oct. 2019 - Mar. 2022, 文部科学省 科学技術・学術政策研究所 科学技術予測センター, 専門調査員
  • 日本薬理学会, 評議員

Research activity information

■ Award
  • Aug. 2005 広島大学医学部医学科広仁会, 広島大学医学部医学科広仁会 「平成17年度 基礎医学研究賞」, 新規NADPH oxidase (Nox4) の活性化機構と神経膠腫浸潤における役割の解明
    UEYAMA TAKEHIKO
    Others

■ Paper
  • Shunkou Kurasawa, Akira Ganaha, Shinya Ayabe, Atsushi Yoshiki, Fumiya Kawama, Shota Kitayama, Keiji Tabuchi, Kouhei Yamashita, Takehiko Ueyama
    DFNA1 (deafness, nonsyndromic autosomal dominant 1), initially identified as nonsyndromic sensorineural hearing loss, has been associated with an additional symptom: macrothrombocytopenia. However, the timing of the onset of hearing loss (HL) and thrombocytopenia has not been investigated, leaving it unclear which occurs earlier. Here, we generated a knock-in (KI) DFNA1 mouse model, diaphanous-related formin 1 (DIA1)KIΔv3/KIΔv3, in which Aequorea coerulescens green fluorescent protein (AcGFP)-tagged human DIA1(p.R1213X) was knocked into the ATG site of Dia1. Additionally, the exon 7 of Dia1 was deleted using genome editing to knock out (KO) Dia1-v3, a specific variant of Dia1. AcGFP-DIA1(p.R1213X) expression and endogenous DIA1 KO were confirmed in cochleae and platelets. Hearing function in DIA1KIΔv3/KIΔv3, but not DIA1KIΔv3/+ mice, evaluated by auditory brainstem response, was significantly worse at low frequencies compared to wild-type (WT) mice starting at 3 months of age (3M), with progressive deterioration. Using confocal microscopy and scanning electron microscopy, various stereociliary deformities were identified in the cochleae of DIA1KIΔv3/KIΔv3 mice. Platelet counts in DIA1KIΔv3/KIΔv3, but not DIA1KIΔv3/+ mice, were significantly lower than those in WT mice at 12M, but not at 6M. Furthermore, in a cohort of eight patients with DFNA1 harboring the p.R1213X mutation, HL preceded thrombocytopenia in three individuals. Thus, in both mice and humans, though HL and thrombocytopenia are progressive, HL manifests earlier than thrombocytopenia. Unlike myosin heavy chain 9 (MYH9)-related diseases, thrombocytopenia cannot be a predictive marker for HL in DFNA1. Nevertheless, monitoring platelet counts could provide insights into the progression of the hearing impairments in patients with DFNA1.
    Corresponding, Jan. 2025, FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 39(2) (2), e70309, English, International magazine
    [Refereed]
    Scientific journal

  • Naoko Adachi, Douglas T Hess, Takehiko Ueyama
    At least 10% of proteins constituting the human proteome are subject to S-acylation by a long-chain fatty acid, thioesterified to a Cys thiol side chain. Fatty S-acylation (prototypically, S-palmitoylation) operates across eukaryotic phylogeny and cell type. S-palmitoylation is carried out in mammalian cells by a family of 23-24 dedicated zDHHC palmitoyl transferase enzymes, and mutation of zDHHCs is associated with a number of human pathophysiologies. Activation of the zDHHCs by auto-S-palmitoylation, the transthioesterification of the active site Cys by fatty acyl-CoA, is the necessary first step in zDHHC-mediated protein S-palmitoylation. Most prior in vitro assessments of zDHHC activation have utilized purified zDHHCs, a time- and effort-intensive approach, which removes zDHHCs from their native membrane environment. We describe here a facile assay for zDHHC activation in native membranes. We overexpressed HA-tagged wild-type or mutant zDHHCs in cultured HEK293 cells and prepared a whole membrane fraction, which was incubated with fluorescent palmitoyl CoA (NBD-palmitoyl-CoA) followed by SDS-PAGE, fluorescence imaging and western blotting for HA. We show by mutational analysis that, as assayed, zDHHC auto-S-palmitoylation by NBD-palmitoyl-CoA is limited to the active site Cys. Application of the assay revealed differential effects on zDHHC activation of posttranslational zDHHC modification, and of zDHHC mutations associated with human disease, in particular cancer. Our assay provides a facile means of assessing zDHHC activation and thus of differentiating the effects of zDHHC mutation and post-translational modification on zDHHC activation versus secondary effects on zDHHC functionality resulting from altered zDHHC interaction with substrate palmitoyl-proteins.
    Corresponding, Jan. 2025, Journal of lipid research, 100743 - 100743, English, International magazine
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Takashi Sasayama, Takeshi Hamada, Kazuhiro Tanaka, Hiroaki Nagashima, Shunsuke Yamanishi, Takehiko Ueyama
    Abstract Glioblastoma is the most common malignant brain tumor in adults, the survival rate of which has not significantly improved over the past three decades. Therefore, there is an urgent need to develop novel treatment modalities. We previously reported that G1 to S phase transition 1 (GSPT1) depletion induces delayed cell cycle in primary astrocytes. Herein, we examined the potential of GSPT1 as a novel target for glioblastoma therapy. CC-885, a cereblon modulator that degrades GSPT1 by bridging GSPT1 to the CRL4 E3 ubiquitin ligase complex, was administered to nude mice with transplanted brain tumors of U87 glioblastoma cells. The survival period was significantly longer in CC-885 treated mice than in control mice. Furthermore, we generated GSPT1-knockout (KO) U87 cells and GSPT1-KO U87 cells with stable overexpression of FLAG-tagged GSPT1 (Rescued GSPT1-KO). Mice with transplanted GSPT1-KO U87 cells and Rescued GSPT1-KO U87 cells showed significantly longer and similar survival periods, respectively, as those with wild-type (WT) U87 cells. GSPT1-KO U87 cells showed enhanced apoptosis, detected by cleaved PARP1, compared to WT U87 cells. Brain tumors with transplantation of GSPT1-KO U87 cells also showed enhanced apoptosis compared to those with transplantation of WT and Rescued GSPT1-KO U87 cells. GSPT1 expression was confirmed in patients with glioblastoma. However, the clinical study using 87 glioblastoma samples showed that GSPT1 mRNA levels were not associated with overall survival. Taken together, we propose that GSPT1 is an essential protein for glioblastoma growth, but not its malignant characteristics, and that GSPT1 is a potential target for developing glioblastoma therapeutics.
    Corresponding, Springer Science and Business Media LLC, Aug. 2024, Cell Death & Disease, 15(8) (8)
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Takeshi Kaizuka, Takehiro Suzuki, Noriyuki Kishi, Kota Tamada, Manfred W. Kilimann, Takehiko Ueyama, Masahiko Watanabe, Tomomi Shimogori, Hideyuki Okano, Naoshi Dohmae, Toru Takumi
    Abstract Postsynaptic proteins play crucial roles in synaptic function and plasticity. During brain development, alterations in synaptic number, shape, and stability occur, known as synapse maturation. However, the postsynaptic protein composition changes during development are not fully understood. Here, we show the trajectory of the postsynaptic proteome in developing male mice and common marmosets. Proteomic analysis of mice at 2, 3, 6, and 12 weeks of age shows that proteins involved in synaptogenesis are differentially expressed during this period. Analysis of published transcriptome datasets shows that the changes in postsynaptic protein composition in the mouse brain after 2 weeks of age correlate with gene expression changes. Proteomic analysis of marmosets at 0, 2, 3, 6, and 24 months of age show that the changes in the marmoset brain can be categorized into two parts: the first 2 months and after that. The changes observed in the first 2 months are similar to those in the mouse brain between 2 and 12 weeks of age. The changes observed in marmoset after 2 months old include differential expression of synaptogenesis-related molecules, which hardly overlap with that in mice. Our results provide a comprehensive proteomic resource that underlies developmental synapse maturation in rodents and primates.
    Springer Science and Business Media LLC, Mar. 2024, Nature Communications, 15(1) (1)
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Shunkou Kurasawa, Hiroaki Mohri, Keiji Tabuchi, Takehiko Ueyama
    Considerable evidence of reactive oxygen species (ROS) involvement in cochlear hair cell (HC) loss, leading to acquired sensorineural hearing loss (SNHL), were reported. Cochlear synaptopathy between HCs and spiral ganglion neurons has been gathering attention as a cochlear HC loss precursor not detectable by normal auditory evaluation. However, the molecular mechanisms linking ROS with HC loss, as well as the relationship between ROS and cochlear synaptopathy have not been elucidated. Here, we examined these linkages using NOX4-TG mice, which constitutively produce ROS without stimulation. mRNA levels of Piccolo 1, a major component of the synaptic ribbon (a specialized structure surrounded by synaptic vesicles in HCs), were decreased in postnatal day 6 NOX4-TG mice cochleae compared to those in WT mice; they were also decreased by noise exposure in 2-week-old WT cochleae. As noise exposure induces ROS production, this suggests that the synaptic ribbon is a target of ROS. The level of CtBP2, another synaptic ribbon component, was significantly lower in NOX4-TG cochleae of 1-month-old and 4-month-old mice compared to that in WT mice, although no significant differences were noted at 1.5- and 2-months. The decrease in CtBP2 plateaued in 4-month-old NOX4-TG, while it gradually decreased from 1 to 6 months in WT mice. Furthermore, CtBP2 level in 2-month-old NOX4-TG mice decreased significantly after exposure to cisplatin and noise compared to that in WT mice. These findings suggest that ROS lead to developmental delays and early degeneration of synaptic ribbons, which could be potential targets for novel therapeutics for ROS-induced SNHL.
    Corresponding, Oct. 2023, Neurobiology of disease, 186, 106280 - 106280, English, International magazine
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Takashi Nakamura, Hirofumi Sakaguchi, Hiroaki Mohri, Yuzuru Ninoyu, Akihiro Goto, Taro Yamaguchi, Yoshitaka Hishikawa, Michiyuki Matsuda, Naoaki Saito, Takehiko Ueyama
    Abstract Rac small GTPases play important roles during embryonic development of the inner ear; however, little is known regarding their function in cochlear hair cells (HCs) after specification. Here, we revealed the localization and activation of Racs in cochlear HCs using GFP-tagged Rac plasmids and transgenic mice expressing a Rac1-fluorescence resonance energy transfer (FRET) biosensor. Furthermore, we employed Rac1-knockout (Rac1-KO, Atoh1-Cre;Rac1flox/flox) and Rac1 and Rac3 double KO (Rac1/Rac3-DKO, Atoh1-Cre;Rac1flox/flox;Rac3−/−) mice, under the control of the Atoh1 promoter. However, both Rac1-KO and Rac1/Rac3-DKO mice exhibited normal cochlear HC morphology at 13 weeks of age and normal hearing function at 24 weeks of age. No hearing vulnerability was observed in young adult (6-week-old) Rac1/Rac3-DKO mice even after intense noise exposure. Consistent with prior reports, the results from Atoh1-Cre;tdTomato mice confirmed that the Atoh1 promoter became functional only after embryonic day 14 when the sensory HC precursors exit the cell cycle. Taken together, these findings indicate that although Rac1 and Rac3 contribute to the early development of sensory epithelia in cochleae, as previously shown, they are dispensable for the maturation of cochlear HCs in the postmitotic state or for hearing maintenance following HC maturation. Key messages Mice with Rac1 and Rac3 deletion were generated after HC specification. Knockout mice exhibit normal cochlear hair cell morphology and hearing. Racs are dispensable for hair cells in the postmitotic state after specification. Racs are dispensable for hearing maintenance after HC maturation.
    Corresponding, Springer Science and Business Media LLC, May 2023, Journal of Molecular Medicine, 101, 843 - 854
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Yoko Niki, Naoko Adachi, Masaki Fukata, Yuko Fukata, Shinichiro Oku, Chieko Makino-Okamura, Seiji Takeuchi, Kazumasa Wakamatsu, Shosuke Ito, Lieve Declercq, Daniel B Yarosh, Tomas Mammone, Chikako Nishigori, Naoaki Saito, Takehiko Ueyama
    Palmitoylation is a lipid modification involving the attachment of palmitic acid to a cysteine residue, thereby affecting protein function. We investigated the effect of palmitoylation of tyrosinase, the rate-limiting enzyme in melanin synthesis, using a human three-dimensional skin model system and melanocyte culture. The palmitoylation inhibitor, 2-bromopalmitate, increased melanin content and tyrosinase protein levels in melanogenic cells by suppressing tyrosinase degradation. The palmitoylation site was Cysteine500 in the C-terminal cytoplasmic tail of tyrosinase. The nonpalmitoylatable mutant, tyrosinase (C500A), was slowly degraded and less ubiquitinated than wild-type tyrosinase. Screening for the Asp-His-His-Cys (DHHC) family of proteins for tyrosinase palmitoylation suggested that DHHC2, 3, 7, and 15 are involved in tyrosinase palmitoylation. Knockdown of DHHC2, 3, or 15 increased tyrosinase protein levels and melanin content. Determination of their subcellular localization in primary melanocytes revealed that DHHC2, 3, and 15 were localized in the endoplasmic reticulum, Golgi apparatus, and/or melanosomes, whereas only DHHC2 was localized in the melanosomes. Immunoprecipitation showed that DHHC2 and DHHC3 predominantly bind to mature and immature tyrosinase, respectively. Taken together, tyrosinase palmitoylation at Cysteine500 by DHHC2, 3, and/or 15, especially DHHC2 in trans-Golgi apparatus and melanosomes and DHHC3 in the endoplasmic reticulum and cis-Golgi apparatus, regulate melanogenesis by modulating tyrosinase protein levels.
    Corresponding, Sep. 2022, The Journal of investigative dermatology, 143(2) (2), 317 - 327, English, International magazine
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Bong Jik Kim, Takushi Miyoshi, Taimur Chaudhry, Thomas B Friedman, Byung Yoon Choi, Takehiko Ueyama
    Diaphanous-related formin 1 (DIAPH1) is a formin homology F-actin elongating protein encoded by DIAPH1. Homozygous recessive variants resulting in the loss of DIAPH1 function cause seizures, cortical blindness, and microcephaly syndrome (SCBMS), but hearing loss has not been reported. In contrast, dominant variants of human DIAPH1 are associated with DFNA1 non-syndromic sensorineural hearing loss. The deafness phenotype is due partly to abnormal F-actin elongation activity caused by disruption of the DIAPH1 autoinhibitory mechanism. We report an elderly female heterozygous for the c.3145C>T: p.R1049X variant who showed late-onset sensorineural hearing loss in her fifth decade. p.R1049X lacks F-actin elongation activity because this variant truncates one-third of the FH2 domain, which is vital for DIAPH1 dimerization and processive F-actin elongation activity. Concordantly, no increase of F-actin or processive F-actin elongation activity was observed after overexpression of p.R1049X DIAPH1 in HeLa cells or by single-molecule microscopy using Xenopus XTC cells. However, overexpression of the p.R1049X variant impairs formation of cell-cell junctions and mitosis. We speculate that late-onset hearing loss is a long-term consequence of heterozygosity for the recessive p.R1049X variant, a phenotype that may have been overlooked among carriers of other recessive alleles of DIAPH1.
    Corresponding, Apr. 2022, Clinical genetics, 101(4) (4), 466 - 471, English, International magazine
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Hiroaki Mohri, Yuzuru Ninoyu, Hirofumi Sakaguchi, Shigeru Hirano, Naoaki Saito, Takehiko Ueyama
    Reactive oxygen species (ROS) produced by NADPH oxidases (Nox) contribute to the development of different types of sensorineural hearing loss (SNHL), a common impairment in humans with no established treatment. Although the essential role of Nox3 in otoconia biosynthesis and its possible involvement in hearing have been reported in rodents, immunohistological methods targeted at detecting Nox3 expression in inner ear cells reveal ambiguous results. Therefore, the mechanism underlying Nox3-dependent SNHL remains unclear and warrants further investigation. We generated Nox3-Cre knock-in mice, in which Nox3 was replaced with Cre recombinase (Cre). Using Nox3-Cre;tdTomato mice of either sex, in which tdTomato is expressed under the control of the Nox3 promoter, we determined Nox3-expressing regions and cell types in the inner ear. Nox3-expressing cells in the cochlea included various types of supporting cells, outer hair cells, inner hair cells, and spiral ganglion neurons. Nox3 expression increased with cisplatin, age, and noise insults. Moreover, increased Nox3 expression in supporting cells and outer hair cells, especially at the basal turn of the cochlea, played essential roles in ROS-related SNHL. The extent of Nox3 involvement in SNHL follows the following order: cisplatin-induced hearing loss > age-related hearing loss > noise-induced hearing loss. Here, on the basis of Nox3-Cre;tdTomato, which can be used as a reporter system (Nox3-Cre +/- ;tdTomato +/+ and Nox3-Cre +/+ ;tdTomato +/+), and Nox3-KO (Nox3-Cre +/+ ;tdTomato +/+) mice, we demonstrate that Nox3 inhibition in the cochlea is a promising strategy for ROS-related SNHL, such as cisplatin-induced HL, age-related HL, and noise-induced HL.SIGNIFICANCE STATEMENT We found Nox3-expressing regions and cell types in the inner ear, especially in the cochlea, using Nox3-Cre;tdTomato mice, a reporter system generated in this study. Nox3 expression increased with cisplatin, age, and noise insults in specific cell types in the cochlea and resulted in the loss (apoptosis) of outer hair cells. Thus, Nox3 might serve as a molecular target for the development of therapeutics for sensorineural hearing loss, particularly cisplatin-induced, age-related, and noise-induced hearing loss.
    Corresponding, May 2021, The Journal of neuroscience : the official journal of the Society for Neuroscience, 41(21) (21), 4716 - 4731, English, International magazine
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Yuzuru Ninoyu, Hirofumi Sakaguchi, Chen Lin, Toshiaki Suzuki, Shigeru Hirano, Yasuo Hisa, Naoaki Saito, Takehiko Ueyama
    Dia1, which belongs to the diaphanous-related formin family, influences a variety of cellular processes through straight actin elongation activity. Recently, novel DIA1 mutants such as p.R1213X (p.R1204X) and p.A265S, have been reported to cause an autosomal dominant sensorineural hearing loss (DFNA1). Additionally, active DIA1 mutants induce progressive hearing loss in a gain-of-function manner. However, the subcellular localization and pathological function of DIA1(R1213X/R1204X) remains unknown. In the present study, we demonstrated the localization of endogenous Dia1 and the constitutively active DIA1 mutant in the cochlea, using transgenic mice expressing FLAG-tagged DIA1(R1204X) (DIA1-TG). Endogenous Dia1 and the DIA1 mutant were regionally expressed at the organ of Corti and the spiral ganglion from early life; alongside cochlear maturation, they became localized at the apical junctional complexes (AJCs) between hair cells (HCs) and supporting cells (SCs). To investigate HC vulnerability in the DIA1-TG mice, we exposed 4-week-old mice to moderate noise, which induced temporary threshold shifts with cochlear synaptopathy and ultrastructural changes in stereocilia 4 weeks post noise exposure. Furthermore, we established a knock-in (KI) mouse line expressing AcGFP-tagged DIA1(R1213X) (DIA1-KI) and confirmed mutant localization at AJCs and the tips of stereocilia in HCs. In MDCKAcGFP-DIA1(R1213X) cells with stable expression of AcGFP-DIA1(R1213X), AcGFP-DIA1(R1213X) revealed marked localization at microvilli on the apical surface of cells and decreased localization at cell-cell junctions. The DIA1-TG mice demonstrated hazy and ruffled circumferential actin belts at AJCs and abnormal stereocilia accompanied with HC loss at 5 months of age. In conclusion, Dia1 plays a pivotal role in the development and maintenance of AJCs and stereocilia, ensuring cochlear and HC integrity. Subclinical/latent vulnerability of HCs may be the cause of progressive hearing loss in DFNA1 patients, thus suggesting new therapeutic targets for preventing HC degeneration and progressive hearing loss associated with DFNA1.
    Corresponding, Jul. 2020, Cell death & disease, 11(7) (7), 536 - 536, English, International magazine
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Shigefumi Morioka, Hirofumi Sakaguchi, Hiroaki Mohri, Mariko Taniguchi-Ikeda, Motoi Kanagawa, Toshiaki Suzuki, Yuko Miyagoe-Suzuki, Tatsushi Toda, Naoaki Saito, Takehiko Ueyama
    Corresponding, Public Library of Science (PLoS), May 2020, PLOS Genetics, 16(5) (5), e1008826 - e1008826, English
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • UEYAMA TAKEHIKO, Megumi Sakuma, Mio Nakatsuji, Tatsuya Uebi, Takeshi Hamada, Atsu Aiba, SAITO NAOAKI
    Jan. 2020, J Investigative Dermatology, 140, 75 - 84, English
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Differential disruption of autoinhibition and defect in assembly of cytoskeleton during cell division decide the fates of human DIAPH1-related cytoskeletopathy
    Bong Jik Kim, UEYAMA TAKEHIKO, Takushi Miyoshi, Shin-ichiro Kitajiri, Byung Yoon Choi
    Dec. 2019, J Medical Genetics, 56(12) (12), 818 - 827, English
    [Refereed]
    Scientific journal

  • Shirafuji T, Shimazaki H, Miyagi T, Ueyama T, Adachi N, Tanaka S, Hide I, Saito N, Sakai N.
    Spinocerebellar ataxia type 14 (SCA14) is an autosomal dominant neurodegenerative disorder characterized by cerebellar ataxia with myoclonus, dystonia, spasticity, and rigidity. Although missense mutations and a deletion mutation have been found in the protein kinase C gamma (PRKCG) gene encoding protein kinase C γ (PKCγ) in SCA14 families, a nonsense mutation has not been reported. The patho-mechanisms underlying SCA14 remain poorly understood. However, gain-of-function mechanisms and loss-of-function mechanisms, but not dominant negative mechanisms, were reported the patho-mechanism of SCA14. We identified the c.226C>T mutation of PRKCG, which caused the p.R76X in PKCγ by whole-exome sequencing in patients presenting cerebellar atrophy with cognitive and hearing impairment. To investigate the patho-mechanism of our case, we studied aggregation formation, cell death, and PKC inhibitory effect by confocal microscopy, western blotting with cleaved caspase 3, and pSer PKC motif antibodies, respectively. PKCγ(R76X)-GFP have aggregations the same as wild-type (WT) PKCγ-GFP. The PKCγ(R76X)-GFP inhibited PKC phosphorylation activity more than GFP alone. It also induced more apoptosis in COS7 and SH-SY5Y cells compared to WT-PKCγ-GFP and GFP. We first reported SCA14 patients with p.R76X in PKCγ who have cerebellar atrophy with cognitive and hearing impairment. Our results suggest that a dominant negative mechanism due to truncated peptides produced by p.R76X may be at least partially responsible for the cerebellar atrophy.
    Jul. 2019, Molecular and Cellular Neuroscience, 98, 46 - 53, English, International magazine
    [Refereed]
    Scientific journal

  • Takehiko Ueyama
    The small GTPases of the Rho-family (Rho-family GTPases) have various physiological functions, including cytoskeletal regulation, cell polarity establishment, cell proliferation and motility, transcription, reactive oxygen species (ROS) production, and tumorigenesis. A relatively large number of downstream targets of Rho-family GTPases have been reported for in vitro studies. However, only a small number of signal pathways have been established at the in vivo level. Cumulative evidence for the functions of Rho-family GTPases has been reported for in vivo studies using genetically engineered mouse models. It was based on different cell- and tissue-specific conditional genes targeting mice. In this review, we introduce recent advances in in vivo studies, including human patient trials on Rho-family GTPases, focusing on highly polarized sensory organs, such as the cochlea, which is the primary hearing organ, host defenses involving reactive oxygen species (ROS) production, and tumorigenesis (especially associated with RAC, novel RAC1-GSPT1 signaling, RHOA, and RHOBTB2).
    Jan. 2019, Cells, 8(2) (2), English, International magazine
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Nakazono A, Adachi Naoko, Takahashi H, Seki T, Hamada D, Ueyama Takehiko, Sakai N, Saito Naoaki
    Sep. 2018, J Biol Chem, 293(38) (38), 14758 - 14774, English
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • MORIOKA SHIGEFUMI, SAKAGUCHI HIROFUMI, Yamaguchi T, Ninoyu Y, Mohri H, Nakamura T, Hisa Y, Ogita K, SAITO NAOAKI, UEYAMA TAKEHIKO
    Previous studies have convincingly argued that reactive oxygen species (ROS) contribute to the development of several major types of sensorineural hearing loss, such as noise-induced hearing loss (NIHL), drug-induced hearing loss, and age-related hearing loss. However, the underlying molecular mechanisms induced by ROS in these pathologies remain unclear. To resolve this issue, we established an in vivo model of ROS overproduction by generating a transgenic (TG) mouse line expressing the human NADPH oxidase 4 (NOX4, NOX4-TG mice), which is a constitutively active ROS-producing enzyme that does not require stimulation or an activator. Overproduction of ROS was detected at the cochlea of the inner ear in NOX4-TG mice, but they showed normal hearing function under baseline conditions. However, they demonstrated hearing function vulnerability, especially at high-frequency sounds, upon exposure to intense noise, which was accompanied by loss of cochlear outer hair cells (OHCs). The vulnerability to loss of hearing function and OHCs was rescued by treatment with the antioxidant Tempol. Additionally, we found increased protein levels of the heat-shock protein 47 (HSP47) in models using HEK293 cells, including H2 O2 treatment and cells with stable and transient expression of NOX4. Furthermore, the up-regulated levels of Hsp47 were observed in both the cochlea and heart of NOX4-TG mice. Thus, antioxidant therapy is a promising approach for the treatment of NIHL. Hsp47 may be an endogenous antioxidant factor, compensating for the chronic ROS overexposure in vivo, and counteracting ROS-related hearing loss.
    Corresponding, Aug. 2018, J Neurochemistry, 146(4) (4), 459 - 473, English, International magazine
    [Refereed]
    International conference proceedings
    Link to Kobe Univ. Repository (Kernel)

  • Isao Sakamoto, Takehiko Ueyama, Masakazu Hayashibe, Takashi Nakamura, Hiroaki Mohri, Hiroshi Kiyonari, Michiko Shigyo, Chihiro Tohda, Naoaki Saito
    Academic Press Inc., Apr. 2018, Experimental Neurology, 302, 57 - 67, English
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Toshihiko Shirafuji, Takehiko Ueyama, Naoko Adachi, Ken-Ichi Yoshino, Yusuke Sotomaru, Junsuke Uwada, Azumi Kaneoka, Taro Ueda, Shigeru Tanaka, Izumi Hide, Naoaki Saito, Norio Sakai
    Society for Neuroscience, Jan. 2018, Journal of Neuroscience, 38(2) (2), 278 - 290, English
    [Refereed]
    Scientific journal

  • Cheryl M. Hanes, Anna E. D'Amico, Takehiko Ueyama, Alexander C. Wong, Xuexin Zhang, W. Frederick Hynes, Margarida M. Barroso, Nathaniel C. Cady, Mohamed Trebak, Naoaki Saito, Michelle R. Lennartz
    Jul. 2017, JOURNAL OF IMMUNOLOGY, 199(1) (1), 271 - 277, English
    [Refereed]
    Scientific journal

  • Takashi Nakamura, Takehiko Ueyama, Yuzuru Ninoyu, Hirofumi Sakaguchi, Narantsog Choijookhuu, Yoshitaka Hishikawa, Hiroshi Kiyonari, Masaaki Kohta, Mizuho Sakahara, Ivan de Curtis, Eiji Kohmura, Yasuo Hisa, Atsu Aiba, Naoaki Saito
    Rac signaling impacts a relatively large number of downstream targets; however, few studies have established an association between Rac pathways and pathological conditions. In the present study, we generated mice with double knockout of Rac1 and Rac3 (Atoh1-Cre;Rac1flox/flox;Rac3-/- ) in cerebellar granule neurons (CGNs). We observed impaired tangential migration at E16.5, as well as numerous apoptotic CGNs at the deepest layer of the external granule layer (EGL) in the medial cerebellum of Atoh1-Cre;Rac1flox/flox;Rac3-/- mice at P8. Atoh1-Cre;Rac1flox/flox;Rac3-/- CGNs differentiated normally until expression of p27kip1 and NeuN in the deep EGL at P5. Primary CGNs and cerebellar microexplants from Atoh1-Cre;Rac1flox/flox;Rac3-/- mice exhibited impaired neuritogenesis, which was more apparent in Map2-positive dendrites. Such findings suggest that impaired tangential migration and final differentiation of CGNs have resulted in decreased cerebellum size and agenesis of the medial internal granule layer, respectively. Furthermore, Rac depleted/deleted cells exhibited decreased levels of Mid1 and impaired mTORC1 signaling. Mid1 depletion in CGNs produced mild impairments in neuritogenesis and reductions in mTORC1 signaling. Thus, a novel Rac-signaling pathway (Rac1-Mid1-mTORC1) may be involved in medial cerebellar development.
    May 2017, Development (Cambridge, England), 144(10) (10), 1863 - 1875, English, International magazine
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Taiji Ishii, Takehiko Ueyama, Michiko Shigyo, Masaaki Kohta, Takeshi Kondoh, Tomoharu Kuboyama, Tatsuya Uebi, Takeshi Hamada, David H Gutmann, Atsu Aiba, Eiji Kohmura, Chihiro Tohda, Naoaki Saito
    Astrogliosis (i.e. glial scar), which is comprised primarily of proliferated astrocytes at the lesion site and migrated astrocytes from neighboring regions, is one of the key reactions in determining outcomes after CNS injury. In an effort to identify potential molecules/pathways that regulate astrogliosis, we sought to determine whether Rac/Rac-mediated signaling in astrocytes represents a novel candidate for therapeutic intervention following CNS injury. For these studies, we generated mice with Rac1 deletion under the control of the GFAP (glial fibrillary acidic protein) promoter (GFAP-Cre;Rac1flox/flox). GFAP-Cre;Rac1flox/flox (Rac1-KO) mice exhibited better recovery after spinal cord injury and exhibited reduced astrogliosis at the lesion site relative to control. Reduced astrogliosis was also observed in Rac1-KO mice following microbeam irradiation-induced injury. Moreover, knockdown (KD) or KO of Rac1 in astrocytes (LN229 cells, primary astrocytes, or primary astrocytes from Rac1-KO mice) led to delayed cell cycle progression and reduced cell migration. Rac1-KD or Rac1-KO astrocytes additionally had decreased levels of GSPT1 (G1 to S phase transition 1) expression and reduced responses of IL-1β and GSPT1 to LPS treatment, indicating that IL-1β and GSPT1 are downstream molecules of Rac1 associated with inflammatory condition. Furthermore, GSPT1-KD astrocytes had cell cycle delay, with no effect on cell migration. The cell cycle delay induced by Rac1-KD was rescued by overexpression of GSPT1. Based on these results, we propose that Rac1-GSPT1 represents a novel signaling axis in astrocytes that accelerates proliferation in response to inflammation, which is one important factor in the development of astrogliosis/glial scar following CNS injury.
    Jan. 2017, The Journal of biological chemistry, 292(4) (4), 1240 - 1250, English, International magazine
    [Refereed]
    Scientific journal

  • Hirofumi Sakaguchi, Takashi Nakamura, Takehiko Ueyama
    Japan Society for Equilibrium Research, 2017, Equilibrium Research, 76(6) (6), 720 - 726, Japanese
    [Refereed]
    Scientific journal

  • Toshihiko Shirafuji, Takehiko Ueyama, Shigeru Tanaka, Izumi Hide, Naoaki Saito, Norio Sakai
    2017, ACTA HISTOCHEMICA ET CYTOCHEMICA, 50(6) (6), 177 - 180, English
    [Refereed]
    Scientific journal

  • Takehiko Ueyama, Yuzuru Ninoyu, Shin-ya Nishio, Takushi Miyoshi, Hiroko Torii, Koji Nishimura, Kazuma Sugahara, Hideaki Sakata, Dean Thumkeo, Hirofumi Sakaguchi, Naoki Watanabe, Shin-ichi Usami, Naoaki Saito, Shin-ichiro Kitajiri
    Nov. 2016, EMBO MOLECULAR MEDICINE, 8(11) (11), 1310 - 1324, English
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Takehiko Ueyama, Megumi Sakuma, Yuzuru Ninoyu, Takeshi Hamada, Corinne Dupuy, Miklós Geiszt, Thomas L. Leto, Naoaki Saito
    American Society for Biochemistry and Molecular Biology Inc., Mar. 2015, Journal of Biological Chemistry, 290(10) (10), 6495 - 6506, English
    [Refereed]
    Scientific journal

  • Hideyuki Takahashi, Naoko Adachi, Toshihiko Shirafuji, Sally Danno, Takehiko Ueyama, Michele Vendruscolo, Anton N. Shuvaev, Takuya Sugimoto, Takahiro Seki, Daizo Hamada, Kazuhiro Irie, Hirokazu Hirai, Norio Sakai, Naoaki Saito
    Jan. 2015, HUMAN MOLECULAR GENETICS, 24(2) (2), 525 - 539, English
    [Refereed]
    Scientific journal

  • RANTES/CCL5 mediated-biological effects depend on the syndecan-4/PKCα signaling pathway
    Maillard L, Saito Naoaki, Hlawaty H, Friand V, Suffee N, Chmilewsky F, Haddad O, Laguillier C, Guyot E, Ueyama Takehiko, Oudar O, Sutton A, Charnaux N
    Sep. 2014, Biol Open, 3(10) (10), 995 - 1004, English
    [Refereed]
    Scientific journal

  • Toshihiko Shirafuji, Takehiko Ueyama, Ken-ichi Yoshino, Hideyuki Takahashi, Naoko Adachi, Yukio Ago, Ken Koda, Tetsuaki Nashida, Naoki Hiramatsu, Toshio Matsuda, Tatsushi Toda, Norio Sakai, Naoaki Saito
    Jul. 2014, JOURNAL OF NEUROSCIENCE, 34(28) (28), 9268 - 9280, English
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Takehiko Ueyama, Hirofumi Sakaguchi, Takashi Nakamura, Akihiro Goto, Shigefumi Morioka, Aya Shimizu, Kazuki Nakao, Yoshitaka Hishikawa, Yuzuru Ninoyu, Hidetoshi Kassai, Shiro Suetsugu, Takehiko Koji, Bernd Fritzsch, Shigenobu Yonemura, Yasuo Hisa, Michiyuki Matsuda, Atsu Aiba, Naoaki Saito
    May 2014, JOURNAL OF CELL SCIENCE, 127(9) (9), 2040 - 2052, English
    [Refereed]
    Scientific journal

  • Hyun Choi, Ju-yearl Park, Hyoung-June Kim, Minsoo Noh, Takehiko Ueyama, Yunsoo Bae, Tae Ryong Lee, Dong Wook Shin
    Apr. 2014, JOURNAL OF DERMATOLOGICAL SCIENCE, 74(1) (1), 56 - 63, English
    [Refereed]
    Scientific journal

  • Megumi Sakuma, Yasuhito Shirai, Takehiko Ueyama, Naoaki Saito
    Mar. 2014, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 445(2) (2), 340 - 345, English
    [Refereed]
    Scientific journal

  • Takehiko Ueyama, Jeonghyun Son, Takeshi Kobayashi, Takeshi Hamada, Takashi Nakamura, Hirofumi Sakaguchi, Toshihiko Shirafuji, Naoaki Saito
    Sep. 2013, JOURNAL OF IMMUNOLOGY, 191(5) (5), 2560 - 2569, English
    [Refereed]
    Scientific journal

  • Daniel Schlam, Michal Bohdanowicz, Alexandros Chatilialoglu, Benjamin E. Steinberg, Takehiko Ueyama, Guangwei Du, Sergio Grinstein, Gregory D. Fairn
    Aug. 2013, Journal of Biological Chemistry, 288(32) (32), 23090 - 23104, English
    [Refereed]
    Scientific journal

  • Michal Bohdanowicz, Daniel Schlam, Martin Hermansson, David Rizzuti, Gregory D. Fairn, Takehiko Ueyama, Pentti Somerharju, Guangwei Du, Sergio Grinstein
    Jun. 2013, Molecular Biology of the Cell, 24(11) (11), 1700 - 1712, English
    [Refereed]
    Scientific journal

  • Kouhei Yamashita, Takashi Miyoshi, Yasuyuki Arai, Kiyomi Mizugishi, Akifumi Takaori-Kondo, Takehiko Ueyama
    Apr. 2013, INTERNATIONAL JOURNAL OF HEMATOLOGY, 97(4) (4), 505 - 510, English
    [Refereed]
    Scientific journal

  • Takehiko Ueyama, Junya Nakakita, Takashi Nakamura, Takeshi Kobayashi, Toshihiro Kobayashi, Jeonghyun Son, Megumi Sakuma, Hirofumi Sakaguchi, Thomas L. Leto, Naoaki Saito
    Nov. 2011, JOURNAL OF BIOLOGICAL CHEMISTRY, 286(47) (47), 40693 - 40705, English
    [Refereed]
    Scientific journal

  • Yasuhito Shirai, Shoko Morioka, Megumi Sakuma, Ken-ichi Yoshino, Chihiro Otsuji, Norio Sakai, Kaori Kashiwagi, Kazuhiro Chida, Ryutaro Shirakawa, Hisanori Horiuchi, Chikako Nishigori, Takehiko Ueyama, Naoaki Saito
    Apr. 2011, MOLECULAR BIOLOGY OF THE CELL, 22(8) (8), 1340 - 1352, English
    [Refereed]
    Scientific journal

  • Takumi Kawasaki, Takehiko Ueyama, Ingo Lange, Stefan Feske, Naoaki Saito
    Aug. 2010, JOURNAL OF BIOLOGICAL CHEMISTRY, 285(33) (33), 25720 - 25730, English
    [Refereed]
    Scientific journal

  • Eisuke Adachi, Yutaka Kazoe, Yohei Sato, Yuko Suzuki, Tetsumei Urano, Takehiko Ueyama, Naoaki Saito, Viacheslav O. Nikolaev, Martin J. Lohse, Makoto Tominaga, Hideo Mogami
    Nov. 2009, PFLUGERS ARCHIV-EUROPEAN JOURNAL OF PHYSIOLOGY, 459(1) (1), 227 - 234, English
    [Refereed]
    Scientific journal

  • Thomas L. Leto, Stanislas Morand, Darrell Hurt, Takehiko Ueyama
    Last, Oct. 2009, ANTIOXIDANTS & REDOX SIGNALING, 11(10) (10), 2607 - 2619, English
    [Refereed]
    Scientific journal

  • Stanislas Morand, Takehiko Ueyama, Satoshi Tsujibe, Naoaki Saito, Agnieszka Korzeniowska, Thomas L. Leto
    Apr. 2009, FASEB JOURNAL, 23(4) (4), 1205 - 1218, English
    [Refereed]
    Scientific journal

  • Noriaki Ashida, Takehiko Ueyama, Kyoko Rikitake, Yasuhito Shirai, Mika Eto, Takeshi Kondoh, Eiji Kohmura, Naoaki Saito
    Dec. 2008, NEUROSCIENCE LETTERS, 446(2-3) (2-3), 123 - 128, English
    [Refereed]
    Scientific journal

  • Sequential binding of cytosolic phox complex to phagosomes through regulated adaptor proteins: Evaluation using the novel monomeric kusabira-green system and live imaging of phagocytosis
    Takehiko Ueyama, Tomoko Kusakabe, Satoshi Karasawa, Takumi Kawasaki, Aya Shimizu, Jeonghyun Son, Thomas L. Leto, Atsushi Miyawaki, Naoaki Saito
    Lead, Jul. 2008, JOURNAL OF IMMUNOLOGY, 181(1) (1), 629 - 640, English
    [Refereed]
    Scientific journal

  • Naoko Adachi, Takeshi Kobayashi, Hideyuki Takahashi, Takumi Kawasaki, Yasuhito Shirai, Takehiko Ueyama, Toshio Matsuda, Takahiro Seki, Norio Sakai, Naoaki Saito
    Jul. 2008, JOURNAL OF BIOLOGICAL CHEMISTRY, 283(28) (28), 19854 - 19863, English
    [Refereed]
    Scientific journal

  • Takumi Kawasaki, Takeshi Kobayashi, Takehiko Ueyama, Yasuhito Shirai, Naoaki Saito
    Jan. 2008, BIOCHEMICAL JOURNAL, 409, 471 - 479, English
    [Refereed]
    Scientific journal

  • Takehiko Ueyama, Toshihiko Tatsuno, Takumi Kawasaki, Satoshi Tsujibe, Yasuhito Shirai, Hideki Sumimoto, Thomas L. Leto, Naoaki Saito
    Lead, Feb. 2007, MOLECULAR BIOLOGY OF THE CELL, 18(2) (2), 441 - 454, English
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Takehiko Ueyama, Kristen Lekstrom, Satoshi Tsujibe, Naoaki Saito, Thomas L. Leto
    Jan. 2007, FREE RADICAL BIOLOGY AND MEDICINE, 42(2) (2), 180 - 190, English
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • T Matsubara, Y Shirai, K Miyasaka, T Murakami, Y Yamaguchi, T Ueyama, M Kai, F Sakane, H Kanoh, T Hashimoto, S Kamada, U Kikkawa, N Saito
    Mar. 2006, JOURNAL OF BIOLOGICAL CHEMISTRY, 281(10) (10), 6152 - 6164, English
    [Refereed]
    Scientific journal

  • T Ueyama, M Geiszt, TL Leto
    Mar. 2006, MOLECULAR AND CELLULAR BIOLOGY, 26(6) (6), 2160 - 2174, English
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • KL Cheeseman, T Ueyama, TM Michaud, K Kashiwagi, D Wang, LA Flax, Y Shirai, DJ Loegering, N Saito, MR Lennartz
    Feb. 2006, MOLECULAR BIOLOGY OF THE CELL, 17(2) (2), 799 - 813, English
    [Refereed]
    Scientific journal

  • Isoform-specific membrane targeting mechanism of Rac during Fc gamma R-mediated phagocytosis: Positive charge-dependent and independent targeting mechanism of Rac to the phagosome
    T Ueyama, M Eto, K Kami, T Tatsuno, T Kobayashi, Y Shirai, MR Lennartz, R Takeya, H Sumimoto, N Saito
    Lead, Aug. 2005, JOURNAL OF IMMUNOLOGY, 175(4) (4), 2381 - 2390, English
    [Refereed]
    Scientific journal

  • S Hayashi, T Ueyama, T Kajimoto, K Yagi, E Kohmura, N Saito
    May 2005, JOURNAL OF NEUROCHEMISTRY, 93(4) (4), 883 - 891, English
    [Refereed]
    Scientific journal

  • Superoxide production at phagosomal cup/phagosome through beta I protein kinase C during Fc gamma R-mediated phagocytosis in microglia
    T Ueyama, MR Lennartz, Y Noda, T Kobayashi, Y Shirai, K Kikitake, T Yamasaki, S Hayashi, N Sakai, H Seguchi, M Sawada, H Sumimoto, N Saito
    Lead, Oct. 2004, JOURNAL OF IMMUNOLOGY, 173(7) (7), 4582 - 4589, English
    [Refereed]
    Scientific journal

  • A Kageyama, M Oka, T Okada, S Nakamura, T Ueyama, N Saito, VJ Hearing, M Ichihashi, C Nishigori
    Jun. 2004, JOURNAL OF BIOLOGICAL CHEMISTRY, 279(26) (26), 27774 - 27780, English
    [Refereed]
    Scientific journal

  • AK Bhattacharjee, T Ueyama, T Kondoh, S Hayashi, A Abouelfetouh, N Sakai, N Saito, E Kohmura
    May 2004, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 317(4) (4), 1144 - 1148, English
    [Refereed]
    Scientific journal

  • K Sakai, C Hasegawa, M Okura, O Morikawa, T Ueyama, Y Shirai, N Sakai, N Saito
    May 2003, NEUROSCIENCE LETTERS, 342(3) (3), 175 - 178, English
    [Refereed]
    Scientific journal

  • Elaine C. Larsen, Takehiko Ueyama, Pamela M. Brannock, Yasuhito Shirai, Naoaki Saito, Christer Larsson, Daniel Loegering, Peter B. Weber, Michelle R. Lennartz
    Dec. 2002, Journal of Cell Biology, 159(6) (6), 939 - 944, English
    Scientific journal

  • Naoki Irie, Norio Sakai, Takehiko Ueyama, Taketoshi Kajimoto, Yasuhito Shirai, Naoaki Saito
    2002, Biochemical and Biophysical Research Communications, 298(5) (5), 738 - 743, English
    [Refereed]
    Scientific journal

  • Generation of a constitutively active fragment of PKN in microglia/macrophages after middle cerebral artery occlusion in rats
    Ueyama T, Y Ren, N Sakai, M Takahashi, Y Ono, T Kondoh, N Tamaki, N Saito
    Lead, Nov. 2001, JOURNAL OF NEUROCHEMISTRY, 79(4) (4), 903 - 913, English
    [Refereed]
    Scientific journal

  • cDNA cloning of an alternative splicing variant of protein kinase C delta (PKC delta III), a new truncated form of PKC delta, in rats
    T Ueyama, Y Ren, S Ohmori, K Sakai, N Tamaki, N Saito
    Lead, Mar. 2000, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 269(2) (2), 557 - 563, English
    [Refereed]
    Scientific journal

  • Non-traumatic acute paraplegia associated with cervical disk herniation: A case report
    UEYAMA TAKEHIKO
    Jan. 1999, Surgical Neurology, 52, 204 - 207, English
    [Refereed]
    Scientific journal

  • Bridging veins on the tentorial surface of the cerebellum: A microsurgical anatomic study and operative considerations
    Ueyama T, Al-Mefty O, Tamaki N
    Nov. 1998, Neurosurgery, 43(5) (5), 1137 - 1145
    [Refereed]

  • UEYAMA T, TAMAKI N, KONDOH T, KURATA H
    Jan. 1998, SURGICAL NEUROLOGY, 50(1) (1), 30 - 32, English
    [Refereed]
    Scientific journal

  • Cerebellopontine angle ependymoma with internal auditory canal enlargement and pineal extension
    Takehiko Ueyama, Norihiko Tamaki, Takeshi Kondoh, Takashi Kokunai, Masahiro Asada
    Oct. 1997, Neurologia Medico-Chirurgica, 37(10) (10), 762 - 765, English
    [Refereed]
    Scientific journal

  • A case of traumatic extracranial internal carotid artery dissecting aneurysm treated by proximal ligation and STA-MCA bypass
    Takehiko Ueyama, Norihiko Tamaki, Yousuke Ishihara, Kazuki Hosoi, Takeshi Kondoh, Mitsugu Nakamura, Masahiro Asada
    Mar. 1997, Neurological Surgery, 25(3) (3), 253 - 258, Japanese
    Scientific journal

  • Transsphenoidal surgery for a case of empty sella syndrome associated with GH secreting pituitary adenoma
    Hideo Aihara, Norihiko Tamaki, Takehiko Ueyama, Yousuke Ishihara, Takeshi Kondoh
    Dec. 1996, Neurological Surgery, 24(12) (12), 1119 - 1123, Japanese
    [Refereed]
    Scientific journal

■ MISC
  • めまい疾患のモデル動物-遺伝子医療・再生医療への扉 聴覚・平衡覚の成立におけるRho-GTPaseの関与
    坂口 博史, 中村 高志, 上山 健彦
    (一社)日本めまい平衡医学会, Dec. 2017, Equilibrium Research, 76(6) (6), 720 - 726, Japanese

  • Cdc42による蝸牛有毛細胞不動毛の形態維持
    中村 高志, 坂口 博史, 森岡 繁文, 久 育男, 上山 健彦, 齋藤 尚亮
    耳鼻咽喉科ニューロサイエンス研究会, Jun. 2015, 耳鼻咽喉科ニューロサイエンス, 29, 24 - 26, Japanese

■ Lectures, oral presentations, etc.
  • 遺伝性難聴の診断と治療法の開発～モデルマウスを用いて
    上山健彦
    第98回日本薬理学会 (APPW2025) シンポジウム, Mar. 2025, Japanese
    [Invited]
    Invited oral presentation

  • NOX由来ROSによる後天性感音難聴の発症機序（標的）と治療法開発戦略
    毛利宏明, 上山健彦
    第96回日本生化学会大会 シンポジウム, Nov. 2023
    [Invited]
    Nominated symposium

  • アクチン細胞骨格異常が関与する感音難聴の発症機序～Rho-family small GTPasesに注目
    上山健彦
    第96回日本生化学会大会 シンポジウム, Oct. 2023
    [Invited]
    Invited oral presentation

  • Acquired hearing loss induced by ROS and challenges for therapy development based on its mechanism
    Takehiko Ueyama
    第95回日本薬理学会年会 シンポジウム, Mar. 2022
    [Invited]
    Public symposium

  • Sensorineural hearing loss: challenges for therapy development based on its mechanisms
    Takehiko Ueyama
    The 8th Neuroscience Network in Kobe symposium, Feb. 2022, English
    [Invited]
    Invited oral presentation

  • 遺伝子改変マウスからの疾患病態解明と治療法開発戦略へのフィードバック
    上山健彦
    第60回日本組織細胞化学会総会 シンポジウム, Sep. 2019, Japanese
    [Invited]
    Nominated symposium

  • Strategy based on knowledge obtained from patients and mice models for development of novel therapy against hereditary sensorineural hearing loss (HSNHL) causing from impaired actin turnover
    Takehiko Ueyama
    第92回日本薬理学会年会 シンポジウム, Mar. 2019
    [Invited]
    Public symposium

  • イメージング手法を駆使した聴・平衡覚機能障害治療法開発への挑戦
    上山健彦
    第59回日本組織細胞化学会総会 シンポジウム, Sep. 2018
    [Invited]
    Nominated symposium

  • Hearing loss causing from impaired actin dynamics: clinical implications
    Takehiko Ueyama
    International Congress of Korean Society of Otorhinolaryngology Head & Neck Surgery (ORL-HNS; ICORL 2018), Apr. 2018, English
    [Invited]
    Invited oral presentation

  • 脳・脊髄損傷後修復時のアストロサイトにおける新規シグナリング
    上山健彦
    第58回日本組織細胞化学会総会 シンポジウム, Sep. 2017
    [Invited]
    Nominated symposium

  • Analysis of hearing loss causing from failed maintenance of auditory hair’s integrity
    Takehiko Ueyama
    第89回日本薬理学会年会 シンポジウム, Mar. 2016
    [Invited]
    Public symposium

  • 生体防御に関する活性酸素産生酵素の食胞・頂側膜へのターゲティング及び会合メカニズム
    Ueyama Takehiko, Saito Naoaki
    第92回日本生理学会大会, Mar. 2015, English, 日本生理学会, 神戸, Domestic conference
    [Invited]
    Public symposium

  • Rho-family small GTPases
    Ueyama Takehiko, Saito Naoaki
    第55回日本組織細胞化学会総会 シンポジウム, Sep. 2014, Japanese, 日本組織細胞化学会, 松本, Domestic conference
    [Invited]
    Nominated symposium

  • Maintenance of stereocillia and apical junctional complexes by Cdc42 in cochlear hair cells.
    Ueyama Takehiko, Sakaguchi, H, Nakamura, T, Goto, A, Morioka, S, Fritzsch, B, Hisa, Y, Matsuda, M, Aiba, A, Saito Naoaki
    Society for Neuroscience 2014, Sep. 2014, English, Society for Neuroscience, Washington D.C., USA, International conference
    Oral presentation

  • 活性酸素産生NADPH oxidasesの食胞・頂側膜へのターゲティング及び会合メカニズム
    Ueyama Takehiko
    第6回 Asia International Institute of Infectious Disease Control (ADC) 研 シンポジウム, Aug. 2014, Japanese, 東京, Domestic conference
    [Invited]
    Invited oral presentation

  • Cdc42 is required for maintenance of stereocilia and apical junction complexes in cochlear hair cells
    Takehiko Ueyama
    Joint Workshop (UCL and Kobe University) in Brussels: Cell polarity and Cell adhesion, Mar. 2014, English
    [Invited]

  • Glial functions of Rho-family small GTPases during cerebellar development and after CNS injury
    Ueyama Takehiko, Kondoh T, Kohta M, Kohmura Eiji, Hayashibe M, Nakamura T, Sakahara M, Kassai H, Nakao K, Aiba A, Saito Naoaki
    The 14th international congress of Histochemistry and Cytochemistry (ICHC 2012), Aug. 2012, English, 京都, International conference
    Poster presentation

  • Brain damage and Rho Family GTPases
    Ueyama Takehiko, Saito Naoaki
    第85回日本薬理学会年会, Mar. 2012, English, 日本薬理学会, 京都, Domestic conference
    [Invited]
    Public symposium

  • Histological analysis of small G proteins using region spscific knock-out mice
    Ueyama Takehiko, Saito Naoaki
    第52回組織細胞化学会総会 シンポジウム, Sep. 2011, Japanese, 日本組織細胞化学会, 金沢, Domestic conference
    [Invited]
    Nominated symposium

  • Regulatory mechanisms of Rac activation by RhoGDI
    Takehiko Ueyama, Naoaki Saito
    第84回日本薬理学会年会 シンポジウム, Mar. 2011, English
    [Invited]
    Public symposium

  • Translocation/dissociation mechanisms of Rac-RhoGDI complex on membranes
    Takehiko Ueyama, Son J., Kobayashi T., Saito N.
    International Symposium on Cell Signaling and Gene Regulation in Taiwan, 2011, English
    [Invited]

  • Translocation/dissociation mechanisms of Rac-RhoGDI complex on membranes
    Takehiko Ueyama, Son J., Kobayashi T., Saito N.
    Gordon Research Conferences (NADPH oxidase), 2010, English

  • Dual oxidases (Duoxs) form cell surface complexes with Duox activators (Duoxas) affecting the specificity of ROS generation
    Takehiko Ueyama, Morand S., Tsujibe S., Korzeniowska A., Saito N., Leto TL
    The 24th International Union of Physiological Sciences (IUPS), 2009, English

  • Sequential targeting of cytosolic phox proteins to phagosomes through regulated adaptor proteins during Fc gamma R-mediated phagocytosis.
    Ueyama Takehiko, Kusakabe Tomoko, Karasawa Satoshi, Kawasaki Takumi, Shimizu Aya, Son Jeong-hyun, Leto L. Thomas
    第81回日本薬理学会年会, Mar. 2008, Japanese, パシフィコ横浜, Domestic conference
    Oral presentation

  • Sequential binding of cytosolic phox complex to phagosomes through
    Ueyama Takehiko, Kusakabe Tomoko, Karasawa Satoshi, Kawasaki Takumi, Shimizu Aya, Son Jeong-hyun, Leto L.Thomas, Miyawaki Atsushi, Saito Naoaki
    Gordon Research Conference (NADPH oxidase), 2008, English, ボストン, International conference
    Poster presentation

  • A regulated adaptor function of p40phox:Intramolecularinteraction(PX-PB1 domain)within p40phox.
    Ueyama Takehiko, Tatsuno Toshihiko, Kawasaki Takumi, Leto L. Thomas, Saito Naoaki
    Gordon Research Conference (Phagocytes), Jun. 2007, English, ロードアイランド, International conference
    Poster presentation

  • Nox2活性化におけるp40phoxのp67phoxに対するキャリア一蛋白としての機能獲得メカニズム：p40phoxのPX-PB1 domainを介した分子内結合の切断
    Ueyama Takehiko, Tatsuno Toshihiko, Kawasaki Takumi, Tsujibe Satoshi, Sumimoto Hideki, Leto L. Thomas, Saito Naoaki
    第80回日本薬理学会年会, Mar. 2007, Japanese, 名古屋国際会議場, Domestic conference
    Oral presentation

  • 貪食細胞(Nox2)と内耳上皮細胞(Nox3)おける活性化型NADPH oxidase複合体形成機構---Nox2:p40phoxのアダプターとしての機能獲得メカニズム---
    Ueyama Takehiko, Tatsuno Toshihiko, Kawasaki Takumi, Thomas L. Leto, Saito Naoaki
    第41回食細胞機能異常症研究会, Dec. 2006, Japanese, 東京経団連会館, Domestic conference
    [Invited]
    Invited oral presentation

  • 貪食細胞における活性化型NADPH oxidase複合体形成機構の可視化---p40phoxのアダプター蛋白としての機能獲得メカニズムの解明---
    Ueyama Takehiko, Tatsuno Toshihiko, Kawasaki Takunmi, Sumimoto Hideki, Tomas L. Leto, Saito Naoaki
    生理研シンポジウム, Oct. 2006, Japanese, 岡崎カンファレンスセンター, Domestic conference
    Oral presentation

  • Rac1 is an essential component for activation of multi-component Nox1-based NADPH oxidase.
    Ueyama Takehiko, Geiszt M, Saito Naoaki, Leto L.Thomas
    20th IUBMB International Congress of Biochemistry and Molecular Biology and 11th FAOBMB Congress, Jun. 2006, English, 国立京都国際会館, Domestic conference
    Poster presentation

  • Nox1, Nox2 and Nox3 require the involvement of Rac for their activation
    Takehiko Ueyama, Geiszt M., Leto TL
    Gordon Research Coference (NADPH oxidase), 2006, English

  • Is membrane-targeting of Nox activator (Noxa1 or p67phox) enough for activation of Nox family NADPH oxidase?
    akehiko Ueyama, Geiszt M., Leto TL
    The 11th MPO meeting, 2005, English

  • Analysis of isoform-specific function of PKC during phagocytosis in microglia
    Takehiko Ueyama, Lennartz MR, Kashiwagi K., Shirai Y., Sumimoto H., Sawada M., Saito N.
    The 33nd Annual Meeting of Society for Neuroscience, 2003, English

  • マイクログリアの貪食細胞における脂質メッセンジャーの役割
    上山健彦
    第75回日本生化学会大会 シンポジウム, Oct. 2002
    [Invited]
    Public symposium

  • Fragmentation of PKN in microglia after middle cerebral artery occlusion in rats
    Takehiko Ueyama, Ren Y., Sakai N., Takahashi M., Ono Y., Kondoh T., Tamaki N., Saito N.
    The 30nd Annual Meeting of Society for Neuroscience, 2000, English

■ Affiliated Academic Society
  • 米国免疫学会
    2013 - Present

  • 日本生化学会
    2009 - Present

  • 日本薬理学会
    2001 - Present

  • 北米神経科学会
    2000 - Present

  • 日本脳神経外科学会
    1992 - Present

■ Research Themes
  • 活性酸素産生酵素NOX3の発現制御機序解明による後天性及び片側性難聴の治療法開発
    上山 健彦, 坂口 博史, 足立 直子, 中村 高志
    日本学術振興会, 科学研究費助成事業, 基盤研究(B), 神戸大学, 01 Apr. 2024 - 31 Mar. 2027

  • 脊髄小脳変性症でのPKCリン酸化を介した神経保護機構の解明と新規治療法への応用
    白藤 俊彦, 上山 健彦
    日本学術振興会, 科学研究費助成事業, 基盤研究(C), 神戸大学, Apr. 2021 - Mar. 2024
    脊髄小脳変性症(SCA)患者の小脳プルキンエ細胞ではPKCリン酸化亢進が共通して起こり、小脳プルキンエ神経保護的に働くことが報告されている。 本研究の目的はSCAでリン酸化が亢進するPKCリン酸化基質やそのシグナル経路を同定し、そのPKCリン酸化経路をターゲットにしたSCA に共通して適用できる新規治療法を開発することである。 本年度は、SCAでPKCリン酸化が亢進する基質の同定を行った。第１に小脳プルキンエ細胞に発現し、SCAや神経変性疾患に重要であるタンパク質を候補として、細胞レベルでPKCによるリン酸化があるのかを同定した。SCA38、46、18 の原因遺伝子ELOVL5、PLD3、 IFRD1や、以前行った小脳をPKC刺激をして得たリン酸化プロテオームで同定した、GOLGA5、MTFR1L、 NSFL1C、TOMM70、VCP、PLEKHG4の9つのタンパク質にFLAGタグを付加した。そのプラスミドを細胞に導入し、その細胞をPKC刺激薬で処置し、pSer PKC AbでIBを行った。その結果、VCP, PLEKHG4がPKCによりリン酸化を受けることを確認した。 また、網羅的に小脳プルキンエ細胞で亢進しているPKCリン酸化基質を同定するための網羅的リン酸化プロテオームについては、マウスの全脳を用いた条件検討で、リン酸化変化の実験間の変動が大きいために脱リン酸化酵素阻害の手法を改善しているところである。

  • Acquired sensorineural hearing loss caused from Nox3-derived ROS
    上山 健彦, 坂口 博史, 足立 直子, 中村 高志
    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. 2024
    1. 自ら開発したNADPH oxidase 3 (Nox3) 発現細胞が赤色蛍光蛋白 tdTomato で標識される（Nox3-CreKI;tdTomato+/+）マウスを用い、耳石形成に必須のNox3由来活性酸素（ROS）発生源細胞として、内リンパ嚢・内リンパ管の管腔に面した上皮細胞を特定した。更に、Nox3が一次聴覚感受器官である蝸牛コルチ器に発現することを発見、Nox3発現細胞として、内・外有毛細胞とその周囲に存在し有毛細胞を機能・構造的に支える種々の支持細胞（内・外指節細胞、外柱細胞、クラウディウス細胞）を特定した。 2. 加えて、上記の蝸牛におけるNox3発現（tdTomato陽性）細胞数が、聴毒性で有名な抗癌剤であるシスプラチンの投与および加齢や騒音不可により上昇する事を発見した。特に、Nox3が発現誘導された外有毛細胞は、アポトーシスに陥ることを明らかにした。 3.自ら開発したNox3-knockout (KO)マウスを用いて、シスプラチン誘発感音難聴、加齢性感音難聴、騒音性感音難聴において、Nox3-KOが聴覚温存に働くことを明らかにした。 4.上記3種の主要後天性感音難聴の中で、シスプラチン誘発感音難聴と加齢性感音難聴においてNox3の関与が非常に高く、騒音性感音難聴ではやや低いが有意に関与することを明らかにした。
    上記の結果は、Nox3の蝸牛コルチ器（特に、外有毛細胞細胞）での発現抑制が、後天性感音難聴の治療法開発の標的になる事を強く示唆している。

  • Challenge for establishing the criteria of laterality-based diseases in hearing and vision
    Ueyama Takehiko
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Challenging Research (Exploratory), Challenging Research (Exploratory), Kobe University, Jun. 2019 - Mar. 2022
    We examined superoxide-producing capabilities in the cochlea and the retina from the age of 14 days after birth to 1-year-old. It was maximum from 2-month-old to 6-month-old, and then reached the plateau (from 6-month-old to 12-month-old). Moreover, it was enhanced/increased by aging and cisplatin, which is a famous anti-cancer drug exhibiting toxicity to hearing and vision. We also found that aging and cisplatin cause loss of outer hair cells in the cochlea and optic ganglion cells in the retina. These new findings suggest that diseases caused by superoxide may most frequently occur in the middle ages in humans.

  • Investigation for gene transfer therapy in inner ears
    SAKAGUCHI HIROFUMI
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), Apr. 2018 - Mar. 2021
    We studied gene transfer methods to inner ears using electroporation technique into mouse embryos. We successfully confirmed exogenous gene expression in the inner ear cells, including hair cells and supporting cells. We also studied molecular pathogenesis and gene expression patterns of dia1, large, POMGnT1 and Nox3 genes which are known to be related to certain types of sensorineural deafness, including hereditary, sound-induced, drug-induced and age-related hearing loss. Our investigation suggested that these genes are excellent candidate for molecular-target therapy to inner ear using gene transfer methods.

  • Development of a novel treatment for Parkinson's disease focusing on the protective effect of CSPa phosphorylation on synaptic terminal
    Shirafuji Toshihiko
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), Kobe University, Apr. 2018 - Mar. 2021
    I generated antibodies against Ser34 phosphorylation of CSPa (CSPa pS34 antibody) and confirmed that CSPa pS34 antibody can indeed specifically identify Ser34 phosphorylation. In CSPa extracted from whole mouse brain, the CSPa pS34 antibody detected a band. On the other hand, this antibody did not work in immunohistochemistry. Four AAV PHP.eB were generated under the control of rTHp promoter: CSPa-WT, phosphorylation-deficient (SA), phosphorylation-mimicking (SD), and H43Q. We confirmed their expression in the substantia nigra striatum by administration via the orbital venous plexus. The ratio of expression to that of endogenous CSPa is currently being verified.

  • SAITO NAOAKI
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (B), Kobe University, Apr. 2017 - Mar. 2020
    Neurodegenerative disease that affects cerebellar Purkinje cells, is characterized by the intracellular formation of neurotoxic amyloid-like aggregates of genetic variants of protein kinase Cgamma (PKCgamma). We studied the effect of Hsp90 inhibitors, such as celastrol and Hsp990, for the PKCgamma aggregation by up-regelation of Hsp70 to develop a new drug for various neurodegenerative diseases. Hsp990, a BBB permeable Hsp90 inhibitor, diminished net PKCgamma aggregation by preventing aggregate formation, resulting in decreased levels of apoptotic cell death among primary cultured Purkinje cells expressing PKCgamma variant. Furthermore, oral administration of Hsp990 decreased PKCgamma aggregation in SCA14 model mice. These compounds may be of utility as therapeutic agents against SCA14.
    Competitive research funding

  • Ueyama Takehiko
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (B), Kobe University, Apr. 2017 - Mar. 2020, Principal investigator
    1. Noise exposure induced significantly decreased number of the ribbon synapse in hair cells in mice expressing a DIA1 mutant (p.R1213X) compared with control mice, suggesting that noise is one of factors leading to progressive hearing loss in DFNA1 patients. 2. In mice expressing the DIA1 mutant, the mutant was localized at the apical junctional complex (AJC) of hair cells, where showed morphological abnormalities by TEM. Thus, AJC is likely the main lesion in DFNA1. 3. In aged mice expressing the DIA1 mutant, size of the platelets was spread over a wider range, compared with control mice. Detailed studies analyzing the mechanism manifesting the phenotype are going on.
    Competitive research funding

  • Ueyama Takehiko, SAITO Naoaki
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), Kobe University, Apr. 2014 - Mar. 2017, Principal investigator
    (1) We generated inner ear hair cell (HC)-specific KO mice to analyze the role of Cdc42 in HCs. HCs of Cdc42-KO mice developed normally but progressively degenerated after maturation, resulting in progressive hearing loss particularly at high frequencies. Adenovirus-encoded GFP-Cdc42 expression in HCs and fluorescence resonance energy transfer (FRET) imaging of HCs from transgenic mice expressing Cdc42-FRET biosensor indicated Cdc42 presence/activation at stereociliary membranes in cochlear HCs. (2) We found that the amount of active RhoA (GTP-form) is increased in Cdc42-KD cells. DIA1 is a downstream molecule of RhoA signaling pathways, and nucleates and elongates unbranched/straight actin. We discovered a novel patient-derived DIAPH1 mutation (c.3610C>T) in two unrelated Japanese families. Mice expressing the DIA1(R1204X) mutant experienced progressive deafness beginning at high frequencies and HC loss with various morphological abnormalities in stereocilia at the basal turn.
    Competitive research funding

  • SAITO NAOAKI, UEYAMA Takehiko
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Challenging Exploratory Research, Kobe University, Apr. 2014 - Mar. 2016
    We studied a new Rac-medicated pathway which can induce the differentiation of adipocytes. Using DNA microarray and RT-PCR, we found 5 factors (a,b,c,d,e) secreted from keraninocytes when Rac-mediated pathway is activated. Differentiation of adipocyte (3T3-L1 cells) was significantly enhanced when the cells were treated with the combination of a+b or a+c, much more than treated with a, b or c alone. We further elucidate whether the signal induces the diffentiation to white or beige adipocyte and we study the new drug target for the disorder of lipid/glucose metabolim.
    Competitive research funding

  • SAITO NAOAKI, ADACHI Naoko, UEYAMA Takehiko, SHIRAFUJI Toshihiko
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (B), Kobe University, Apr. 2013 - Mar. 2016
    We identified substrate proteins for PKCg as a new drug target for Parkinson disease. Nine substrate proteins were identified: Connexin-43, Disk1, MADD, CSPa, Calnexin, Stathmin, bPIX, NogoA, Adducin. Furthermore, 1) Phosphorylation of Ser583, Ser340 of bPIX is involved in DA release, 2) Phosphorylation of Connexin-43, MADD, Calnexin and Adducin is also related to DA release, 3) Phosphorylation of Ser10 of CSPa is important for cell viability.
    Competitive research funding

  • SAITOU Naoaki, UEYAMA Takehiko, KUSUNOKI Masato
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Challenging Exploratory Research, Kobe University, 2011 - 2012
    On the basis of our hypothesis that the etiology of Crohn’s disease is related with that of CGD enteritis due to p40phox gene abnormality, we examined the new cause of Crohn disease and its treatment. We examined the production of reactive oxygen, and p40phox gene mutations in treatment-resistant of the six cases the (refractory) patients with Crohn's disease, but we were not able to discover the CGD enteritis patients due to p40phox gene abnormality.
    Competitive research funding

  • UEYAMA Takehiko, SAKAGUCHI Hirofumi
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), Kobe University, 2010 - 2012, Principal investigator
    1. We engineered a genetically modified mice-line producing ROS, and found that the mice-line was vulnerable to noise exposure. The vulnerability to ROS was inhibited by the scavenger of ROS. 2. We engineered a knockout (KO) mice-line manifesting progressive hearing loss. Now we are extensively analyzing the molecular mechanism of deafness in the KO mice.
    Competitive research funding

  • 二国間交流「活性酸素種(ROS)特異的プローブの開発とDuoxによるROSメカニズムの解明」
    上山 健彦
    二国間交流事業（ハンガリーとの共同研究）, 2012, Principal investigator
    Competitive research funding

  • SAITO Naoaki, SHIRAI Yasuhito, UEYAMA Takehiko
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (B), Kobe University, 2010 - 2011
    Functional involvement of PKCγin the etiology of SCA14 and Parkinson disease was studied in cells and animals. We found 1) mutant PKCγs form amyloid-like fibrils in Purkinje cells and the formation was inhibitied by trehalose, 2) dysfunction of PKCγcould cause Parkinson disease by the inhibition of dopamine release and the loss of dopaminergic neurons in the substantia nigra.
    Competitive research funding

  • 二国間交流「活性酸素種(ROS)特異的プローブの開発とDuoxによるROSメカニズムの解明」
    上山 健彦
    二国間交流事業（ハンガリーとの共同研究）, 2011, Principal investigator
    Competitive research funding

  • 上山 健彦
    日本学術振興会, 科学研究費補助金／特定領域研究, 特定領域研究, 神戸大学, 2008 - 2009, Principal investigator
    【研究の背景と目的】貪食細胞により貪食された微生物は、細胞膜と細胞内膜器官の膜成分により形成される食胞膜に取り込まれ、食胞膜上で形成されるNADPH oxidase(Nox)の機能的複合体形成により産生される活性酸素種により殺菌・消化される。貪食細胞におけるNoxであるNox2の機能的複合体は、細胞膜成分であるcytochrome b_<558>と4つの細胞質成分(p47^, p67^, p40^, Rac)により成る。このように、食胞は、複数の細胞内コンパーメントが、個々の独立した機序により集合し、貪食時に食胞膜で始めて機能を発現する"活性酸素産生トランスポートソーム"と見なし得る。本研究は、この"活性酸素産生トランスポートソーム"の分子構成、脂質-蛋白質結合、時空間動態、複合体形成にかかわる分子間相互作用を解析することにより、膜上でのNoxの活性化型複合体による活性酸素を産生するメカニズムの解明を目指すものである。 【研究方法・研究内容】(1)p40^の構造変化を誘導する生理的機構のイメージング解析、(2) RhoGDI分子種による活性酸素産生制御機構のイメージング解析 を行った。 【研究結果】1. 生細胞を用いて、共焦点レーザー顕微鏡下での可視化により、刺激時にp40^の構造変化が起こるメカニズムとp40^がNox2の活性化促進因子として機能するメカニズムについて解明した。2. RhoGDI分子種の違いにより活性酸素産生抑制能力に違いがあることを突きとめ、そのメカニズムが、RhoGDI-Rac複合体の膜ターゲット様式の違いに起因することを明らかにした。これらの両研究成果については、現在論文作成中である。
    Competitive research funding

  • UEYAMA Takehiko
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), Kobe University, 2007 - 2008, Principal investigator
    近年、上皮細胞における活性酸素の産生が報告され、続いて組織特異的に存在する新規NADPH oxidase(Nox: 7種類) が確認され、その生理機能やその機能異常によって引き起こされる疾患などに注目が集っている。本研究では、新規Noxの活性化機構の詳細を、遺伝子操作マウスや操作マウス由来の細胞などを用いて解明することを試みた。 その結果、 1. 生細胞を用いて、共焦点レーザー顕微鏡下で、可視化により蛋白質相互作用を検知できる新規蛍光蛋白質システム(complementation-based method using a monomeric coral fluorescent protein: mKG system)を開発し、その有用性をNox複合体の相互作用を用いて証明し、報告した。 2. 遺伝子操作マウス由来の初代培養細胞とレンチウイルスを用いた Nox の再構築系の確立に成功した。
    Competitive research funding

  • SAITO Naoaki, SHIRAI Yasuhito, UEYAMA Takehiko
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (B), Kobe University, 2007 - 2008
    脊髄小脳変性症14型の発症メカニズムとして、変異型PKCγには細胞膜に十分な時間存在する能力が欠如しTRPCチャネルをリン酸化することができないため細胞外からのCa^<2+> 流入を抑制する機構が働かないことを示した。この過剰なCa^<2+> 流入が神経細胞死が招くと推測された。また他の要因として初代培養プルキンエ細胞において変異型PKCγは凝集体の形成・樹状突起の伸展・シナプス形成異常を引き起こす事を示した。
    Competitive research funding

  • 齋藤 尚亮
    日本学術振興会, 科学研究費補助金／特定領域研究, 特定領域研究, 神戸大学, 2006 - 2007
    脳虚血障害は発症後、現有の最高の治療法を用いても重度の障害を残す例は多くあり、新たな治療法の開発が期待されている。我々は、エストロゲンが細胞膜型エストロゲン受容体を介して神経細胞特異的なプロテインキナーゼCであるγPKCを活性化することにより、虚血後においても神経保護作用を示すことを見出した。一方、グリア細胞に発現するδPKCは、そのノックアウトマウスにおいて明らかな虚血巣の縮小が見られることから、γPKCと異なり神経障害作用を示すと考えられている。我々は、脳虚血による神経障害からの保護作用を示すエストロゲンの作用機序に着目し、この神経保護作用に対する神経、グリアそれぞれの独立した関与、およびニューロンーグリアのネットワークの役割について検討してきた。 今回、脳虚血時に神経細胞保護的に作用する細胞膜型エストロゲン受容体を新たに同定した。この細胞膜型エストロゲン受容体は、G蛋白質結合型受容体であり、エストロゲンによる神経保護作用に必須であるPKCγの活性化とともに、細胞内カルシウム濃度上昇を導くものであつた。また、本受容体は脳内に広く分布していることも免疫組織化学的に明らかにした。従来知られていた細胞質型とは異なる細胞膜型のエストロゲン受容体の研究は、性ステロイドのシグナル伝達機構の解明にとどまらず、脳虚血障害の新治療薬の開発につながると考えられる。
    Competitive research funding

  • 上山 健彦
    日本学術振興会, 科学研究費補助金／若手研究(B), 若手研究(B), 神戸大学, 2005 - 2006, Principal investigator
    非貧食細胞及び貧食細胞における活性酸素産生の制御機構の解明を行った。 1)貧食作用時におけるp47^-p67^-p40^複合体の食胞膜への移行メカニズムをp40^に注目して行った。p40^が、それ自身では膜移行能を持たないp67^を食胞膜に移行させる"アダプター蛋白"として機能すること明らかにした。さらに、p40^がアダプター蛋白として機能するメカニズムは、p40^の分子内結合(PX-PBI)の切断による、PI(3)Pに特異的結合能を持つPXdomainの露出ためであることを見出し、世界に先駆けて報告した。 2)非貧食細胞(上皮細胞)における活性酸素産生機構の解明に取り組み、酵素本体であるNADPH oxidase(Nox)の新規ホモログであるNox1とNox3の活性化機構の詳細とその活性化にはRac1が必要であることを見出し、世界に先駆けて報告した。 3)さらに、非貧食細胞(上皮細胞)における活性酸素産生機構の解明において、Noxを活性化する細胞質因子Noxo1(貧食細胞における細胞質因子であるp47^の上皮細胞におけるホモログ)の4種のアイソザイムの機能発現機構の詳細を報告した。 4)貧食作用時において貧食作用自体を制御する因子として、我々が報告しているePKCの食胞膜への集積機構の詳細を報告した。 5)貧食作用時におけるRacの食胞膜への集積機序について、Rac2は直接食胞膜に移動するのではなく、まず細胞内器官膜に移動し、その器官がRacが持つ器官移動促進作用を利用して食胞に癒合することにより、異物を包むのに必要な膜成分とRac自身の両方を食胞に供給するという、新説を提唱し報告した。
    Competitive research funding

  • SAITO Naoaki, SHIRAI Yasuhito, UEYAMA Takehiko
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (B), Kobe University, 2005 - 2006
    Recent technical development has enabled us to monitor the production, movement and signaling of lipid messengers in living cells using probes for specific lipids. In this project, we performed the spatio-temporal analysis using various probes for lipid messengers to elucidate when and where the lipid signaling occurs within the cells in physiological cellular responses. As a result, we obtained the results below. 1) Live imaging of signaling molecules (Protein kinase C (PKC) subtypes, Rac subtypes, p47^, p40^) in phagocytosis. Each molecule having different lipid binding ability translocated to the specific intracellular compartment in different time course. The finely tuned control of targeting mechanism of signaling molecules to lipid regulate the superoxide production in phagocytosis. 2) Functional and molecular interaction of Diacylglycerol (DG)-PKC-DG kinase (DGK) pathway. Direct binding between PKC and DGK and the regulation of each enzymatic activity by the binding were demonstrated using live-imaging techniques. PKC binds to DGK on the plasma membrane and phosphorylates DGK at S776and S779. The phosphorylated DGK is then activated and terminates PKC pathway by convert DG to PA on the plasma membrane. 3) Nuclear translocation of DGKgamma requires its Cl domain but not the kinase activity. Nuclear DGKgamma controls cell cycle. 4) Aggregation of mutant PKCgamma within cells may cause Spinocerebellar ataxia type 14 (SCA14).
    Competitive research funding

  • 齋藤 尚亮
    日本学術振興会, 科学研究費補助金／特定領域研究, 特定領域研究, 神戸大学, 2005 - 2005
    貪食細胞での微生物の殺菌は、Noxの細胞膜成分(cytochromeb558)と4つ細胞質成分(p47phox,p67phox,p40phox,Rac)により構成されるNADPHoxidase(Nox)に由来する活性酸素種により行われる。本研究は、この厳密に制御された活性酸素産生の分子機構を、機能蛋白質特にPKCおよびRacに焦点を当て、リアルタイムで可視化することにより解明することを目的とした。【今年度の成果】(1)Rac分子種の貪食IgGビーズへの集積度に違いがあることを見出した(Rac1>Rac3>Rac2)。この違いは、C末側の塩基性配列(PB)の違いに依存することを、イメージング解析により示した。また、protein-lipid overlay assayにより、この領域に脂質結合特異性があることを示した。さらに、Rac分子種のPBを入れ替えたキメラ蛋白を用いることにより、各分子種のPBの活性酸素産生に果たす重要性を明らかにした。また、Rac2の活性型変異体(Rac(Q61L))の集積を観察すると、小胞体などの細胞内器官の膜に局在し、貪食時にそれらの小器官膜が線状に貪胞膜へ癒合するのが観察された。(2)GFP融合PKCを貪食細胞に発現させIgGビーズを貧食させると、7種のPKC分子種の中beta I-PKCとepsilon-PKCのみが食胞に集積する。その結果、貧食時のepsilon-PKCの集積は、そのC1BドメインにPLC-gamma1経由で形成されるdiacylglycerol(DAG)が結合することにより起こることを明らかにした。【考察】(1)Rac1は貪食時にPBの高い正電荷とその脂質結合能を利用し食胞膜に集積する。Rac2はPBの正電荷を利用した弱い集積機序の他に、まず細胞内器官膜にターゲットし、この膜が貪胞膜に融合することを利用した、PBの正電荷に依存しない集積機序を持つことがわかった。これらのことから、3種すべてのRac分子種が活性酸素産生に関与する能力を有するが、マクロファージではRac1が、白血球ではRac2が優位に関与することが推察された。(2)同じC1BドメインをもつPKC分子種の中でも、特にepsilon-PKCが、貪食時食胞に強く集積することは、C1Bの脂質結合性の違いがPKCの分子種特異的シグナリングに重要な役割を果たしていることが推察された。
    Competitive research funding

  • 白井 康仁
    科学研究費補助金／特定領域研究, 2005
    Competitive research funding

  • ファゴサイトーシス時に働くシグナルカスケードのリアルタイム解析
    上山 健彦
    日本学術振興会, 科学研究費助成事業, 若手研究(B), 神戸大学, 2003 - 2004
    Fcγ receptorを介したファゴサイトーシス時におけるシグナルカスケードの解明を、PKC分子種、Diacylglycerol kinase (DGK)分子種にGFPを標織しmicrogliaに発現させ、IgGで標識したビーズを貪食させ、共焦点コンフォーカル顕微鏡下でリアルタイム観察することにより行った。 1,PKC分子種の中でβIPKCとεPKCのみが、食胞膜に限局して一時的に集積すること、この2種の集積様式が異なっていることをつきとめた。さらに、βIPKCの集積機序の詳細な解明に努め、βIPKCは食胞膜で限局的に産生されるdiacylglycerolとオシレーションを伴う細胞内のカルシウムの上昇により、オシレーションを伴って食胞膜に限局的に集積することがわかった。 2,βIPKCのファゴサイトーシス時における機能を解明するため、カルシウム依存性PKCの選択的阻害剤を用いて、Fcγ receptor刺激時の活性酸素産生を測定した。活性酸素の産生を(1)細胞外への活性酸素産生、(2)食胞内への活性酸素産生に分けて考えると、(1)はβIPKCに完全に依存したが、(2)は部分的に依存していた。 3,さらにPKCの機能を調節していると考えられるDGKのなかでカルシウム依存性のある分子種に注目したところ、DGKβのみがファゴサイトーシス時に集積した。活性酸素の産生は、DGK選択的阻害剤で処置すると、特に細胞外への活性酸素産生が著名に増加した。さらにこの阻害剤処置時のβIPKCの集積は著名に増強した。以上のことから、Fcγ receptorを介したファゴサイトーシスにおいて、細胞外への活性酸素の産生は、βIPKCにより制御されており、その調節をDGKβが行っていることがわかった。 4,さらにβIPKCの基質であるp47^も食胞膜に限局的に集積することをつきとめた。以上のことから、Fcγ receptorを介したファゴサイトーシス時における活性酸素の産生は、食胞で限局するように厳密に制御されていることが推測された。そこで電子顕微鏡を用いてFcγ receptorを介したファゴサイトーシス時における活性酸素の産生部位を検索したところ、食胞に限局していた。 以上の成果は、現在投稿中である。

TOP