Research activity information

• Mar. 2004 日本薬理学会, 日本薬理学会奨励賞, ライブイメージングを用いたプロテインキナーゼＣおよびジアシルグリセロールキナーゼの機能解析

  Yasuhito SHirai


• 2004 日本薬理学会, 薬理学会奨励賞, ライブイメージングを用いたプロテインキナーゼＣ及びジアシルグリセロールキナーゼの機能解析

  Shirai Yasuhito

• Tocotrienols activate diacylglycerol kinase a via 67 KDa laminin receptor

  Tomoka Namba, Daiki Hayashi, Itsuko Fukuda, Shuji Ueda, Yasuhito Shirai

  Corresponding, May 2025, Functional Food Science, 5(5) (5), 160 - 169, No password

  [Refereed]

  Scientific journal

  添付ファイルのURL


• Metabolites and Free Fatty Acids in Japanese Black Beef During Wet Aging.

  Shuji Ueda, Yuka Yoshida, Yuka Tateoka, Biniam Kebede, Masakazu Shinohara, Hiroki Nakanishi, Itsuko Fukuda, Yasuhito Shirai

  Background: Japanese Black beef is known for its high intramuscular fat content, an important factor in its distinctive Wagyu aroma. Wet aging, which involves vacuum-packing meat and storing it at low temperatures, enhances flavor, texture, and tenderness and is essential for maintaining and improving meat quality. In this study, changes in metabolites and lipid profiles were investigated during the wet aging of Japanese Black and Holstein beef. Methods/Results: Gas chromatography-mass spectrometry identified 113 metabolites in Japanese Black beef and 94 in Holstein beef, with significant increases in metabolites like aspartic acid and maleic acid over the aging period. Regarding lipid composition, total free fatty acids significantly increased with wet aging, with Japanese Black beef showing significantly higher concentrations of oleic and linoleic acids than Holstein beef. Additionally, lipid analysis by liquid chromatography-mass spectrometry revealed a reduction in specific phospholipids, particularly lysophosphatidylcholine (LPC) and lysophosphatidylethanolamine (LPE), with notable decreases in LPC (18:1), LPC (18:2), LPE (18:1), and LPE (18:2). Conclusions: These results suggest that wet aging influences the stability of membrane lipids, facilitating the degradation of phospholipids into free fatty acids, and improving the flavor of Japanese Black beef.

  Feb. 2025, Metabolites, 15(2) (2), English, International magazine

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Evaluating functionalities of food components by a model simulating human intestinal microbiota constructed at Kobe University.

  Ro Osawa, Itsuko Fukuda, Yasuhito Shirai

  In this era of pandemics, reducing the risk of lifestyle-related diseases (LRD) by functional foods is of paramount importance. The conventional process of functional food development almost invariably involves in vitro, animal, and human intervention trials, but differences in intestinal environments between humans and experimental animals make it difficult to develop functional foods that are truly effective in humans. Thus, it is necessary to construct a model that simulates the human intestinal environment to evaluate the functionality of any food component before subjecting it to a human intervention trial. In this review, we provide an overview of a model simulating human intestinal microbiota constructed at Kobe University and its use as a tool to identify food components that contribute to the prevention and treatment of LRD.

  Mar. 2024, Current opinion in biotechnology, 87, 103103 - 103103, English, International magazine

  Scientific journal


• New Implications of Metabolites and Free Fatty Acids in Quality Control of Crossbred Wagyu Beef during Wet Aging Cold Storage.

  Shuji Ueda, Yuka Yoshida, Biniam Kebede, Chiaki Kitamura, Ryo Sasaki, Masakazu Shinohara, Itsuko Fukuda, Yasuhito Shirai

  Efficient cold-chain delivery is essential for maintaining a sustainable global food supply. This study used metabolomic analysis to examine meat quality changes during the "wet aging" of crossbred Wagyu beef during cold storage. The longissimus thoracic (Loin) and adductor muscles (Round) of hybrid Wagyu beef, a cross between the Japanese Black and Holstein-Friesian breeds, were packaged in vacuum film and refrigerated for up to 40 days. Sensory evaluation indicated an increase in the umami and kokumi taste owing to wet aging. Comprehensive analysis using gas chromatography-mass spectrometry identified metabolite changes during wet aging. In the Loin, 94 metabolites increased, and 24 decreased; in the Round, 91 increased and 18 decreased. Metabolites contributing to the umami taste of the meat showed different profiles during wet aging. Glutamic acid increased in a cold storage-dependent manner, whereas creatinine and inosinic acid degraded rapidly even during cold storage. In terms of lipids, wet aging led to an increase in free fatty acids. In particular, linoleic acid, a polyunsaturated fatty acid, increased significantly among the free fatty acids. These results provide new insight into the effects of wet aging on Wagyu-type beef, emphasizing the role of free amino acids, organic acids, and free fatty acids generated during cold storage.

  Jan. 2024, Metabolites, 14(2) (2), English, International magazine

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Isolation and identification of hyaluronan-degrading bacteria from Japanese fecal microbiota.

  Hazuki Akazawa, Itsuko Fukuda, Haruna Kaneda, Shoichi Yoda, Mamoru Kimura, Ryohei Nomoto, Shuji Ueda, Yasuhito Shirai, Ro Osawa

  Hyaluronan (HA) is a high-molecular-weight glycosaminoglycan and widely distributed in all connective tissues and organs with diverse biological functions. HA has been increasingly used as dietary supplements targeted to joint and skin health for humans. We here first report isolation of bacteria from human feces that are capable of degrading HA to lower molecular weight HA oligosaccharides (oligo-HAs). The bacteria were successfully isolated via a selective enrichment method, in which the serially diluted feces of healthy Japanese donors were individually incubated in an enrichment medium containing HA, followed by the isolation of candidate strains from streaked HA-containing agar plates and selection of HA-degrading strains by measuring HA using an ELISA. Subsequent genomic and biochemical assays identified the strains as Bacteroides finegoldii, B. caccae, B. thetaiotaomicron, and Fusobacterium mortiferum. Furthermore, our HPLC analysis revealed that the strains degraded HA to oligo-HAs of various lengths. Subsequent quantitative PCR assay targeting the HA degrading bacteria showed that their distribution in the Japanese donors varied. The evidence suggests that dietary HA is degraded by the human gut microbiota with individual variation to oligo-HAs components, which are more absorbable than HA, thereby exerting its beneficial effects.

  2023, PloS one, 18(5) (5), e0284517, English, International magazine

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Vitamin E functions by association with a novel binding site on the 67 kDa laminin receptor activating diacylglycerol kinase.

  Daiki Hayashi, Varnavas D Mouchlis, Seika Okamoto, Tomoka Namba, Liuqing Wang, Sheng Li, Shuji Ueda, Minoru Yamanoue, Hirofumi Tachibana, Hiroyuki Arai, Hitoshi Ashida, Edward A Dennis, Yasuhito Shirai

  It is generally recognized that the main function of α-tocopherol (αToc), which is the most active form of vitamin E, is its antioxidant effect, while non-antioxidant effects have also been reported. We previously found that αToc ameliorates diabetic nephropathy via diacylglycerol kinase alpha (DGKα) activation in vivo, and the activation was not related to the antioxidant effect. However, the underlying mechanism of how αToc activates DGKα have been enigmatic. We report that the membrane-bound 67 kDa laminin receptor (67LR), which has previously been shown to serve as a receptor for epigallocatechin gallate (EGCG), also contains a novel binding site for vitamin E, and its association with Vitamin E mediates DGKα activation by αToc. We employed hydrogen-deuterium exchange mass spectrometry (HDX/MS) and molecular dynamics (MD) simulations to identify the specific binding site of αToc on the 67LR and discovered the conformation of the specific hydrophobic pocket that accommodates αToc. Also, HDX/MS and MD simulations demonstrated the detailed binding of EGCG to a water-exposed hydrophilic site on 67LR, while in contrast αToc binds to a distinct hydrophobic site. We demonstrated that 67LR triggers an important signaling pathway mediating non-antioxidant effects of αToc, such as DGKα activation. This is the first evidence demonstrating a membrane receptor for αToc and one of the underlying mechanisms of a non-antioxidant function for αToc.

  Dec. 2022, The Journal of nutritional biochemistry, 110, 109129 - 109129, English, International magazine

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• The Role of Diacylglycerol Kinase in the Amelioration of Diabetic Nephropathy.

  Daiki Hayashi, Yasuhito Shirai

  The drastic increase in the number of patients with diabetes and its complications is a global issue. Diabetic nephropathy, the leading cause of chronic kidney disease, significantly affects patients' quality of life and medical expenses. Furthermore, there are limited drugs for treating diabetic nephropathy patients. Impaired lipid signaling, especially abnormal protein kinase C (PKC) activation by de novo-synthesized diacylglycerol (DG) under high blood glucose, is one of the causes of diabetic nephropathy. DG kinase (DGK) is an enzyme that phosphorylates DG and generates phosphatidic acid, i.e., DGK can inhibit PKC activation under diabetic conditions. Indeed, it has been proven that DGK activation ameliorates diabetic nephropathy. In this review, we summarize the involvement of PKC and DGK in diabetic nephropathy as therapeutic targets, and its mechanisms, by referring to our recent study.

  Oct. 2022, Molecules (Basel, Switzerland), 27(20) (20), English, International magazine

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Polo-Like Kinase 2 Plays an Essential Role in Cytoprotection against MG132-Induced Proteasome Inhibition via Phosphorylation of Serine 19 in HSPB5.

  Shuji Ueda, Moeka Nishihara, Yuuki Hioka, Ken-Ichi Yoshino, Soichiro Yamada, Minoru Yamanoue, Yasuhito Shirai

  Protein homeostasis, including protein folding, refolding, and degradation, is thought to decline with aging. HSPB5 (also known as αB-crystallin) prevents target protein aggregation as a molecular chaperone and exhibits a cytoprotective function against various cell stresses. To elucidate the effect of HSPB5 on endoplasmic reticulum (ER) stress, we searched for novel binding proteins of HSPB5 using the proximity-dependent biotin labeling method. Proteins presumed to interact with HSPB5 in cells treated with the proteasome inhibitor MG132 were identified by a reversible biotin-binding capacity method combining tamavidin2-REV magnetic beads and mass spectrometry. We discovered a new binding protein for HSPB5, polo-like kinase 2 (PLK2), which is an apoptosis-related enzyme. The expression of PLK2 was upregulated by MG132 treatment, and it was co-localized with HSPB5 near the ER in L6 muscle cells. Inhibition of PLK2 decreased ER stress-induced phosphorylation of serine 19 in HSPB5 and increased apoptosis by activation of caspase 3 under ER stress. Overexpression of HSPB5 (WT) suppressed the ER stress-induced caspase 3 activity, but this was not observed with phospho-deficient HSPB5 (3A) mutants. These results clarify the role of HSPB5 phosphorylation during ER stress and suggest that the PLK2/HSPB5 pathway plays an essential role in cytoprotection against proteasome inhibition-induced ER stress.

  Sep. 2022, International journal of molecular sciences, 23(19) (19), English, International magazine

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Application of Mass Spectrometry for Determining the Geographic Production Area of Wagyu Beef

  Shuji Ueda, Yasuharu Takashima, Yunosuke Gotou, Ryo Sasaki, Rio Nakabayashi, Takeshi Suzuki, Shinji Sasazaki, Ituko Fukuda, Biniam Kebede, Yuki Kadowaki, Maiko Tamura, Hiroki Nakanishi, Yasuhito Shirai

  Japanese Black cattle (Japanese Wagyu) beef is attracting attention for its aroma and marbling, and its handling is increasing worldwide. Here, we focused on the origin discrimination of Wagyu beef and analyzed the nutritional components of Japanese Wagyu (produced in multiple prefectures of Japan), Hybrid Wagyu (a cross between Angus and Wagyu cattle born in Australia and transported to Japan), and Australian Wagyu beef using mass spectrometry (MS). Triple-quadrupole liquid chromatography–MS was used to clarify the molecular species of lipids in Wagyu beef. Fourteen classes of lipids were separated, and 128 different triacylglycerides (TGs) were detected. A simple comparative analysis of these TGs using high-performance liquid chromatography revealed significantly higher levels of triolein (C18:1/C18:1/C18:1; abbreviated OOO) and C18:1/C18:1/C16:1 (OOPo) in Japanese Wagyu. Wagyu elements beef were comprehensively analyzed using inductively coupled plasma (ICP)–MS and ICP–optical emission spectrometry. We found significant differences in the rubidium, cesium, and lithium levels of Japanese and Australian Wagyu beef. On comparing metabolites using gas chromatography–MS, we identified significant differences in the levels of amino acids and other components of the Japanese and Australian Wagyu beef. These results suggest the possibility of determining the origin of Wagyu cattle breeds using MS and genetic discrimination.

  MDPI AG, Aug. 2022, Metabolites, 12(9) (9), 777 - 777

  [Refereed]

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Almond Skin Polyphenol Extracts Stimulate the Activation of Diacylglycerol Kinase alpha via a 67 kDa Laminin Receptor

  Boonyaporn Chinthammit, Seika Okamoto, Yasuhito Shirai

  Background: Almond skins are the byproduct of the almond industry that are rich in dietary fibers and polyphenols. Recently, there has been an increasing interest in utilizing the phenolic compounds from almond skins for their functional benefits. Galloylated catechins activate diacylglycerol kinase α (DGKα) which is involved in the amelioration of diabetic nephropathy. Therefore, in this study, we investigated whether almond skin polyphenol extracts can also induce the activation of DGKα. Methods: Phenolic contents in the almond skin polyphenol extracts were identified by liquid chromatography (LC)-time of flight mass spectrometry (TOFMS). Using confocal microscopy, the translocation of green fluorescent protein (GFP)-DGKα to the cell membrane was observed upon stimulation with almond skin polyphenol extracts. To check the involvement of 67 kDa laminin receptor (67LR), pre-treatment of anti-67LR antibody was used.Results: We identified that naringenin and flavanone (2,3-dihydroflavone), which are among the phenolic contents in the almond skin polyphenol extracts identified, can also induce the activation of DGKα. In addition, we also investigated whether the pathway involves the same receptor as that of epigallocatechin-3-gallate (EGCg); the 67LR. Naringenin stimulated through the 67LR, while flavanone mainly used the 67LR-independent pathways.Conclusion: These findings were additional function of almond skin polyphenol extracts and may implicate the benefits of the intake of nuts in daily diets. Polyphenols can be extracted from almond skins as an inexpensive source. The various health benefits of polyphenols can be applied to functional foods and supplements.Keywords: flavonoid, lipid kinase, imaging, catechin, vitamin E, diabetic nephropathy

  Corresponding, Functional Food Center, Apr. 2022, Functional Foods in Health and Disease, 12(4) (4), 151 - 160

  [Refereed]

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Production of Hydroxy Fatty Acids, Precursors of γ-Hexalactone, Contributes to the Characteristic Sweet Aroma of Beef.

  Shuji Ueda, Mana Hosoda, Kumi Kasamatsu, Masahiro Horiuchi, Rio Nakabayashi, Bubwoong Kang, Masakazu Shinohara, Hiroki Nakanishi, Takayo Ohto-Nakanishi, Minoru Yamanoue, Yasuhito Shirai

  Aroma is an essential factor for meat quality. The meat of Japanese Black cattle exhibits fine marbling and a rich and sweet aroma with a characteristic lactone composition. The mechanism of lactone formation associated with beef aroma has not been elucidated. In this study, we examined the precursors of γ-hexalactone, an indicator of the sweet aroma of beef and identified the mechanism underlying γ-hexalactone production. A low-temperature vacuum system was used to prepare beef tallow from Japanese Black cattle and Holstein cattle. The odor components were identified using headspace-gas chromatography. The analysis revealed that γ-hexalactone, γ-dodecalactone, δ-tetradecalactone, and δ-hexadecalactone were present as sweet aroma components of beef tallow prepared from marbling and muscle. Since we previously reported that γ-hexalactone formation correlates with linoleic acid content in beef, we analyzed ten oxidized fatty acids derived from linoleic acid by liquid chromatography-triple quadrupole mass spectrometry and detected two hydroxy-octadecadienoic acids (9S-HODE and 13S-HODE) in beef tallow. Significant differences in arachidonic acid 15-lipoxygenase and cyclooxygenase protein expression levels among subcutaneous fat, intramuscular fat, and muscle tissue were observed. Our results suggest that the combination of linoleic acid and the expression of lipid oxidase derived from beef muscle and intramuscular fat produce hydroxy fatty acids that result in a sweet aroma.

  Apr. 2022, Metabolites, 12(4) (4), English, International magazine

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Combination therapy for hepatocellular carcinoma with diacylglycerol kinase alpha inhibition and anti-programmed cell death-1 ligand blockade.

  Naoki Okada, Ko Sugiyama, Shunsuke Shichi, Yasuhito Shirai, Kaoru Goto, Fumio Sakane, Hidemitsu Kitamura, Akinobu Taketomi

  Activation of diacylglycerol kinase alpha (DGKα) augments proliferation and suppresses apoptosis of cancer cells and induces T lymphocyte anergy. We investigated the dual effects of DGKα inhibition on tumorigenesis and anti-tumor immunity with the aim of establishing a novel therapeutic strategy for cancer. We examined the effects of a DGKα inhibitor (DGKAI) on liver cancer cell proliferation and cytokine production by immune cells in vitro and on tumorigenesis and host immunity in a hepatocellular carcinoma (HCC) mouse model. Oral DGKAI significantly suppressed tumor growth and prolonged survival in model mice. Tumor infiltration of T cells and dendritic cells was also enhanced in mice treated with DGKAI, and the production of cytokines and cytotoxic molecules by CD4+ and CD8+ T cells was increased. Depletion of CD8+ T cells reduced the effect of DGKAI. Furthermore, interferon-γ stimulation augmented the expression of programmed cell death-1 ligand (PD-L1) on cancer cells, and DGKAI plus an anti-PD-L1 antibody strongly suppressed the tumor growth. These results suggest that DGKα inhibition may be a promising new treatment strategy for HCC.

  Apr. 2022, Cancer immunology, immunotherapy : CII, 71(4) (4), 889 - 903, English, International magazine

  [Refereed]

  Scientific journal


• RalA, PLD and mTORC1 Are Required for Kinase-Independent Pathways in DGKβ-Induced Neurite Outgrowth.

  Takuya Kano, Ryosuke Tsumagari, Akio Nakashima, Ushio Kikkawa, Shuji Ueda, Minoru Yamanoue, Nobuyuki Takei, Yasuhito Shirai

  Diacylglycerol kinase β (DGKβ) is an enzyme that converts diacylglycerol to phosphatidic acid and is mainly expressed in the cerebral cortex, hippocampus and striatum. We previously reported that DGKβ induces neurite outgrowth and spinogenesis, contributing to higher brain functions, including emotion and memory. To elucidate the mechanisms involved in neuronal development by DGKβ, we investigated the importance of DGKβ activity in the induction of neurite outgrowth using human neuroblastoma SH-SY5Y cells. Interestingly, both wild-type DGKβ and the kinase-negative (KN) mutant partially induced neurite outgrowth, and these functions shared a common pathway via the activation of mammalian target of rapamycin complex 1 (mTORC1). In addition, we found that DGKβ interacted with the small GTPase RalA and that siRNA against RalA and phospholipase D (PLD) inhibitor treatments abolished DGKβKN-induced neurite outgrowth. These results indicate that binding of RalA and activation of PLD and mTORC1 are involved in DGKβKN-induced neurite outgrowth. Taken together with our previous reports, mTORC1 is a key molecule in both kinase-dependent and kinase-independent pathways of DGKβ-mediated neurite outgrowth, which is important for higher brain functions.

  Dec. 2021, Biomolecules, 11(12) (12), English, International magazine

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Milk oligosaccharide-mediated cross-feeding between Enterococcus gallinarum and lactobacilli in the gut microbiota of infant rats

  Saki MATSUI, Hazuki AKAZAWA, Yuji TSUJIKAWA, Itsuko FUKUDA, Yoshihiro SUZUKI, Yuji YAMAMOTO, Takao MUKAI, Yasuhito SHIRAI, Ro OSAWA

  We investigated bacteria that have a nutritional symbiotic relationship with respect to milk oligosaccharides in gut microbiota of suckling rats, with specific reference to sialyllactose (SL) degrading Enterococcus gallinarum. Our next generation sequencing analysis of the colonic contents of 12-day-old suckling rats revealed that almost half of the bacteria in the microbiota belonged to the Lactobacillaceae family. Major Lactobacillus species in the contents were identified as L. johnsonii, L. murinus, and L. reuteri. We then monitored changes in numbers of the above Lactobacillus species, E. gallinarum, and the bacteria belonging to the family Enterobacteriaceae (i.e., enterobacteria) in the colonic contents of infant rats at 7, 12, 21, 28, and 35 days of age by using real-time PCR assays targeting these bacterial groups. The 7-day-old infant rats had a gut microbiota in which enterobacteria were predominant. Such dominance was replaced by L. johnsonii and the concomitant E. gallinarum markedly increased in those of 12 and 21 days of ages. During this period, the number of enterobacteria declined dramatically, but that of L. reuteri surged dramatically. Our separate in vitro experiment showed that supplementation of culture media with SL promoted the growth of L. johnsonii and E. gallinarum, with marked production of lactic acid. These findings revealed possible milk oligosaccharide-mediated cross-feeding between E. gallinarum and L. johnsonii, with the former degrading SL to release lactose to be utilized by the latter.

  BMFH Press, Oct. 2021, Bioscience of Microbiota, Food and Health, 40(4) (4), 204 - 211, English, Domestic magazine

  [Refereed]

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Gene Expression Analysis Provides New Insights into the Mechanism of Intramuscular Fat Formation in Japanese Black Cattle

  Shuji Ueda, Mana Hosoda, Ken-ichi Yoshino, Minoru Yamanoue, Yasuhito Shirai

  Japanese Black cattle (Japanese Wagyu) have a unique phenotype in which ectopic intramuscular fat accumulates in skeletal muscle, producing finely marbled beef. However, the mechanism of intramuscular fat formation in Japanese Black cattle remains unclear. To investigate the key genes involved in intramuscular fat accumulation, we comprehensively analyzed mRNA levels in subcutaneous and intramuscular fat tissues using RNA sequence (RNA-seq) analysis, which detected 27,606 genes. We identified eight key genes, namely carboxypeptidase E, tenascin C, transgelin, collagen type IV alpha 5 (COL4A5), cysteine and glycine-rich protein 2, PDZ, and LIM domain 3, phosphatase 1 regulatory inhibitor subunit 14A, and regulator of calcineurin 2. These genes were highly and specifically expressed in intramuscular fat tissue. Immunohistochemical analysis revealed a collagen network, including COL4A5, in the basement membrane around the intramuscular fat tissue. Moreover, pathway analysis revealed that, in intramuscular fat tissue, differentially expressed genes are related to cell adhesion, proliferation, and cancer pathways. Furthermore, pathway analysis showed that the transforming growth factor-β (TGF-β) and small GTPases regulators RASGRP3, ARHGEF26, ARHGAP10, ARHGAP24, and DLC were upregulated in intramuscular fat. Our study suggests that these genes are involved in intramuscular fat formation in Japanese Black cattle.

  MDPI AG, Jul. 2021, Genes, 12(8) (8), 1107 - 1107

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Exploring the Lipids Involved in the Formation of Characteristic Lactones in Japanese Black Cattle

  Shuji Ueda, Ryo Sasaki, Rio Nakabayashi, Minoru Yamanoue, Yasuhito Sirai, Eiji Iwamoto

  The meat from Japanese Black cattle (Japanese Wagyu) is finely marbled and exhibits a rich and sweet aroma known as Wagyu beef aroma. To clarify the key metabolites involved in the aroma, we analyzed the correlation between lactone and lipid composition in Japanese Black cattle. Using gas chromatography-olfactometry, we identified 39 characteristic odorants of the intermuscular fat. Seven characteristic lactones considered to be involved in Wagyu beef aroma were quantified and compared in the marbled area and intermuscular fat using a stable isotope dilution assay. Among them, γ-hexalactone was the only lactone whose level was significantly higher in the marbled area. To explore the lipid species involved in lactone formation, we analyzed samples with different aroma characteristics. Liquid chromatography-mass spectrometry revealed eight lipid classes and showed significant differences in triacylglycerides (TAGs). To determine the molecular species of TAGs, we performed high-performance liquid chromatography analysis and identified 14 TAG species. However, these analyses showed that seven lactones had a low correlation with the TAGs. However, γ-hexalactone showed a positive correlation with linoleic acid. This study suggests that lipid composition affects the characteristic lactone profile involved in the Wagyu beef aroma.

  MDPI AG, Mar. 2021, Metabolites, 11(4) (4), 203

  [Refereed]

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Motor Dyscoordination and Alteration of Functional Correlation Between DGKγ and PKCγ in Senescence-Accelerated Mouse Prone 8 (SAMP8)

  Ryosuke Tsumagari, Kenta Maruo, Takaaki Nakao, Shuji Ueda, Minoru Yamanoue, Yasuhito Shirai

  Senescence-accelerated mouse prone 8 (SAMP8) is an animal model of age-related central nervous system (CNS) disorders. Although SAMP8 shows deficits in learning, memory, and emotion, its motor coordination has not been clarified. We have recently reported that DGKγ-regulated PKCγ activity is important for cerebellar motor coordination. However, involvement of the functional correlation between the kinases in age-related motor dyscoordination still remains unknown. Therefore, we have investigated the motor coordination in SAMP8 and involvement of the functional correlation between DGKγ and PKCγ in the age-related motor dyscoordination. Although 6 weeks old SAMP8 showed equivalent motor coordination with control mice (SAMR1) in the rotarod test, 24 weeks old SAMP8 exhibited significantly less latency in the rotarod test and more frequent slips in the beam test compared to the age-matched SAMR1. Furthermore, 24 weeks old SAMP8 showed the higher locomotor activity in open field test and Y-maze test. Western blotting revealed that DGKγ expression decreased in the cerebellum of 24 weeks old SAMP8, while PKCγ was upregulated. These results suggest that SAMP8 is a useful model of age-related motor dysfunction and that the DGKγ-regulated PKCγ activity is involved in the age-related motor dyscoordination.

  Frontiers Media SA, Jan. 2021, Frontiers in Aging Neuroscience, 13

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Exploring the Characteristic Aroma of Beef from Japanese Black Cattle (Japanese Wagyu) via Sensory Evaluation and Gas Chromatography-Olfactometry

  Shuji Ueda, Minoru Yamanoue, Yasuhito Sirai, Eiji Iwamoto

  Beef from Japanese Black cattle (Japanese Wagyu) is renowned for its flavor characteristics. To clarify the key metabolites contributing to this rich and sweet aroma of beef, an omics analysis combined with GC-olfactometry (GC-O) and metabolomics analysis with gas chromatography–mass spectrometry (GC-MS) were applied. GC-O analysis identified 39 odor-active odorants from the volatile fraction of boiled beef distilled by solvent-assisted flavor evaporation. Eight odorants predicted to contribute to Wagyu beef aroma were compared between Japanese Black cattle and Holstein cattle using a stable isotope dilution assay with GC–tandem quadrupole mass spectrometry. By correlating the sensory evaluation values of retronasal aroma, γ-hexalactone, γ-decalactone, and γ-undecalactone showed a high correlation with the Wagyu beef aroma. Metabolomics data revealed a high correlation between the amounts of odorants and multiple metabolites, such as glutamine, decanoic acid, lactic acid, and phosphoric acid. These results provide useful information for assessing the aroma and quality of beef.

  MDPI AG, Jan. 2021, Metabolites, 11(1) (1), 56

  [Refereed]

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Protective effect of Zingiber zerumbet Smith extract on thermotolerance

  Yasuhito Shirai, Hisakazu Kobayashi, Shuji Ueda, Yun Sang Soon, Akiho Kushiya, Ryosuke Tsumagari, Minoru Yamanoue

  Background: Global warming causes severe heat conditions. Heat stress contributes to higher morbidity of heatstroke in human and mortality in livestock. To protect them from heat stress, thermotolerance mechanisms were widely studied, and some studies suggest relationship between heat shock proteins (HSPs) and thermotolerance. HSPs were not induced by only heat shock but also some stimulations including bioactive compounds from plants. Zingiber zerumbet is a perennial herb found in many tropical countries, including Thailand. The rhizome of Zingiber zerumbet contains zerumbone that is a bioactive compound to induce HSPs expression in animal cells.Objective: To prevent higher morbidity of heatstroke in human and mortality in livestock by the heat stress, we investigated the effect of zerumbone, the extract of Zingiber zerumbet Smith, on thermotolerance, using a cell line and mice.Methods: The murine liver hepatoma cell line, Hepa1c1c7 cells, were incubated in medium supplemented with extract from rhizome of Zingiber zerumbet Simith containing zerumbone, and then the expression of heat shock proteins (HSP) 40, 70 and 90 were investigated by western blotting. Furthermore, we established the evaluation system of thermotolerance using mice, and studied the effect of the extract on the growth rate of mice under the heat shock treatment. Briefly, 4 weeks old C57BL6 mice were fed that with the extract (or vehicle) for a week before the first heat shock treatment (38 °C for an hour). Before and after five days heat treatment, body weights were measured. The protein expressions of heat shock proteins in liver were measured by western blotting using HSPs antibodies.Results: The extract of Zingiber zerumbet rhizome, equivalent to 50 μM zerumbone, significantly increased the expression of heat shock proteins (HSP40, HSP70, HSP90). The growth rate of the mice under the heat treatment were lower than control. The feeding with the extract containing 25 ppm zerumbone have significantly attenuated the decline of the growth rate led by the heat treatment, whereas there was little effect on mouse growth rate grown under normal conditions. The protein expression of HSP70 in the liver of zerumbone-fed mice was upregulated compared with control mice, equivalent to heat treatment without zerumbone. On the other hand, both treatments of zerumbone and heat resulted in highest HSP70 expression among four groups.Conclusion: Our study demonstrated that oral administration of the extract of Zingiber zerumbet Smith led to the attenuation of decline of growth rate induced by heat treatment. HSP70 expression in murine liver was enhanced by either feeding the extract or heat treatment. More interestingly, HSP70 expression was further enhanced by both treatments of zerumbone and heat. These results suggested that zerumbone may contribute to thermotolerance via, at least, HSP70 expression.Keywords: Zingiber zerumbet, thermotolerance, heat shock protein

  Functional Food Center, Jan. 2021, Bioactive Compounds in Health and Disease, 4(1) (1), 1 - 1

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• The mechanisms of ameliorating effect of a green tea polyphenol on diabetic nephropathy based on diacylglycerol kinase α

  Daiki Hayashi, Liuqing Wang, Shuji Ueda, Minoru Yamanoue, Hitoshi Ashida, Yasuhito Shirai

  Significant efforts have been made to ameliorate diabetic nephropathy (DN) by inhibiting protein kinase C. However, these efforts have not been successful in human trials, suggesting that novel therapeutic strategies are required. Thus far, it has been reported that green tea polyphenol epigallocatechin gallate (EGCg) improved albuminuria in DN in a human trial. Our previous study revealed that activation of diacylglycerol kinase α (DGKα) plays a crucial role in the amelioration of DN and that EGCg activates DGKα. Here, we investigated whether and how DGKα contributes to the amelioration of DN upon stimulation by EGCg by using streptozotocin-induced type 1 diabetic model mice. Our results revealed that EGCg ameliorated albuminuria in DN through DGKα in vivo, and methylated EGCg, which has higher absorption in the plasma improved albuminuria in DN effectively. Additionally, we showed that c-Src mediated EGCg-induced DGKα translocation and colocalized with the 67 kDa laminin receptor, which is an EGCg receptor. Furthermore, EGCg attenuated the loss of podocytes in DN by preventing a decrease in focal adhesion under high glucose conditions. Our results indicate that the DGKα pathway is an attractive therapeutic target and that activating this pathway is a novel strategy for treating DN.

  Springer Science and Business Media LLC, Dec. 2020, Scientific Reports, 10(1) (1)

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Precise Regulation of the Basal PKCγ Activity by DGKγ Is Crucial for Motor Coordination.

  Ryosuke Tsumagari, Kenta Maruo, Sho Kakizawa, Shuji Ueda, Minoru Yamanoue, Hiromitsu Saito, Noboru Suzuki, Yasuhito Shirai

  Diacylglycerol kinase γ (DGKγ) is a lipid kinase to convert diacylglycerol (DG) to phosphatidic acid (PA) and indirectly regulates protein kinase C γ (PKCγ) activity. We previously reported that the basal PKCγ upregulation impairs cerebellar long-term depression (LTD) in the conventional DGKγ knockout (KO) mice. However, the precise mechanism in impaired cerebellar LTD by upregulated PKCγ has not been clearly understood. Therefore, we first produced Purkinje cell-specific DGKγ KO (tm1d) mice to investigate the specific function of DGKγ in Purkinje cells and confirmed that tm1d mice showed cerebellar motor dysfunction in the rotarod and beam tests, and the basal PKCγ upregulation but not PKCα in the cerebellum of tm1d mice. Then, the LTD-induced chemical stimulation, K-glu (50 mM KCl + 100 µM, did not induce phosphorylation of PKCα and dissociation of GluR2 and glutamate receptor interacting protein (GRIP) in the acute cerebellar slices of tm1d mice. Furthermore, treatment with the PKCγ inhibitor, scutellarin, rescued cerebellar LTD, with the phosphorylation of PKCα and the dissociation of GluR2 and GRIP. In addition, nonselective transient receptor potential cation channel type 3 (TRPC3) was negatively regulated by upregulated PKCγ. These results demonstrated that DGKγ contributes to cerebellar LTD by regulation of the basal PKCγ activity.

  Oct. 2020, International journal of molecular sciences, 21(21) (21), English, International magazine

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• 67kDaラミニンレセプターはビタミンEとの結合によりパルミトイル化修飾を受ける

  岡本 聖香, 林 大輝, 足立 直子, Varnavas Mouchlis, 上田 修司, 山之上 稔, Dennis Edward, 斎藤 尚亮, 伊藤 俊樹, 白井 康仁

  (公社)日本生化学会, Sep. 2020, 日本生化学会大会プログラム・講演要旨集, 93回, [1Z09 - 161)], Japanese


• mTORC1 is involved in DGKβ-induced neurite outgrowth and spinogenesis.

  Hiroko Nakai, Ryosuke Tsumagari, Kenta Maruo, Akio Nakashima, Ushio Kikkawa, Shuji Ueda, Minoru Yamanoue, Naoaki Saito, Nobuyuki Takei, Yasuhito Shirai

  Diacylglycerol kinase β (DGKβ) is an enzyme converting DG to phosphatidic acid (PA) and is specifically expressed in neurons, especially those in the cerebral cortex, hippocampus and striatum. We previously reported that DGKβ induces neurite outgrowth and spinogenesis, contributing to higher brain function including emotion and memory, and plasma membrane localization of DGKβ via the C1 domain and a cluster of basic amino acids at the C-terminus is necessary for its function. To clarify the mechanisms involved in neuronal development by DGKβ, we investigated whether DGKβ activity induces neurite outgrowth using human neuroblastoma SH-SY5Y cells. DGKβ induced neurite outgrowth by activation of mammalian target of rapamycin complex 1 (mTORC1) through a kinase-dependent pathway. In addition, in primary cultured cortical and hippocampal neurons, inhibition of mTORC1 abolished DGKβ induced-neurite outgrowth, branching and spinogenesis. These results indicated that DGKβ induces neurite outgrowth and spinogenesis by activating mTORC1 in a kinase-dependent pathway.

  Mar. 2020, Neurochemistry international, 134, 104645 - 104645, English, International magazine

  [Refereed]


• Comparative Study of the Effects of Different Pretreatment Procedures on Beef Taste-traits Using an Electronic Taste Sensing System

  Yanan Zhao, Masahiro Nishida, Shuji Ueda, Yasuhito Shirai, Masaaki Habara, Hidekazu Ikezaki, Minoru Yamanoue

  Japanese Society for Food Science and Technology, 2020, Food Science and Technology Research, 26(3) (3), 329 - 338

  Scientific journal


• Dgkg knock-out mice show impairments in cerebellar motor coordination, Ltd, and the dendritic development of purkinje cells through the activation of pkcg

  Ryosuke Tsumagari, Sho Kakizawa, Sakiko Kikunaga, Yoshitaka Fujihara, Shuji Ueda, Minoru Yamanoue, Naoaki Saito, Masahito Ikawa, Yasuhito Shirai

  2020, eNeuro, 7(2) (2)

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Screening of subtype-specific activators and inhibitors for diacylglycerol kinase.

  Hayashi D, Tsumagari R, Liu K, Ueda S, Yamanoue M, Sakane F, Shirai Y

  Jun. 2019, Journal of biochemistry, 165(6) (6), 517 - 522, English

  [Refereed]

  Scientific journal


• DzMab-1: Anti-Human Diacylglycerol Kinaseζ Monoclonal Antibody for Immunocytochemistry.

  Nakano T, Ogasawara S, Tanaka T, Hozumi Y, Sano M, Sayama Y, Yamada S, Shirai Y, Kaneko MK, Kato Y, Goto K

  2019, Monoclonal antibodies in immunodiagnosis and immunotherapy, 38, 179 - 182, English

  [Refereed]

  Scientific journal


• Comparative metabolomics of Japanese Black cattle beef and other meats using gas chromatography–mass spectrometry

  UEDA SYUJI, IWAMOTO EIJI, KATO YOSHIKI, SHINOHARA MASAKAZU, SHIRAI YASUHITO, YAMANOUE MINORU

  Progress in metabolomic analysis now allows the evaluation of food quality. This study aims to identify the metabolites in meat from livestock using a metabolomic approach. Using gas chromatography-mass spectrometry (GC/MS), many metabolites were reproducibly detected in meats, and distinct differences between livestock species (cattle, pigs, and chickens) were indicated. A comparison of metabolites between tissues types (muscle, intramuscular fat, and intermuscular fat) in marbled beef of Japanese Black cattle revealed that most metabolites are abundant in the muscle tissue. Several metabolites (medium-chain fatty acids, etc.) involved in triacylglycerol synthesis were uniquely detected in fat tissue. Additionally, the results of multivariate analysis suggest that GC/MS analysis of metabolites can distinguish between cattle breeds. These results provide useful information for the analysis of meat quality using GC/MS-based metabolomic analysis.ABBREVIATIONS: GC/MS: gas chromatography-mass spectrometry; NMR: nuclear magnetic resonance; MS: mass spectrometry; IS: 2-isopropylmalic acid; MSTFA: N-Methyl-N-trimethylsilyltrifluoroacetamide; CV: coefficient of variation; TBS: Tris-buffered saline; MHC: myosin fast type; PCA: principal component analysis; OPLS-DA: orthogonal partial least-squares discriminant analysis; O2PLS: two-way orthogonal partial least-squares.

  Japan Society for Bioscience, Biotechnology, and Agrochemistry, Jan. 2019, Bioscience, Biotechnology, and Biochemistry, 83(1) (1), 137 - 147, English, International magazine

  [Refereed]

  Scientific journal


• Lentinan Exerts its Anti-Inflammatory Activity by Suppressing TNFR1 Transfer to the Surface of Intestinal Epithelial Cells through Dectin-1 in an in vitro and mice model

  SAKAGUCHI KANA, SHIRAI YASUHITO, ITOH TOSHIKI, MIZUNO MASASHI

  Dec. 2018, Immunome Research, (14) (14), 165, English

  [Refereed][Invited]

  Scientific journal


• DgMab-6: Antihuman DGKγ Monoclonal Antibody for Immunocytochemistry.

  Tomoyuki Nakano, Satoshi Ogasawara, Toshiaki Tanaka, Yasukazu Hozumi, Atsumi Yamaki, Fumio Sakane, Yasuhito Shirai, Takuro Nakamura, Miyuki Yanaka, Shinji Yamada, Mika K Kaneko, Yukinari Kato, Kaoru Goto

  Diacylglycerol kinase (DGK) phosphorylates diacylglycerol (DG) to produce phosphatidic acid (PA). Since both DG and PA serve as lipidic second messengers, DGK plays a pivotal role in regulating the balance of two signaling pathways mediated by DG and PA in cellular functions. Reportedly, DGKγ, one of the 10 mammalian DGK isozymes, is involved in leukemic cell differentiation, mast cell function, and membrane traffic. Transfection studies using tagged expression vectors and immunohistochemistry on rat tissues revealed that DGKγ localizes to the cytoplasm, plasma membrane, and Golgi apparatus. However, a limited number of studies reported the detailed localization of native protein of DGKγ in human tissues and cells. In this study, we developed a novel anti-DGKγ monoclonal antibody, DgMab-6, which is very useful in immunocytochemistry of human cultured cells.

  Nov. 2018, Monoclonal antibodies in immunodiagnosis and immunotherapy, 37(5) (5), 229 - 232, English, International magazine

  [Refereed]

  Scientific journal


• Comparative metabolomics of Japanese Black cattle beef and other meats using gas chromatography-mass spectrometry

  Ueda S, Iwamoto E, Kato Y, Shinohara M, SHIRAI YASUHITO, Yamanoue M

  Oct. 2018, Biosci Biotechnol Biochem, 18, 1 - 11, English

  [Refereed]

  Scientific journal


• RhoA結合蛋白質として同定されたSTE20 like kinaseの筋細胞の肥大化における役割

  櫛谷 晃帆, 上田 修司, 前田 愛実, 加藤 良毅, 吉野 健一, 竹内 敦子, 山之上 稔, 白井 康仁

  (公社)日本生化学会, Sep. 2018, 日本生化学会大会プログラム・講演要旨集, 91回, [1T14e - 370)], Japanese


• [Importance of diacylglycerol signaling in cerebellar motor coordination].

  Ryosuke Tsumagari, Yasuhito Shirai

  Brain can be roughly divided into two parts, cerebrum and cerebellum. Cerebrum controls higher brain functions including memory, emotion and cognition, while cerebellum is important for motor coordination. The only output neuron in cerebellum, Purkinje cell, regulates long term depression (LTD). LTD and morphology of Purkinje cells are important for motor function. So far, disorder of protein kinase C (PKC) α and γ, which are expressed in Purkinje cells, impaired LTD, morphology of Purkinje cells and motor coordination. Diacylglycerol kinase (DGK) γ phosphorylates diacylglycerol (DG) and is abundantly expressed in Purkinje cells. In other words, DGKγ can attenuate PKC activity by reducing amount of DG and may contribute to motor coordination. However, its physiological role has not been elucidated. Therefore, we developed DGKγ knockout (KO) mice and investigated their LTD, morphology of Purkinje cells, and cerebellar motor coordination. We found that cerebellar motor coordination and LTD were impaired in the DGKγ KO mice and the morphology of Purkinje cells from DGKγ KO mice was significantly retracted. Interestingly, abnormal activation of PKCγ was involved in impairment of the morphology of Purkinje cells from DGKγ KO mice. These results indicated that DGKγ was involved in cerebellar LTD and morphology of Purkinje cells, and DG signaling is important for cerebellar motor coordination.

  2018, Nihon yakurigaku zasshi. Folia pharmacologica Japonica, 152(2) (2), 90 - 93, Japanese, Domestic magazine

  Scientific journal


• Epigallocatechin gallate induces GLUT4 translocation in skeletal muscle through both PI3K- and AMPK-dependent pathways.

  UEDA-WAKAGI Manabu, HAYASHIBARA Kaori, NAGANO Tomoya, IKEDA Masaki, YUAN Sihao, UEDA Shuji, SHIRAI Yasuhito, YOSHIDA Ken-ichi, ASHIDA Hitoshi

  Our previous report demonstrated that epigallocatechin gallate (EGCg) promotes translocation of glucose transporter 4 (GLUT4) in skeletal muscle. In this study, we investigated the molecular mechanism of GLUT4 translocation by EGCg at the physiological concentration range. In L6 cells, EGCg induced phosphorylation of phosphatidylinositide 3'-kinase (PI3K) and downstream protein kinase C (PKC) λ/ξ without affecting the phosphorylation of insulin receptor and Akt. EGCg-induced GLUT4 translocation was suppressed by RNA interference-mediated knockdown of PI3K and treatment with PKC inhibitor Go6983. Moreover, EGCg increased Rac1 activity and actin remodelling as downstream events of PKCλ/ξ. These results indicate that EGCg induced GLUT4 translocation through a PI3K-dependent pathway, but its mode of action differed from that of insulin. EGCg also induced GLUT4 translocation through a 5'-adenosine monophosphate-activated protein kinase (AMPK)-dependent pathway. 67 kDa laminin receptor, which is a target molecule of EGCg, was not involved in EGCg-induced glucose uptake in L6 cells. The oral administration of EGCg suppressed postprandial hyperglycaemia accompanied by GLUT4 translocation through both PI3K- and AMPK-dependent pathways, and promoted glycogen accumulation in skeletal muscle of ICR mice. EGCg promotes GLUT4 translocation through both PI3K- and AMPK-dependent pathways and glycogen accumulation in skeletal muscle.

  2018, Food and Function, 9(8) (8), 4223 - 4233, English, International magazine

  [Refereed]

  Scientific journal


• Amelioration of diabetic nephropathy by oral administration of d-α-Tocopherol and its mechanisms

  Daiki Hayashi, Shuji Ueda, Minoru Yamanoue, Hitoshi Ashida, Yasuhito Shirai

  Japan Society for Bioscience Biotechnology and Agrochemistry, 2018, Bioscience, Biotechnology and Biochemistry, 82(1) (1), 65 - 73, English

  [Refereed]

  Scientific journal


• Diacylglycerol Kinase alpha is Involved in the Vitamin E-Induced Amelioration of Diabetic Nephropathy in Mice

  Daiki Hayashi, Keiko Yagi, Chihong Song, Shuji Ueda, Minoru Yamanoue, Matthew Topham, Toshinobu Suzaki, Naoaki Saito, Noriaki Emoto, Yasuhito Shirai

  Jun. 2017, SCIENTIFIC REPORTS, 7, English

  [Refereed]

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Enzymatically synthesized glycogen inhibits colitis through decreasing oxidative stress.

  SHIRAI YASUHITO

  Feb. 2017, Free Radic Biol Med, 106, 355–367, English

  [Refereed]

  Scientific journal


• 2)Effects of vitamin E and its derivatives on diabetic nephropathy in Rats and identification of diacylglycerol kinase subtype involved in the improvement of diabetic nephropathy

  Kakehi Tomoko, Yagi Keiko, Saito Naoaki, SHIRAI YASUHITO

  2017, Functional Foods in Health and Disease, 7(10) (10), 816 - 832, English

  [Refereed]

  Scientific journal


• A novel diacylglycerol kinase alpha-selective inhibitor, CU-3, induces cancer cell apoptosis and enhances immune response

  Ke Liu, Naoko Kunii, Megumi Sakuma, Atsumi Yamaki, Satoru Mizuno, Mayu Sato, Hiromichi Sakai, Sayaka Kado, Kazuo Kumagai, Hirotatsu Kojima, Takayoshi Okabe, Tetsuo Nagano, Yasuhito Shirai, Fumio Sakane

  Mar. 2016, JOURNAL OF LIPID RESEARCH, 57(3) (3), 368 - 379, English

  [Refereed]

  Scientific journal


• A novel diacylglycerol kinase alpha-selective inhibitor, CU-3, induces cancer cell apoptosis and enhances immune response

  Ke Liu, Naoko Kunii, Megumi Sakuma, Atsumi Yamaki, Satoru Mizuno, Mayu Sato, Hiromichi Sakai, Sayaka Kado, Kazuo Kumagai, Hirotatsu Kojima, Takayoshi Okabe, Tetsuo Nagano, Yasuhito Shirai, Fumio Sakane

  Mar. 2016, JOURNAL OF LIPID RESEARCH, 57(3) (3), 368 - 379, English

  [Refereed]

  Scientific journal


• Epigallocatechin-3-gallate activates diacylglycerol kinase alpha via a 67 kDa laminin receptor: A possibility of galloylated catechins as functional food to prevent and/or improve diabetic renal dysfunctions

  Daiki Hayashi, Shuji Ueda, Minoru Yamanoue, Naoki Saito, Hitoshi Ashida, Yasuhito Shirai

  May 2015, JOURNAL OF FUNCTIONAL FOODS, 15, 561 - 569, English

  [Refereed]

  Scientific journal


• Chicken heat shock protein HSPB1 increases and interacts with alpha B-crystallin in aged skeletal muscle

  Shuji Ueda, Yoshito Kokaji, Shunsaku Simizu, Kazuhisa Honda, Ken-ichi Yoshino, Hiroshi Kamisoyama, Yasuhito Shirai, Minoru Yamanoue

  2015, BIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY, 79(11) (11), 1867 - 1875, English

  [Refereed]

  Scientific journal


• 脂質結合ドメインを用いた新規ホスファチジン酸可視化プローブの開発

  OKIMOTO KO, NAKAI HIROKO, ITOH TOSHIKI, UEDA SHUJI, YAMANOUE MINORU, SHIRAI YASUHITO

  Dec. 2014, 日本農芸化学会関西支部講演会講演要旨集, 487th, 11, Japanese

  Research society


• 筋細胞におけるHSPB1の結合分子の探索とその機能解析

  小鍛治 泰斗, 上田 修司, 吉野 健一, 山之上 稔, 白井 康仁

  (公社)日本生化学会, Oct. 2014, 日本生化学会大会プログラム・講演要旨集, 87回, [4P - 154], Japanese


• Diacylglycerol kinase as a possible therapeutic target for neuronal diseases

  Yasuhito Shirai, Naoaki Saito

  Apr. 2014, JOURNAL OF BIOMEDICAL SCIENCE, 21, 28, English

  [Refereed][Invited]

  Scientific journal


• Both the C1 domain and a basic amino acid cluster at the C-terminus are important for the neurite and branch induction ability of DGK beta

  Takuya Kano, Takeshi Kouzuki, Satoru Mizuno, Shuji Ueda, Minoru Yamanoue, Fumio Sakane, Naoaki Saito, Yasuhito Shirai

  Apr. 2014, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 447(1) (1), 89 - 94, English

  [Refereed]

  Scientific journal


• Diacylglycerol kinase γ regulates antigen-induced mast cell degranulation by mediating Ca(2+) influxes.

  SAKUMA MEGUMI, SHIRAI YASUHITO, UEYAMA TAKEIKO, SAITO NAOAKI

  Feb. 2014, Biochem Biophys Res Commun, 445, 340 - 350, English

  [Refereed]

  Scientific journal


• The expression of diacylglycerol kinase theta during the organogenesis of mouse embryos

  Shuji Ueda, Becky Tu-Sekine, Minoru Yamanoue, Daniel M. Raben, Yasuhito Shirai

  Oct. 2013, BMC DEVELOPMENTAL BIOLOGY, 13, 35, English

  [Refereed]

  Scientific journal


• Depression of type I diacylglycerol kinases in pancreatic -cells from male mice results in impaired insulin secretion

  Kaneko Yukiko, Kobayashi Yusuke, Motoki Keisuke, Nakata Kunihito, Miyagawa Shoko, Yamamoto Mao, Hayashi Daiki, SHIRAI YASUHITO, Sakane Fumio, Ishikawa Tomohisa

  Endocrinology, Sep. 2013, Depression of type I diacylglycerol kinases in pancreatic -cells from male mice results in impaired insulin secretion, 154, 4089 - 4098, English

  [Refereed]

  Scientific journal


• PKC-epsilon pseudosubstrate and catalytic activity are necessary for membrane delivery during IgG-mediated phagocytosis

  Tiffany R. Wood, Rachel Y. Chow, Cheryl M. Hanes, Xuexin Zhang, Kaori Kashiwagi, Yasuhito Shirai, Mohamed Trebak, Daniel J. Loegering, Naoaki Saito, Michelle R. Lennartz

  Jul. 2013, JOURNAL OF LEUKOCYTE BIOLOGY, 94(1) (1), 109 - 122, English

  [Refereed]

  Scientific journal


• Both C1B domain and pseudosubstrate region are necessary for saturated fatty acid-induced translocation of epsilon PKC to the plasma membrane: Distinct role of intramolecular domains for different translocation

  Takaaki Nishimoto, Kaori Kashiwagi, Naoaki Saito, Yasuhito Shirai

  Mar. 2013, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 432(2) (2), 384 - 388, English

  [Refereed]

  Scientific journal


• Increased seizure susceptibility in a mouse with diacylglycerol kinase beta deficiency

  Mitsue Ishisaka, Kazuhiro Tsuruma, Masamitsu Shimazawa, Yasuhito Shirai, Naoaki Saito, Hideaki Hara

  2013, JOURNAL OF PHARMACOLOGICAL SCIENCES, 121, 181P - 181P, English

  [Refereed]

  Scientific journal


• Novel PKC alpha-mediated phosphorylation site(s) on cofilin and their potential role in terminating histamine release

  Megumi Sakuma, Yasuhito Shirai, Ken-ichi Yoshino, Maho Kuramasu, Tomofumi Nakamura, Toshihiko Yanagita, Kensaku Mizuno, Izumi Hide, Yoshihiro Nakata, Naoaki Saito

  Sep. 2012, MOLECULAR BIOLOGY OF THE CELL, 23(18) (18), 3707 - 3721, English

  [Refereed]

  Scientific journal


• c-Abl Tyrosine Kinase Regulates Serum-induced Nuclear Export of Diacylglycerol Kinase alpha by Phosphorylation at Tyr-218

  Takehiro Matsubara, Momo Ikeda, Yuko Kiso, Megumi Sakuma, Ken-ichi Yoshino, Fumio Sakane, Isabel Merida, Naoaki Saito, Yasuhito Shirai

  Feb. 2012, JOURNAL OF BIOLOGICAL CHEMISTRY, 287(8) (8), 5507 - 5517, English

  [Refereed]

  Scientific journal


• Regulation of diacylglycerol kinase by phosphorylation

  Y. Shirai, M. Ikeda, N. Saito

  Elsevier Ltd, 2012, Advances in Biological Regulation, 52(1) (1), 239 - 247, English

  [Refereed][Invited]

  Scientific journal


• Diacylglycerol kinase (DGK) as a regulator of PKC

  SHIRAI YASUHITO, Saito Naoaki

  2012, Neuromethods, 68, 259 - 271, English

  [Refereed][Invited]

  Scientific journal


• Streptococcus mutans diacylglycerol kinase homologue: a potential target for anti-caries chemotherapy

  Yukie Shibata, Miki Kawada-Matsuo, Yasuhito Shirai, Naoaki Saito, Dan Li, Yoshihisa Yamashita

  May 2011, JOURNAL OF MEDICAL MICROBIOLOGY, 60(5) (5), 625 - 630, English

  [Refereed]

  Scientific journal


• Direct binding of RalA to PKC eta and its crucial role in morphological change during keratinocyte differentiation

  Shirai, Yasuhito, Morioka, Shoko, Sakuma, Megumi, Yoshino, Ken-ichi, Otsuji, Chihiro, Sakai, Norio, Kashiwagi, Kaori, Chida, Kazuhiro, Shirakawa, Ryutaro, Horiuchi, Hisanori e

  During differentiation, keratinocytes undergo a dramatic shape change from small and round to large and flat, in addition to production of proteins necessary for the formation of epidermis. It has been shown that protein kinase C (PKC) eta is crucial for keratinocyte differentiation. However, its role in this process has yet to be fully elucidated. Here, we show that catalytic activity is not necessary for enlarged and flattened morphology of human keratinocytes induced by overexpression of PKC eta, although it is important for gene expression of the marker proteins. In addition, we identify the small G protein RalA as a binding partner of PKC eta, which binds to the C1 domain, an indispensable region for the morphological change. The binding led activation of RalA and actin depolymerization associated with keratinocyte differentiation. siRNA techniques proved that RalA is involved in not only the keratinocyte differentiation induced by PKC eta overexpression but also normal keratinocyte differentiation induced by calcium and cholesterol sulfate. These results provide a new insight into the molecular mechanism of cytoskeletal regulation leading to drastic change of cell shape.

  AMER SOC CELL BIOLOGY, Apr. 2011, MOLECULAR BIOLOGY OF THE CELL, 22(8) (8), 1340 - 1352

  [Refereed]

  Scientific journal


• Diacylglycerol Kinase beta Knockout Mice Exhibit Lithium-Sensitive Behavioral Abnormalities

  Kenichi Kakefuda, Atsushi Oyagi, Mitsue Ishisaka, Kazuhiro Tsuruma, Masamitsu Shimazawa, Koichi Yokota, Yasuhito Shirai, Kyoji Horie, Naoaki Saito, Junji Takeda, Hideaki Hara

  Oct. 2010, PLOS ONE, 5(10) (10), e13447, English

  [Refereed]

  Scientific journal


• Suppression mechanisms of flavonoids on aryl hydrocarbon receptor-mediated signal transduction

  Rie Mukai, Yasuhito Shirai, Naoaki Saito, Itsuko Fukuda, Shin Nishiumi, Ken-ichi Yoshida, Hitoshi Ashida

  Sep. 2010, ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS, 501(1) (1), 134 - 141, English

  [Refereed]

  Scientific journal


• Essential Role of Neuron-Enriched Diacylglycerol Kinase (DGK), DGK beta in Neurite Spine Formation, Contributing to Cognitive Function

  Yasuhito Shirai, Takeshi Kouzuki, Kenichi Kakefuda, Shigeki Moriguchi, Atsushi Oyagi, Kyoji Horie, Shin-ya Morita, Masamitsu Shimazawa, Kohji Fukunaga, Junji Takeda, Naoaki Saito, Hideaki Hara

  Jul. 2010, PLOS ONE, 5(7) (7), e11602, English

  [Refereed]

  Scientific journal


• A critical role of conventional protein kinase C in morphological changes of rodent mast cells

  Yanase Y, Hide I, Mihara S, Shirai Yasuhito, Saito N, Nakata Y, Hide M, Sakai N

  Feb. 2010, Immunol. Cell. Biol, 89, 149 - 159, English

  [Refereed]

  Scientific journal


• Inflammatory Cytokine Tumor Necrosis Factor-alpha Enhances Nerve Growth Factor Production in Human Keratinocytes, HaCaT Cells

  Koji Takaoka, Yasuhito Shirai, Naoaki Saito

  Dec. 2009, JOURNAL OF PHARMACOLOGICAL SCIENCES, 111(4) (4), 381 - 391, English

  [Refereed]

  Scientific journal


• An essential role of the aPKC-Aurora A-NDEL1 pathway in neurite elongation by modulation of microtubule dynamics

  Daisuke Mori, Masami Yamada, Yuko Mimori-Kiyosue, Yasuhito Shirai, Atsushi Suzuki, Shigeo Ohno, Hideaki saya, Anthony Wynshaw-Boris, Shinji Hirotsune

  Sep. 2009, NATURE CELL BIOLOGY, 11(9) (9), 1057 - U47, English

  [Refereed]

  Scientific journal


• Subcellular localization of flavonol aglycone in hepatocytes visualized by confocal laser scanning fluorescence microscope

  Rie Mukai, Yasuhito Shirai, Naoaki Saito, Ken-ichi Yoshida, Hitoshi Ashida

  Apr. 2009, CYTOTECHNOLOGY, 59(3) (3), 177 - 182, English

  [Refereed]

  Scientific journal


• Kinase activity of the dgk gene product is involved in the virulence of Streptococcus mutans

  Yukie Shibata, Jan R. van der Ploeg, Takeshi Kozuki, Yasuhito Shirai, Naoaki Saito, Miki Kawada-Matsuo, Toru Takeshita, Yoshihisa Yamashita

  Feb. 2009, MICROBIOLOGY-SGM, 155, 557 - 565, English

  [Refereed]

  Scientific journal


• Ca2+ oscillation induced by P2Y2 receptor activation and its regulation by a neuron-specific subtype of PKC (gamma PKC)

  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


• Reactive oxygen species derived from NOX1/NADPH oxidase enhance inflammatory pain

  Masakazu Ibi, Kuniharu Matsuno, Dai Shiba, Masato Katsuyama, Kazumi Iwata, Tomoko Kakehi, Takayuki Nakagawa, Kazunori Sango, Yasuhito Shirai, Takahiko Yokoyama, Shuji Kaneko, Naoaki Saito, Chihiro Yabe-Nishimura

  Sep. 2008, JOURNAL OF NEUROSCIENCE, 28(38) (38), 9486 - 9494, English

  [Refereed]

  Scientific journal


• Protein kinase C epsilon: function in neurons

  Yasuhito Shirai, Naoko Adachi, Naoaki Saito

  Aug. 2008, FEBS JOURNAL, 275(16) (16), 3988 - 3994, English

  [Refereed]

  Scientific journal


• Enzymological analysis of mutant protein kinase C gamma causing spinocerebellar ataxia type 14 and dysfunction in Ca2+ homeostasis

  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


• Lck-dependent tyrosine phosphorylation of diacylglycerol kinase alpha regulates its membrane association in T cells

  Ernesto Merino, Antonia Avila-Flores, Yasuhito Shirai, Ignacio Moraga, Naoaki Saito, Isabel Merida

  May 2008, JOURNAL OF IMMUNOLOGY, 180(9) (9), 5805 - 5815, English

  [Refereed]

  Scientific journal


• Regulation of clathrin-dependent endocytosis by diacylglycerol kinase delta: importance of kinase activity and binding to AP2 alpha

  Takumi Kawasaki, Takeshi Kobayashi, Takehiko Ueyama, Yasuhito Shirai, Naoaki Saito

  Jan. 2008, BIOCHEMICAL JOURNAL, 409, 471 - 479, English

  [Refereed]

  Scientific journal


• A novel PIP2 binding of epsilon PKC and its contribution to the neurite induction ability

  Yasuhito Shirai, Takuya Murakami, Maho Kuramasu, Leo Iijima, Naoaki Saito

  Sep. 2007, JOURNAL OF NEUROCHEMISTRY, 102(5) (5), 1635 - 1644, English

  [Refereed]

  Scientific journal


• A regulated adaptor function of p40(phox): Distinct p67(phox) membrane targeting by p40(phox) and by p47(phox)

  Takehiko Ueyama, Toshihiko Tatsuno, Takumi Kawasaki, Satoshi Tsujibe, Yasuhito Shirai, Hideki Sumimoto, Thomas L. Leto, Naoaki Saito

  Feb. 2007, MOLECULAR BIOLOGY OF THE CELL, 18(2) (2), 441 - 454, English

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• A regulated adaptor function of p40(phox): Distinct p67(phox) membrane targeting by p40(phox) and by p47(phox)

  Takehiko Ueyama, Toshihiko Tatsuno, Takumi Kawasaki, Satoshi Tsujibe, Yasuhito Shirai, Hideki Sumimoto, Thomas L. Leto, Naoaki Saito

  Feb. 2007, MOLECULAR BIOLOGY OF THE CELL, 18(2) (2), 441 - 454, English

  [Refereed]

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Phosphorylation and up-regulation of diacylglycerol kinase gamma via its interaction with protein kinase C gamma

  Yasuto Yamaguchi, Yasuhito Shirai, Takehiro Matsubara, Koichi Sanse, Masamitsu Kuriyama, Noriko Oshiro, Ken-ichi Yoshino, Kazuyoshi Yonezawa, Yoshitaka Ono, Naoaki Saito

  Oct. 2006, JOURNAL OF BIOLOGICAL CHEMISTRY, 281(42) (42), 31627 - 31637, English

  [Refereed]

  Scientific journal


• Phosphorylation and up-regulation of diacylglycerol kinase gamma via its interaction with protein kinase C gamma

  Yasuto Yamaguchi, Yasuhito Shirai, Takehiro Matsubara, Koichi Sanse, Masamitsu Kuriyama, Noriko Oshiro, Ken-ichi Yoshino, Kazuyoshi Yonezawa, Yoshitaka Ono, Naoaki Saito

  Oct. 2006, JOURNAL OF BIOLOGICAL CHEMISTRY, 281(42) (42), 31627 - 31637, English

  Scientific journal


• Nuclear transportation of diacylglycerol kinase gamma and its possible function in the nucleus

  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


• Nuclear transportation of diacylglycerol kinase gamma and its possible function in the nucleus

  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

  Scientific journal


• Targeting of protein kinase C-epsilon during Fc gamma receptor-dependent phagocytosis requires the epsilon C1B domain and phospholipase C-gamma 1

  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


• Local disassembly of actin stress fibers induced by selected release of intracellular tension in Osteoblastic cell

  Katsuya Sato, Taiji Adachi, Yasuhito Shirai, Naoaki Saito, Yoshihiro Tomita

  2006, Journal of biomechanical science and engineering, 1, 204--214, English

  [Refereed]

  Scientific journal


• Spatiotemporal analysis of the molecular interaction between PICK1 and PKC

  Kenji Masukawa, Norio Sakai, Shiho Ohmori, Yasuhito Shirai, Naoaki Saito

  2006, ACTA HISTOCHEMICA ET CYTOCHEMICA, 39(6) (6), 173 - 181, English

  Scientific journal


• Spatiotemporal analysis of the molecular interaction between PICK1 and PKC

  Kenji Masukawa, Norio Sakai, Shiho Ohmori, Yasuhito Shirai, Naoaki Saito

  2006, ACTA HISTOCHEMICA ET CYTOCHEMICA, 39(6) (6), 173 - 181, English

  [Refereed]

  Scientific journal


• Immunocytochemical localization of a neuron-specific diacylglycerol kinase beta and gamma in the developing rat brain

  N Adachi, M Oyasu, T Taniguchi, Y Yamaguchi, R Takenaka, Y Shirai, N Saito

  Oct. 2005, MOLECULAR BRAIN RESEARCH, 139(2) (2), 288 - 299, 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

  Aug. 2005, JOURNAL OF IMMUNOLOGY, 175(4) (4), 2381 - 2390, English

  Scientific journal


• Localization and functional interrelationships among cytosolic Group IV, secreted Group V, and Ca2+-independent group VI phospholipase A(2)s in P388D(1) macrophages using GFP/RFP constructs

  Y Shirai, J Balsinde, EA Dennis

  Jul. 2005, BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR AND CELL BIOLOGY OF LIPIDS, 1735(2) (2), 119 - 129, English

  [Refereed]

  Scientific journal


• Protein kinase C-alpha a mediates TNF release process in RBL-2H3 mast cells

  IT Abdel-Raheem, Hide, I, Y Yanase, YS Shigemoto-Mogami, N Sakai, Y Shirai, N Saito, FM Hamada, NA El-Mahdy, N Sakai, Y Shirai, N Saito, FMH Hamada, NA El-Mahdy, AEDE Elsisy, SS Sokar, Y Nakata

  Jun. 2005, BRITISH JOURNAL OF PHARMACOLOGY, 145(4) (4), 415 - 423, English

  Scientific journal


• Importance of chroman ring and tyrosine phosphorylation in the subtype-specific translocation and activation of diacylglycerol kinase alpha by D-alpha-tocopherol

  R Fukunaga-Takenaka, Y Shirai, K Yagi, N Adachi, N Sakai, E Merino, Merida, I, N Saito

  Apr. 2005, GENES TO CELLS, 10(4) (4), 311 - 319, English

  [Refereed]

  Scientific journal


• Phosphorylation of PKC activation loop plays an important role in receptor-mediated translocation of PKC

  Takahiro Seki, Hiroaki Matsubayashi, Taku Amano, Yasuhito Shirai, Naoaki Saito, Norio Sakai

  Mar. 2005, Genes to Cells, 10(3) (3), 225 - 239

  Scientific journal


• Activation and translocation of PKC delta is necessary for VEGF-induced ERK activation through KDR in FIEK293T cells

  M Kuriyama, T Taniguchi, Y Shirai, A Sasaki, A Yoshimura, N Saito

  Dec. 2004, BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS, 325(3) (3), 843 - 851, 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

  Oct. 2004, JOURNAL OF IMMUNOLOGY, 173(7) (7), 4582 - 4589, English

  [Refereed]

  Scientific journal


• Propagation of gamma PKC translocation along the dendrites of Purkinje cell in gamma PKC-GFP transgenic mice

  N Sakai, H Tsubokawa, M Matsuzaki, T Kajimoto, E Takahashi, Y Ren, S Ohmori, Y Shirai, H Matsubayashi, JS Chen, RS Duman, H Kasai, N Saito

  Oct. 2004, GENES TO CELLS, 9(10) (10), 945 - 957, English

  [Refereed]

  Scientific journal


• Phospholipase A2 products retain a neuron specific gamma isoform of PKC on the plasma membrane through the C1 domain - a molecular mechanism for sustained enzyme activity

  K Yagi, Y Shirai, M Hirai, N Sakai, N Saito

  Jul. 2004, NEUROCHEMISTRY INTERNATIONAL, 45(1) (1), 39 - 47, English

  [Refereed]

  Scientific journal


• Analysis of molecular mechanism regulating spatio-temporal localization and activity of protein kinase C and diacylglycerol kinase using live imaging

  SHIRAI Yasuhito

  Protein kinase C (PKC) changes its subcellular localization depending on extracellular signals including hormones and neurotransmitters. Such translocation is referred to as "targeting" in this review. The live imaging technique using GFP has allowed the dynamic movement of PKC to be visualized in living cells and revealed a remarkable diversity in PKC targeting. These studies indicate an importance of targeting in regulating the physiological and isotype-specific function of PKC. Like PKC, diacylglycerol kinase (DGK), which phosphorylates diacylglycerol resulting in attenuation of PKC, subtype-specifically translocates to particular subcellular compartments including the plasma membrane and Golgi complex. In addition, it has been shown that the localization and activation of the two functionally-related kinases are well organized by direct interaction and phosphorylation. This review summarizes diversity in targeting of PKC and DGK and the molecular mechanisms regulating their targeting.
  

  The Japanese Pharmacological Society, Mar. 2004, Folia Pharmacologica Japonica, 123(3) (3), 189 - 196, Japanese


• Ceramide-induced apoptosis by translocation, phosphorylation, and activation of protein kinase Cdelta in the Golgi complex

  Kajimoto Taketoshi, SHIRAI, Yasuhito, SAKAI,Norio, YAMAMOTO, Toshiyoshi, MATSUZAKI, Hidenori, KIKKAWA, Ushio, SAITO,Naoaki

  Mar. 2004, The Journal Of Biological Chemistry, Vol. 279, No. 13, pp. 12668-12, 12668 - 12676


• Isoform-specific phosphorylation of metabotropic glutamate receptor 5 by protein kinase C (PKC) blocks Ca2+ oscillation and oscillatory translocation of Ca2+-dependent PKC

  M Uchino, N Sakai, K Kashiwagi, Y Shirai, Y Shinohara, K Hirose, M Iino, T Yamamura, N Saito

  Jan. 2004, JOURNAL OF BIOLOGICAL CHEMISTRY, 279(3) (3), 2254 - 2261


• Phosphorylation of PKC activation loop plays an important role in receptor-mediated translocation of PKC

  T Seki, H Matsubayashi, T Amano, N Saito, N Sakai

  2004, JOURNAL OF PHARMACOLOGICAL SCIENCES, 94, 97P - 97P


• リチウム慢性投与は神経伝達物質によるαPKCのトランスロケーションを抑制する

  阪井一雄, SHIRAI, Yasuhito, SAITO,Naoaki, MAEDA, Kiyoshi

  Dec. 2003, 日本神経精神薬理学雑誌, 23巻, 6号, pp. 271-271


• Synthesis and phorbol ester binding of the cysteine-rich domains of diacylglycerol kinase (DGK) isozymes - DGK gamma and DGK gamma are new targets of tumor-promoting phorbol esters

  M Shindo, K Irie, A Masuda, H Ohigashi, Y Shirai, K Miyasaka, N Saito

  May 2003, JOURNAL OF BIOLOGICAL CHEMISTRY, 278(20) (20), 18448 - 18454


• Novel variants of murine serotonin transporter mRNA and the promoter activity of its upstream site

  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


• プロテインキナーゼＣのターゲティング機構

  白井康仁, 斎藤尚亮

  Apr. 2003, 蛋白質 核酸 酵素, 48, 1241 - 1247


• ターゲティングを指標としたプロテインキナーゼＣ(PKC)の機能解析

  白井康仁, 斎藤尚亮

  Apr. 2003, 日薬理誌, 121, 421 - 434


• A role for PKC-ε in FcγR-mediated phagocytosis by RAW 264.7 cells

  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


• Subtype- and species-specific knockdown of PKC using short interfering RNA.

  Irie N, Sakai N, Ueyama T, Kajimoto T, Shirai Y, Saito N

  Nov. 2002, Biochemical and biophysical research communications, 298(5) (5), 738 - 743


• 蛍光蛋白質融合プロテインキナーゼC(PKC)を用いたPKCの機能解析 (研究)

  酒井 規雄, 白井 康仁, 斎藤 尚亮

  神緑会, Aug. 2002, 神戸大学医学部神緑会学術誌, 18, 88 - 92


• Importance of C1B domain for lipid messenger-induced targeting of PKC.

  Kashiwagi, K, Shirai, Y, Kuriyama, M, Sakai, N, Saito, N

  Apr. 2002, J. Biol. Chem., 277, 18037 - 18045


• Distinct regulatory mechanism for p70 S6 kinase beta from that for p70 S6 kinase alpha.

  Taichi Minami, Kenta Hara, Noriko Oshiro, Sachiko Ueoku, Ken-ichi Yoshino, Chinaru Tokunaga, Yasuhito Shirai, Naoaki Saito, Ivan Gout, Kazuyoshi Yonezawa

  BACKGROUND: A novel ribosomal S6 kinase, termed p70 S6 kinase beta (p70beta), has a highly homologous amino acid sequence to that of p70/p85 S6 kinase (p70alpha). This includes the critical phosphorylation sites, Thr252, Ser394 and Thr412 in p70alpha1, which correspond to Thr241, Ser383 and Thr401 in p70beta1, respectively. However, the regulatory mechanism for p70beta remains to be elucidated. RESULTS: We report here the expression and the mechanism of in vivo regulation of p70beta. Two isoforms, p70beta1 and p70beta2, were expressed in a variety of tissues at a different level. p70beta1 was mainly targeted to the nucleus, whereas p70beta2 dispersed throughout the cytoplasm including nucleoplasm. The kinase activity of p70beta1 was less sensitive to the inhibition induced by rapamycin, wortmannin and amino acid withdrawal than that of p70alpha. The portion of p70beta activity inhibited by rapamycin was rescued by the rapamycin-resistant mutant of the mammalian target of rapamycin (mTOR). Mutational analysis revealed that the phosphorylation of Thr241 and Thr401 in p70beta1 was indispensable for the kinase activity. In contrast, a p70beta1 mutant in which Ser383 was substituted with Gly (S383G) still retained nearly the half maximal activity. Sequential phosphorylation of wild-type and S383G mutant of p70beta1 with mTOR and 3-phosphoinositide-dependent protein kinase 1 (PDK1) in vitro synergistically activated their kinase activities. CONCLUSION: These results indicate that p70beta is regulated by the mTOR- and PDK1-signalling pathways through a synergistic interaction between phosphorylated Thr241 and Thr401, while Ser383 plays minor role in their activation mechanism. Activated p70beta may be less sensitive to dephosphorylation mediated by putative phosphatases activated by rapamycin, amino acid withdrawal, and probably wortmannin.

  Nov. 2001, Genes to Cells, 6(11) (11), 1003 - 1015


• Subtype-specific translocation of the delta subtype of protein kinase C and its activation by tyrosine phosphorylation induced by ceramide in HeLa cells.

  Kajimoto T, Ohmori S, Shirai Y, Sakai N, Saito N

  Mar. 2001, Molecular and cellular biology, 21(5) (5), 1769 - 1783


• Subtype-specific translocation of diacylglycerol kinase alpha and gamma and its correlation with protein kinase C.

  Shirai Y, Segawa S, Kuriyama M, Goto K, Sakai N, Saito N

  Apr. 2000, The Journal of Biological Chemistry, 275(32) (32), 24760 - 24766


• Induction of 55 kDa protein of PKN by ischemia/reperfusion model of rat retina.

  Sumioka, K, Shirai, Y, Sakai, N, Hashimoto, T, Tanaka, C, Yamamoto M, Takahashi, M, Ono, Y, Saito, N

  Apr. 2000, Invest. Ophthal. Vis. Sci., 41, 29 - 35


• Phospholipase A2 and its products are involved in the purinergic receptor-mediated translocation of protein kinase C in CHO-K1 cells.

  Shirai, Y, Kashiwagi, K, Sakai, N, Saito, N

  Apr. 2000, J. Cell Sci., 113, 1335 - 1343


• Regulation of nuclear translocation of Forkhead transcription factor AFX by protein kinase B

  Hiroshi Takaishi, Hiroaki Konishi, Hidenori Matsuzaki, Yoshitaka Ono, Yasuhito Shirai, Naoaki Saito, Tadahiro Kitamura, Wataru Ogawa, Masato Kasuga, Ushio Kikkawa, Yasutomi Nishizuka

  Oct. 1999, Proceedings of the National Academy of Sciences of the United States of America, 96(21) (21), 11836 - 11841


• プロテインキナーゼＣのターゲッテイング機構の解析

  斎藤尚亮, 白井康仁

  Apr. 1999, 実験医学, 17, 1867 - 1873


• Molecular Cloning of cDNA for BRab from the Brain of Bombyx mori and Biochemical Properties of BRab Expressed in Escherichia coli

  UNO Tomohide, UENO Mayumi, NAKAJIMA Ayumi, SHIRAI Yasuhito, AIZONO Yasuo

  From a brain cDNA library of Bombyx mori, we cloned cDNA for BRab, which encoded a 202-amino-acid polypeptide sharing 60-80% similarity with rab1 family members. To characterize its biochemical properties, cDNA for BRab was inserted into an expression vector (pGEX2T) and expressed in Escherichia coli as a glutathione S-trans-ferase (GST) fusion protein. The recombinant protein was purified to homogeneity with glutathione S-Sepharose. The purified GST-BRab bound [^<35>S]-GTPγS and [^3H]-GDP with association constants of 1.5×10^6M^<-1> and 0.58×10^6M^<-1>, respectively. The binding of [^<35>S]-GTPγS was inhibited with GTP and GDP, but with no other nucleotides. The GTP-hydrolysis activity was evaluated to be 5 m mole/min/mole of BRab. In the presence of 6 mM MgCl_2,bound [^<35>S]-GTPγS and [^3H]-GDP were exchanged with GTPγS most efficiently. These results suggest that BRab, having a higher affinity for GTP than GDP, converts from the GTP-bound state into the GDP-bound state by intrinsic GTP hydrolysis activity and returns to the GTP-bound state with the exchange of GDP with GTP.

  Japan Society for Bioscience, Biotechnology, and Agrochemistry, Oct. 1998, Bioscience, biotechnology, and biochemistry, 62(10) (10), 1885 - 1891


• Amyloid beta protein (25-35) phosphorylates MARCKS through tyrosine kinase-activated protein kinase C signaling pathway in microglia.

  Shirai Y, Kashiwagi K, Yagi K, Sakai N, Saito N

  Oct. 1998, J Cell Biol., 143(2) (2), 511 - 521


• 神経系のトランスポーターと疾患 セロトニントランスポーターの転写及び機能の制御機構

  斎藤 尚亮, 白井 康仁, 酒井 規雄, 大倉 睦美, 上山 建彦

  日本神経化学会, Sep. 1998, 神経化学, 37(3) (3), 283 - 283


• Three distinct mechanisms for translocation and activation of δ-subspecies of protein kinase C.

  Ohmori, S, Shirai, Y, Sakai, N, Fujii, M, Konishi, H, Kikkawa, U, Saito, N

  Apr. 1998, Mol. Cell. Biol., 18, 5263 - 5271


• Subspecies-specific targeting mechanism of protein kinase C.

  Shirai, Y, Sakai, N, Saito, N

  Apr. 1998, Jpn J Pharmacol, 78, 411 - 417


• Distinct effects of fatty acids on translocation of γ- and ε- protein kinase C.

  Shirai, Y, Kashiwagi, K, Yagi, K, Sakai, N, Saito, N

  Apr. 1998, J. Cell Biol., 143, 511 - 521


• Direct visualization of the translocation of the γ-subspecies of protein kinase C in living cells using fusion protein with green fluorescent protein.

  Sakai, N, Sasaki, K, Ikegaki, N, Shirai, Y, Ono, Y, Saito, N

  Apr. 1997, J. Cell Biol., 139, 1465 - 1476

• 筋細胞の小胞体ストレス応答におけるHSPB5の結合分子の探索と機能解析

  西原萌華, 上田修司, 吉野健一, 吉野健一, 竹内敦子, 山之上稔, 白井康仁

  2020, 日本農芸化学会関西支部講演会講演要旨集, 512th


• Diacylglycerol kinase α-selective inhibitors induce apoptosis and reduce viability of melanoma and several other cancer cell lines.

  Atsumi Yamaki, Rino Akiyama, Chiaki Murakami, Saki Takao, Yuki Murakami, Satoru Mizuno, Daisuke Takahashi, Sayaka Kado, Akinobu Taketomi, Yasuhito Shirai, Kaoru Goto, Fumio Sakane

  Diacylglycerol (DG) kinase (DGK), which phosphorylates DG to generate phosphatidic acid (PA), consists of ten isozymes (α-к). Recently, we identified a novel small molecule inhibitor, CU-3, that selectively inhibits the activity of the α isozyme. In addition, we newly obtained Compound A, which selectively and strongly inhibits type I DGKs (α, β, and γ). In the present study, we demonstrated that both CU-3 and Compound A induced apoptosis (caspase 3/7 activity and DNA fragmentation) and viability reduction of AKI melanoma cells. Liquid chromatography-mass spectrometry revealed that the production of 32:0- and 34:0-PA species was commonly attenuated by CU-3 and Compound A, suggesting that lower levels of these PA molecular species are involved in the apoptosis induction and viability reduction of AKI cells. We determined the effects of the DGKα inhibitors on several other cancer cell lines derived from refractory cancers. In addition to melanoma, the DGKα inhibitors enhanced caspase 3/7 activity and reduced the viability of hepatocellular carcinoma, glioblastoma, and pancreatic cancer cells, but not breast adenocarcinoma cells. Interestingly, Western blot analysis indicated that the DGKα expression levels were positively correlated with the sensitivity to the DGK inhibitors. Because both CU-3 and Compound A induced interleukin-2 production by T cells, it is believed that these two compounds can enhance cancer immunity. Taken together, our results suggest that DGKα inhibitors are promising anticancer drugs.

  Jun. 2019, Journal of cellular biochemistry, 120(6) (6), 10043 - 10056


• BioID法を用いたHSPB5の新規相互作用分子の探索

  西原萌華, 上田修司, 吉野健一, 吉野健一, 竹内敦子, 山之上稔, 白井康仁

  2019, 日本分子生物学会年会プログラム・要旨集(Web), 42nd


• ジアシルグリセロールキナーゼαの新規結合分子の探索

  木本裕子, 渡辺真以, 上田修司, 吉野健一, 山之上稔, 白井康仁

  2019, 日本農芸化学会関西支部講演会講演要旨集, 507th


• リチウムによるDGKβKOマウスの記憶及び感情障害の改善には、PIP2の減少とPKCβの活性抑制が関与している

  沖本 航, 中井 寛子, 石坂 光絵, 中西 広樹, 森口 茂樹, 上田 修司, 山之上 稔, 福永 浩司, 佐々木 雄彦, 原 英彰, 白井 康仁

  日本脂質生化学会, May 2016, 脂質生化学研究, 58, 229 - 233


• 多成分一斉解析技術による黒毛和種牛肉の食品成分の検討

  上田修司, 岩本英治, 正木達規, 谷元哲則, 渡代勝之, 白井康仁, 山之上稔

  2016, 日本畜産学会大会講演要旨, 121st


• Screening of novel diacylglycerol kinase alpha (DGK alpha) activator to develop drugs improving diabetic nephropathy

  Hayashi Daiki, Liu Ka, Ueda Shuji, Yamanoue Minoru, Sakane Fumio, Shirai Yasuhito

  Jul. 2015, JOURNAL OF PHARMACOLOGICAL SCIENCES, 128(3) (3), S102


• ジアシルグリセロールキナーゼαの218番目のチロシンリン酸化はドメイン間相互作用と高次構造を変化させる

  脇阪昌明, 岩下智秋, 濱田大三, 濱田大三, 祇園景子, 上田修司, 山之上稔, 鶴田宏樹, 白井康仁

  2015, 日本生化学会大会(Web), 88th


• チロシンリン酸化がジアシルグリセロールキナーゼαのドメイン間相互作用及び高次構造に与える影響

  脇阪昌明, 岩下智秋, 濱田大三, 濱田大三, 祇園景子, 上田修司, 山之上稔, 鶴田宏樹, 白井康仁

  2015, 日本農芸化学会関西支部講演会講演要旨集, 492nd


• Function of Diacylglycerol kinase in the brain

  SHIRAI YASUHITO

  May 2014, 生化学, 86, 518 - 522


• Function of Diacylglycerol kinase beta in neurons

  SHIRAI YASUHITO

  ジアシルグリセロールキナーゼ（DGK）はジアシルグリセロール（DG）をリン酸化し，ホスファチジン酸（PA）に変換する脂質キナーゼであり，これまでに10種のサブタイプが報告されている．周知のようにDGはPKCの活性化因子であり，産生されるPAも様々な酵素の活性を調節することから，DGKはPKCの抑制やPAの産生を介して生体内において重要な働きをしていると考えられている．しかし，神経系に多く存在するβサブタイプの機能は長い間不明であった．そこで，我々はDGKβのノックアウト（KO）マウスを作製し，その神経系における機能を調べた．その結果，DGKβKOマウスは，記憶障害と感情障害を示した．また，この感情障害は10日間のリチウム処理で改善した．一方，DGKβKOマウスから調製した海馬および大脳皮質初代培養細胞は，突起の分岐の数およびスパイン数が有意に減少していたが，DGKβを過剰発現させることで形態異常は回復した．さらに， DGKβKOマウスの海馬および大脳皮質において，スパイン密度が減少していることを確認した．これらのことから，DGKβは神経細胞の形態を調節・維持することにより神経ネットワークの形成，ひいては記憶や感情などの脳高次機能において重要な働きをしていることが明らかになった．本総説では，このDGKβKOマウスから得られた知見を中心に，神経系におけるDGKβについて概説するとともに，DGKβの記憶障害や感情障害の予防薬および改善薬のターゲットとしての可能性と問題点について論ずる．

  The Japanese Pharmacological Society, Apr. 2014, Folia Pharmacol. Jpn, 143(3) (3), 131 - 136


• The effects of valproate on the abnormal behaviors in diacylglycerol kinase beta knockout mice

  Mitsue Ishisaka, Kazuhiro Tsuruma, Masamitsu Shimazawa, Yasuhito Shirai, Hideaki Hara

  2014, JOURNAL OF PHARMACOLOGICAL SCIENCES, 124, 214P - 214P


• mTOR is involved in both kinase activity-dependent and -independent pathways of DGK beta-regulated neurite and branch induction

  Takuya Kano, Hiroko Nakai, Akio Nakashima, Ushio Kikkawa, Naoaki Saito, Nobuyuki Takei, Shuji Ueda, Minoru Yamanoue, Yasuhito Shirai

  2014, JOURNAL OF PHARMACOLOGICAL SCIENCES, 124, 100P - 100P


• ジアシルグリセロールキナーゼβによる特徴的な神経突起伸にはcPKCも関与する

  中井 寛子, 加野 拓也, 上田 修司, 山之上 稔, 斎藤 尚亮, 白井 康仁

  28 May 2013, 脂質生化学研究, 55, 44 - 47


• Involvement of mTOR in diacylglycerol kinase beta (DGK beta)-induced neurite blanching and spine formation

  H. Nakai, N. Takei, A. Nakashima, U. Kikkawa, N. Saito, S. Ueda, M. Yamanoue, Y. Shirai

  Sep. 2012, FEBS JOURNAL, 279, 383 - 384


• ジアシルグリセロールキナーゼβによる特徴的な神経突起伸にはmTORが関与する

  白井 康仁, 中井 寛子, 中西 広樹, 中嶋 昭雄, 吉川 潮, 上田 修司, 山之上 稔, 斎藤 尚亮

  28 May 2012, 脂質生化学研究, 54, 98 - 100


• The role of diacylglycerol kinase in histamine release

  Megumi Sakuma, Yasuhito Shirai, Takehiko Ueyama, Naoaki Saito

  2012, JOURNAL OF PHARMACOLOGICAL SCIENCES, 118, 255P - 255P


• Physiological function of nuclear DGK alpha under serum depleted conditions

  Yuko Kiso, Yasuhito Shirai, Megumi Sakuma, Momo Ikeda, Naoaki Saito

  2012, JOURNAL OF PHARMACOLOGICAL SCIENCES, 118, 166P - 166P


• Diacylglycerol kinase as a possible drug target

  Yasuhito Shirai, Naoaki Saito

  2012, JOURNAL OF PHARMACOLOGICAL SCIENCES, 118, 31P - 31P


• ジアシルグリセロールキナーゼαの核移行と細胞周期制御に関する研究2 : c-Ablによるチロシンリン酸化サイトの同定

  白井 康仁, 池田 もも, 木曽 裕子, 佐久間 恵, 齋藤 尚亮

  28 Apr. 2011, 脂質生化学研究, 53, 191 - 194


• Novel binding of PKC eta to RalA and its crucial role in morphological change during keratinocytes differentiation

  Yasuhito Shirai, Naoaki Saito

  2011, JOURNAL OF PHARMACOLOGICAL SCIENCES, 115, 32P - 32P


• c-Abl phosphorylates DGK alpha at Y218 during the serum-induced nuclear export

  Momo Ikeda, Yasuhito Shirai, Takehiro Matsubara, Megumi Sakuma, Yuko Kiso, Naoaki Saito

  2011, JOURNAL OF PHARMACOLOGICAL SCIENCES, 115, 167P - 167P


• The function of P40(phox) in Nox2 activation

  Jun-ya Nakakita, Takehiko Ueyama, Toshihiro Kobayashi, Takeshi Kobayashi, Jon-hyun Son, Mio Nakatsuji, Yasuhito Shirai, Naoaki Saito

  2010, JOURNAL OF PHARMACOLOGICAL SCIENCES, 112, 118P - 118P


• Novel phosphorylation site(s) of cofilin by PKC alpha and its role in the degranulation from RBL-2H3 mast cells

  Megumi Sakuma, Yasuhito Shirai, Ken-ichi Yoshino, Maho Kuramasu, Tomofumi Nakamura, Izumi Hide, Yoshihiro Nakata, Naoaki Saito

  2010, JOURNAL OF PHARMACOLOGICAL SCIENCES, 112, 54P - 54P


• Mania-like behavioural abnormalities of diacylglycerol kinase beta knockout mice

  K. Kakefuda, A. Oyagi, K. Tsuruma, M. Shimazawa, K. Yokota, Y. Shirai, K. Horie, N. Saito, J. Takeda, H. Hara

  Sep. 2009, EUROPEAN NEUROPSYCHOPHARMACOLOGY, 19, S471 - S471


• ジアシルグリセロールキナーゼβノックアウトマウスはスパイン形成異常と記憶障害を示す : スパイン形成における脂質シグナリングの重要性

  白井 康仁, 上月 健, 掛札 賢一, 大八木 篤, 堀江 恭二, 森口 茂樹, 嶋澤 雅光, 福永 浩司, 竹田 潤二, 齋藤 尚亮, 原 英彰

  10 Jul. 2009, 脂質生化学研究, 51, 98 - 100


• Role of diacylglycerol kinase beta in neural function

  Takeshi Kozuki, Yasuhito Shirai, Kenichi Kakefuda, Atsushi Ohyagi, Kyoji Horie, Masamitsu Shimazawa, Kohji Fukunaga, Junji Takeda, Naoaki Saito, Hideaki Hara

  2009, JOURNAL OF PHARMACOLOGICAL SCIENCES, 109, 70P - 70P


• Mood-disorder and neuron specific isoform of diacylglycerol kinase, DGK beta

  Yasuhito Shirai, Naoaki Saito

  2009, JOURNAL OF PHARMACOLOGICAL SCIENCES, 109, 30P - 30P


• Diacylglycerol kinase beta (DGK beta) knock out mice show mania-like behaviors

  Kenichi Kakefuda, Atsushi Oyagi, Kazuhiro Tsuruma, Masamitsu Shimazawa, Koichi Yokota, Yasuhito Shirai, Kyoji Horie, Naoaki Saito, Junji Takeda, Hideaki Hara

  2009, JOURNAL OF PHARMACOLOGICAL SCIENCES, 109, 156P - 156P


• Cofilin is phosphorylated by cPKC in the degranulation from RBL-2H3 mast cells

  Megumi Sakuma, Yasuhito Shirai, Maho Kuramasu, Kenichi Yoshino, Tomofumi Nakamura, Izumi Hide, Yoshihiro Nakata, Naoaki Saito

  2009, JOURNAL OF PHARMACOLOGICAL SCIENCES, 109, 236P - 236P


• Important role of diacylglycerol kinase beta in spine formation and memory

  Yasuhito Shirai, Takeshi Kozuki, Kenichi Kakefuda, Atsushi Ohyagi, Kyouji Horie, Shigeki Moriguchi, Masamitsu Simazawa, Kohji Fukunaga, Jyunnji Takeda, Naoaki Saito, Hideaki Hara

  2009, NEUROSCIENCE RESEARCH, 65, S68 - S68


• Differential regulation of Dual oxidase 1 by Duox activator 1 splicing variants

  Satoshi Tsujibe, Takehiko Ueyama, Yasuhito Shirai, Leto Thomas, Naoaki Saito

  2008, JOURNAL OF PHARMACOLOGICAL SCIENCES, 106, 53P - 53P


• PKC induced phosphorylation of Orai1 regulates intracellular Ca2+ level from store operated channel

  Takumi Kawasaki, Takehiko Ueyama, Yasuhito Shirai, Naoaki Saito

  2008, JOURNAL OF PHARMACOLOGICAL SCIENCES, 106, 116P - 116P


• DG-PKCシグナル系の可視化とその異常による疾患

  斎藤 尚亮, 白井 康仁, 松原 岳大, 足立 直子, 高橋 英之, 酒井 規雄

  05 Jun. 2007, 脂質生化学研究, 49, 15 - 15


• 核内脂質シグナル及び核移行蛋白質の可視化システムを用いた細胞周期機構の解明 : DGKの核-細胞質間移行機構と核内における機能に着目して

  白井 康仁, 松原 岳大, 齋藤 尚亮

  05 Jun. 2007, 脂質生化学研究, 49, 305 - 307


• 脂質代謝酵素DGKαの核-細胞質間 shuttling 機構と細胞周期制御への関与

  松原 岳大, 白井 康仁, 斎藤 尚亮

  05 Jun. 2007, 脂質生化学研究, 49, 308 - 311


• Diacylglycerol kinase(DGK) ジアシルグリセロールキナーゼ

  Shirai Yasuhito

  Mar. 2007, 日本薬理学会雑誌, 129巻, 4号, pp. 311-313


• Analysis of inter- and intra-molecular interactions of phox proteins using a new fluorescent protein at cellular levels

  Tomoko Kusakabe, Aya Shimizu, Takehiko Ueyama, Takumi Kawasaki, Yasuhito Shirai, Satosi Karasawa, Atsusi Miyawaki, Naoaki Saito

  2007, JOURNAL OF PHARMACOLOGICAL SCIENCES, 103, 54P - 54P


• A regulated adaptor function of p40phox: a head-to-tail (PX-PB1 domain) intramolecular interaction of p40phox in its resting state

  Takehiko Ueyama, Toshihiko Tatsuno, Takumi Kawasaki, Satosi Tsujibe, Yasuhito Shirai, Hideki Sumimoto, Thomas. L. Leto, Naoaki Saito

  2007, JOURNAL OF PHARMACOLOGICAL SCIENCES, 103, 95P - 95P


• The functional relation between PKC and actin arrangement in the degranulation from RBL-2H3 cells.

  Maho Kuramasu, Yasuhito Shirai, Tomofumi Nakamura, Izumi Hide, Yoshihiro Nakata, Naoaki Saito

  2007, JOURNAL OF PHARMACOLOGICAL SCIENCES, 103, 215P - 215P


• Spatial and temporal controls of PKC isoform activation

  Yasuhito Shirai, Naoaki Saito

  Jul. 2006, ACTA PHARMACOLOGICA SINICA, 27, 3 - 3


• プロテインキナーゼCεのフォスファチジルイノシトール2リン酸結合能と神経突起伸長能との相関

  白井 康仁, 村上 卓也, 倉増 真帆, 齋藤 尚亮

  08 Jun. 2006, 脂質生化学研究, 48, 181 - 183


• 脂溶性リガンドの細胞質膜レセプター

  白井 康仁, 齋藤 尚亮

  01 Mar. 2005, 日本薬理学雑誌 : FOLIA PHARMACOLOGICA JAPONICA, 125(3) (3), 166 - 167


• 糖尿病性血管合併症とPKC/DGKシグナリング

  白井 康仁, 竹中 利佳, 齋藤 尚亮

  01 Nov. 2004, 日本薬理学雑誌 : FOLIA PHARMACOLOGICA JAPONICA, 124, 35P - 36P


• 糖尿病性血管合併症とジアシルグリセロールキナーゼ

  白井 康仁, 齋藤 尚亮

  01 Aug. 2004, 日本薬理学雑誌 : FOLIA PHARMACOLOGICA JAPONICA, 124(2) (2), 128 - 128


• 脳部位特異的Tet制御γPKC-GFP発現トランスジェニックマウスを用いたγPKCの in vivo ライブイメージング

  酒井規雄, 坪川宏, 松崎政紀, 松林弘明, 梶本武利, 大森志保, 白井康仁, 河西春郎, 斎藤尚亮

  25 Dec. 2003, 日本神経精神薬理学雑誌 = Japanese journal of psychopharmacology, 23(6) (6)


• Analysis of PKC targeting mechanism using PKC fused with fluorescent proteins

  Norio Sakai, Yasuhito Shirai, Naoaki Saito

  2003, Folia Pharmacologica Japonica, 121(6) (6), 421 - 434


• Protein kinase Cγ (PKCγ): Function of neuron specific isotype

  Naoaki Saito, Yasuhito Shirai

  Japanese Biochemical Society, 01 Nov. 2002, Journal of Biochemistry, 132(5) (5), 683 - 687


• Activation mechanisms of protein kinase C: Maturation, catalytic activation, and targeting

  Yasuhito Shirai, Naoaki Saito

  Japanese Biochemical Society, 01 Nov. 2002, Journal of Biochemistry, 132(5) (5), 663 - 668


• The silencer activity of the novel human serotonin transporter linked polymorphic regions

  Kazuo Sakai, Masayuki Nakamura, Shu Ichi Ueno, Akira Sano, Norio Sakai, Yasuhito Shirai, Naoaki Saito

  12 Jul. 2002, Neuroscience Letters, 327(1) (1), 13 - 16


• Oscillatory translocation of gamma PKC by the activation of metabotropic glutamate receptor and its regulation by delta PKC in a subtype specific manner.

  M Uchino, N Sakai, Y Shirai, K Hirose, M Iino, Y Shinohara, S Nakanishi, N Saito

  2002, JAPANESE JOURNAL OF PHARMACOLOGY, 88, 175P - 175P


• W2-1 Analysis of PKC targeting mechanism in living cells for understanding PKC signaling :

  Sakai Norio, Shirai Yasuhito, Saito Naoaki

  Japan Society of Histochemistry and Cytochemistry, 2001, Acta histochemica et cytochemica, 34(1) (1), 53 - 53


• Serotonin transporter

  N Sakai, Y Shirai, N Saito

  Oct. 2000, SEIKAGAKU, 72(10) (10), 1215 - 1229


• W2-1 PKCシグナル伝達のさらなる理解を求めて : PKCターゲティング機構の解析(細胞内情報伝達への形態学的アプローチ:リン酸化酵素を中心として,ワークショップ2,第41回 日本組織細胞化学会総会・学術集会)

  酒井 規雄, 白井 康仁, 斎藤 尚亮

  日本組織細胞化学会, 2000, 日本組織細胞化学会総会プログラムおよび抄録集, (41) (41), 59 - 59


• 蛋白質キナーゼのターゲッティングのイメージングーGFP融合プロテインキナーゼＣを用いた研究を中心にー

  Sakai Norio, SHIRAI YASUHITO, Saito Naoaki

  2000, 脳の科学, 22, 783 - 789


• Analysis of signal transduction using GFP-tagged protein kinase C. :

  Shirai Yasuhito, Saito Naoaki

  Japan Society of Histochemistry and Cytochemistry, 1999, Acta histochemica et cytochemica, 32(6) (6), 513 - 513


• Visualization of stimulus-and subtype-specific translocation of protein kinase C(PKC)in living cells : Molecular mechanism of PKC targeting :

  SAITO Naoaki, SAKAI Norio, SHIRAI Yasuhito

  Japan Society of Histochemistry and Cytochemistry, 1999, Acta histochemica et cytochemica, 32(6) (6), 517 - 517


• WS I-2 プロテインキナーゼCのターゲッティング機構と生理機能

  斎藤 尚亮, 酒井 規雄, 白井 康仁

  日本組織細胞化学会, 1999, 日本組織細胞化学会総会プログラムおよび抄録集, (40) (40), 122 - 122


• S XI-4 GFP標識タンパク質を用いた細胞内情報伝達機構の解析

  白井 康仁, 斎藤 尚亮

  日本組織細胞化学会, 1999, 日本組織細胞化学会総会プログラムおよび抄録集, (40) (40), 112 - 112


• GFPを用いた細胞内情報伝達系の可視化--PKC translocationを中心として (日本電子顕微鏡学会第43回シンポジウム論文集--21世紀へ向けての新技術の展開--平成10年10月28日(水)〜30日(金),千葉大学けやき会館〔含 著者名索引〕) -- (ワ-クショップ(第6回コンフォ-カル488シンポジウム) 生細胞・生組織のイメ-ジング技術のフロンティア)

  斎藤 尚亮, 白井 康仁

  日本顕微鏡学会, 1998, 電子顕微鏡, 33(2) (2), 177 - 180

• ビタミンEの非抗酸化的な生理作用を仲介する受容体

  白井康仁, 林大輝

  建帛社, May 2025


• 小脳が司る協調運動におけるジアシルグリセロールシグナリングの重要性

  津曲 涼介, SHIRAI YASUHITO

  日本薬理学雑誌, Aug. 2018


• Nutritional and Therapeutic Interventions for Diabetes and Metabolic Syndrome

  SHIRAI YASUHITO, Daiki Hayashi

  ELSEVIER, Jun. 2018


• 分子生物学

  SHIRAI YASUHITO

  化学同人, Feb. 2011


• Determination of subcellular localization of flavonol in cultured cells by laser scanning. In "Laser Scanning, Theory and Applications!", Ed. by Chau-Chang Wang, pp.215-232.

  MUKAI Rie, TERAO Junji, SHIRAI Yasuhito, SAITO Naoaki, ASHIDA Hitoshi

  Intec-Open Access Publisher, 2011


• Diacylglycerol kinase

  Shirai Yasuhito

  日本生物工学会, Aug. 2008


• プロテインキナーゼＣ及びジアシルグリセロールキナーゼのライブイメージング

  Yasuhito Shirai, Naoaki Saito

  エヌ・テイー・エス, Jun. 2007


• ジアシルグリセロールキナーゼ

  Yasuhito Shirai, Naoaki Saito

  日本薬理学会誌, Mar. 2007


• プロテインキナーゼＣのターゲティング機構

  Yasuhito Shirai, Norio Sakai, Naoaki Saito

  共立出版, Nov. 2003


• 蛋白質・核酸･酵素 / プロテインキナーゼＣのターゲティング

  Shirai Yasuhito

  共立出版, 2003


• シグナル伝達 総集編

  Saito Naoaki, Sakai Norio, SHIRAI YASUHITO

  羊土社, Aug. 1999


• 細胞のシグナリング

  SHIRAI YASUHITO, Saito Naoaki

  中山書店, May 1999


• 無脊椎動物のホルモン

  SHIRAI YASUHITO, Aizono Yasuo

  学会出版センター, Dec. 1998

• Effects of adding FFAs on beef taste-traits analyzed by electronic taste sensing system and sensory evaluation

  Zhao Yanan, Di Yindi, Ueda Shuji, SHIRAI YASUHITO, HABARA MASAAKI, IKEZAKI HIDEKAZU, YAMANOUE MINORU

  日本畜産学会第125回大会, Mar. 2019, 麻布大学


• Methylated epigallocatechin gallate (EGCg) as functional food for diabetic nephropathy

  SHIRAI YASUHITO

  the 25th International conference of functional food center, Oct. 2018, Ritsumeikan Univ


• Effect of Zingiber Zerumbet Smith extract on thermotolerance

  Kobayashi H, Ueda S, Kato Y, Yun Sang Soon, Tsumagari R, Yamanoue M, SHIRAI YASUHITO

  the 25th International conference of functional food center, Oct. 2018, Ritsumeikan Univ


• Effects of intramuscular FFAs on beef taste-traits analyzed by electronic taste sensing system and sensory evaluation

  ZHAO YANAN, NAKADA YUSUKE, UEDA SYUJI, SHIRAI YASUHITO, ICHIMURA SAYAKA, YOSHIDA YUKA, HABARA MASAAKI, IKEZAKI HIDEKAZU, YAMANOUE MINORU

  The 64th ICoMST, Aug. 2018, Melbourne, Australia


• ジアシルグリセロールキナーゼをターゲットとする抗アレルギー薬の開発

  倉橋 航平, 菊永 佐紀子, 上田修司, 山之上稔, SHIRAI YASUHITO

  第133回日本薬理学会近畿部会, Jun. 2018


• alpha-tocopherol と67 kDa ラミニンレセプターの直接的相互作用の検証

  林 大輝, Varnavas Mouchlis, 王 柳青, 上田 修司, 山之上 稔, 芦田 均, Edward Dennis, SHIRAI YASUHITO

  第65回日本生化学会近畿支部例会, May 2018, 兵庫医大


• 味覚センサ分析および官能評価による牛肉呈味性への筋内脂肪の影響

  趙 婭楠, 中田悠介, UEDA SHUJI, SHIRAI YASUHITO, 市村 さやか, 吉田 由香, 羽原正秋, 池崎秀和, YAMANOUE MINORU

  第59回日本食肉研究会大会, Mar. 2018, 東京大学農学部


• 熟成中の牛肉の筋内脂肪変化に影響される呈味性の味覚センサおよび官能評価による分析 (第2報)

  趙 婭楠, 中田悠介, UEDA SHUJI, SHIRAI YASUHITO, 市村 さやか, 吉田 由香, 羽原正秋, 池崎秀和, YAMANOUE MINORU

  日本畜産学会第124回大会, Mar. 2018, 東京大学農学部


• ジアシルグリセロールキナーゼgammaを作用点とする分子標的機能性食品の開発に向けた研究

  倉橋 弘平, 菊永 佐紀子, UEDA SHUJI, YAMANOUE MINORU, SHIRAI YASUHITO

  日本農芸化学会関西支部第501回講演会, Dec. 2017, 神戸大学農学部


• 緑茶カテキン EGCg のメチル化による糖尿病性腎症改善効果の増強

  HAYASHI Daiki, UEDA Shuji, YAMANOUE Minoru, ASHIDA Hitoshi, SHIRAI Yasuhito

  日本農芸化学会 関西・中四国・西日本支部 2017年度合同大阪大会（第49回講演会）, Sep. 2017, Sakai


• 熟成中の牛肉における筋内脂肪変化に影響される呈味性の味覚センサおよび官能評価による分析

  趙 婭楠, 中田悠介, UEDA SHUJI, SHIRAI YASUHITO, 羽原正秋, 池崎秀和, YAMANOUE MINORU

  日本畜産学会第123回大会, Sep. 2017, 信州大学伊那キャンパス


• 質量分析計による黒毛和種牛肉の栄養成分のノンターゲット分析による検討

  UEDA SHUJI, 岩本英治, SHIRAI YASUHITO, YAMANOUE MINORU

  日本畜産学会第123回大会, Sep. 2017, 信州大学伊那キャンパス


• 黒毛和種牛肉の赤身部位の各種分析技術の検討

  UEDA SHUJI, 岩本 英治, 吉田 和利, SHINOHARA MASAKAZU, 中田 悠介, 高杉 瑠美, 岩本 英樹, 野村 郁代, 小川 隆文, SHIRAI YASUHITO, YAMANOUE MINORU

  日本畜産学会第122回大会, Mar. 2017, 神戸大学鶴甲第1キャンパス


• Importance of DG signaling in cerebellar motor coordination

  SHIRAI YASUHITO

  第80回 薬理学会年会, Mar. 2017, 長崎 ブリックホール


• Importance of functional correlation between protein kinase C and diacylglycerol kinase in cerebellar motor coordination

  SHIRAI YASUHITO

  The 1st Biosignal Research Center Int. Symposium, 2017, Jan. 2017, 神戸大学 六甲ホール


• 皮膚におけるジアシルグリセロールキナーゼγの機能解析

  菊永佐紀子, 横谷美由希, 藤原祥高, UEDA SYUJI, YAMANOUE MINORU, 高岡幸二, 伊川正人, SHIRAI YASUHITO

  日本農芸化学会関西支部例会 第497回講演会, Dec. 2016


• 皮膚におけるジアシルグリセロールキナーゼgammaの機能解析

  菊永佐紀子, 横谷美由紀, 藤原祥高, 上田修司, 山之上稔, 高岡幸二, 伊川正人, SHIRAI YASUHITO

  日本農芸化学会関西支部第497回講演会, Dec. 2016, 神戸大学


• Identification of novel RhoA binding proteins by using BioID system

  櫛谷 晃帆, 加藤 良毅, Soichiro Yamada, YOSHINO KEN-ICHI, 竹内 敦子, YAMANOUE MINORU, SHIRAI YASUHITO, UEDA SHUJI

  The American Society for Cell Biology 2016 ASCB/IFCB Meeting, Dec. 2016, Moscone Center San Francisco, California


• Identification of novel HSPB1 binding proteins in skeletal muscle

  加藤 良毅, 小鍛治 泰斗, 櫛谷 晃帆, YOSHINO KEN-ICHI, 竹内 敦子, YAMANOUE MINORU, SHIRAI YASUHITO, UEDA SHUJI

  The American Society for Cell Biology 2016 ASCB/IFCB Meeting, Dec. 2016, Moscone Center San Francisco, California


• Identification of novel RhoA binding protein by usinBioID system

  櫛谷晃帆, UEDA SHUJI, 加藤良毅, YOSHINO KEN-ICHI, 竹内敦子, YAMANOUE MINORU, SHIRAI YASUHITO

  第39回日本分子生物学会年会, Dec. 2016, パシフィコ横浜 展示ホール


• DGKγ contributes cerebellar motor coordination through regulation of protein kinase C

  SHIRAI YASUHITO, Ryousuke Tsumagari, Sakiko Kikunaga, Yoshitaka Fujihara, Shuji Ueda, Minoru Yamanoue, Sho Kakizawa, Hideaki Hara, Masahito Ikawa

  Neuroscience meeting 2016, Nov. 2016, San Diego Convention center


• 骨格筋におけるHSPB1結合蛋白質の探索と

  Kokaji Yoshito, Kato Yoshiki, Kushiya Akiho, YOSHINO KEN-ICHI, Takeuchi Atsuko, UEDA SHUJI, YAMANOUE MINORU, SHIRAI YASUHITO

  日本農芸化学会平成28年度関西支部大会, Sep. 2016, 滋賀県立大学（滋賀県彦根市八坂町2500）


• The elucidation of the mechanism of lentinan-induced inflammatory suppression by visualizimg TNFR1 on the membrane

  SAKAGUCHI KANA, HASHIMOTO TAKASHI, ITO TOSHIKI, SHIRAI YASUHIDE, MIZUNO MASASHI

  Functional and Medical Foods for Chronic Diseases: Bioactive Compounds and Biomarkers, Sep. 2016, Harvard Medical School


• Possibility of epigallocatechin gallate and a-tocopherol as functional food for diabetic nephropathy and their molecular mechanism

  SHIRAI YASUHITO

  20th International conference of Functional Food in health disease, Sep. 2016, Harbard medical school, Boston


• レンチナンによって誘導されるTNFR1発現変化の可視化による炎症抑制機構の解明

  SAKAGUCHI KANA, HASHIMOTO TAKASHI, ITO TOSHIKI, SHIRAI YASUHITO, MIZUNO MASASHI

  日本食品科学工学会第63回大会, Aug. 2016, 名城大学


• リチウムによるDGKbetaKOマウスの記憶及び感情障害の改善には、PIP2の減少とPKCgammaの活性抑制が関与している

  Wataru Okimoto, Horoko Nakai, Mitsue Ishisaka, Hiroki Nakanishi, Shigeki Moriguchi, Shuji Ueda, Minoru Yamanoue, Kouji Fukunaga, Takehiko Sasaki, Hideaki Hara, SHIRAI YASUHITO

  第79回脂質生化学会, Jun. 2016


• 筋肉の増加減退に関わるRhoAの機能解析と相互作用分子の探索

  櫛谷 晃帆, UEDA SHUJI, 加藤 良毅, 桐村 悠佑, 石政 碧, YOSHINO KEN-ICHI, 竹内 敦子, YAMANOUE MINORU, SHIRAI YASUHITO

  第６３回日本生化学会近畿支部例会, May 2016, 神戸薬科大学


• カテキン類によるジアシルグリセロキナーゼα活性化機構と、糖尿病性腎症の予防・改善の可能性について

  SHIRAI YASUHITO

  第70回日本栄養・食糧学会大会, May 2016, 武庫川女子大学


• Function of diacylglycerol kinase

  SHIRAI YASUHITO

  Lipid mediator in health and disease, May 2016, UCSD, San Diego


• Diacylglycerol kinase gamma KO mouse shows impairment of motor coordination

  津曲 涼介, 菊永 佐紀子, 藤原 祥高, 上田 修司, 山之上 稔, 伊川 正人, SHIRAI YASUHITO

  第８９日本薬理学会, Mar. 2016, パシフィコ横浜


• ジアシルグリセロールキナーゼalphaの核－細胞質間シャトリングの生理的意義の解明

  渡辺 真以, 木曽 裕子, 上田 修司, 山之上 稔, 齋藤 尚亮, SHIRAI YASUHITO

  農芸化学会関西支部例会493回講演会, Feb. 2016, 京都大学


• 神経系におけるジアシルグリセロールキナーゼgammaの機能解析

  津曲 涼介, 菊永 佐紀子, 藤原 祥高, 上田 修司, 山之上 稔, 伊川 正人, SHIRAI YASUHITO

  農芸化学会関西支部例会（492回講演会）, Dec. 2015, 神戸大学 農学部


• チロシンリン酸化がジアシルグリセロールキナーゼのドメイン間相互作用及び高次構造に与える影響

  脇阪 昌明, 岩下 智秋, 濱田 大三, 祇園 景子, 上田 修司, 山之上 稔, 鶴田 宏樹, SHIRAI YASUHITO

  農芸化学会関西支部例会（492回講演会）, Dec. 2015, 神戸大学 農学部


• ジアシルグリセロールキナーゼの218番目のチロシンリン酸化はドメイン間相互作用と高次構造を変化させる

  脇阪 昌明, 岩下 智秋, 濱田 大三, 祇園 景子, 上田 修司, 山之上 稔, 鶴田 宏樹, SHIRAI YASUHITO

  BMB2015, Dec. 2015, 神戸国際会議場


• Vitamin E activates diacylglycerol kinase alpha through 67LR and Src family tyrosine kinase

  Hayashi D, Wang L, Ueda S, Yamanoue M, Ashida H, SHIRAI YASUHITO

  annual meeting of ASCB 2015, Dec. 2015, San Diego convention center


• Mechanism of lithium-induced recovery of memory and emotional impairment in DGKbetaKO mice

  Okimoto W, Ishisaka M, Nakanishi H, Ueda S, Yamanoue M, Sasaki T, Hara H, SHIRAI YASUHITO

  Neuroscience meeting 2015, Dec. 2015, Chicago convention center


• Insulin-like growth factor-1 induces activation and upregulation of RhoA via PI3K/AKT/mTOR pathway in masucular hypertrophy mode of C2C12 cells

  Kushiya A, Ueda S, Kirimura Y, Katoh Y, Ishimasa M, Yamanoue M, Satoh T, SHIRAI YASUHITO

  annual meeting of ASCB 2015, Dec. 2015, San Diego convention center


• Functional analysis of diacylglycerol kinase theta deficient mice

  Koizumi N, Ueda S, Fujihara Y, Yamanoue M, Ikawa M, SHIRAI YASUHITO

  19th international conference of functional food center, Nov. 2015, 神戸大学 百年記念会館


• Effect of Vitamin E on diabetic nephropathy in DGKa knockout mice

  Hayashi D, Yagi K, Ueda S, Yamanoue M, Emoto N, Saito N, SHIRAI YASUHITO

  19th international conference of functional food center, Nov. 2015, 神戸大学 百年記念会館


• 細胞内ホスファチジン酸産生を時・空間的に解析できる簡便かつ新規な可視化プローブの開発

  沖本 航, 中井 寛子, 伊藤 俊樹, 森田 真也, 上田 修司, 山之上 稔, SHIRAI YASUHITO

  第５７回日本脂質生化学会, Jun. 2015, 東京


• 熟成中の牛肉筋漿における筋原線維Z線近傍コネクチン断片のELISA法による定量

  YAMANOUE MINORU, MARUYAMA MASAYA, UEDA SYUJI, SIRAI YASUHITO, SASAKI KANA

  第119回日本畜産学会大会, Mar. 2015, 日本畜産学会, 宇都宮大学


• Screening of novel diacylglycerol kinase a activator to develop drugs improving diabetic nephropathy

  Daiki Hayashi, Ke Kiu, Shuji Ueda, Minoru Yamanoue, Fumio Sakane, SHIRAI YASUHITO

  第８８回 日本薬理学会年会, Mar. 2015, 名古屋国際会議場


• Expression analysis of diacylglycerol kinase theta in mouse tissues by immunostaining and LacZ reporter assay

  UEDA SHUJI, Nao Koizumi, Minoru Yamanoue, Yasuhito Shirai

  The 88th Annual Meeting of The Japanese Pharmacological Society, Mar. 2015, Nagoya Congress Corporation, Diacylglycerol kinase (DGK) regulates diacylglycerol signaling, which controls the variety of cell function. DGKθ belongs to the sole member of type V DGK. Several in vitro studies have suggested important role of DGKθ, but the physiological role of the enzyme is still unclear. In this study, to reveal the spatial expression of DGKθ, we performed the immunostaining on paraffin


• DGKb as a possible therapeutic target for memory loss and mania

  SHIRAI YASUHITO

  第８８回 日本薬理学会年会, Mar. 2015


• DGKa欠損マウスにおける糖尿病性腎症に対するビタミンＥの効果

  林 大輝, 八木 敬子, 上田 修司, 山之上 稔, 江本 憲昭, 齋藤 尚亮, SHIRAI YASUHITO

  日本農芸化学会2015大会, Mar. 2015, 岡山大学


• Comprehensive understanding of lipids roles in life science

  SHIRAI YASUHITO, Edward Dennis

  UC San Diego x Kobe University joint research kick off symposium, Feb. 2015, Kobe Univ


• 脂質結合ドメインを用いた新規ホスファチジン酸可視化プローブの開発

  OKIMOTO WATARU, NAKAI HIROKO, ITOH TOSHIKI, UEDA SHUJI, YAMANOUE MINORU, SHIRAI YASUHITO

  第487回 日本農芸化学会関西支部例会, Dec. 2014, 神戸大学


• A role of RhoA activation in IGF-I-induced muscular hypertrophy model.

  Shuji Ueda, Soichiro Yamada, Yusuke Kirimura, Akiho Kushiya, Minoru Yamanoue, Takaya Satoh, SHIRAI YASUHITO

  ASCB meeting 2014, Dec. 2014, Philadelphia


• Novel aspects of galloyl catechins: development of functional food targeting on diacylglycerol kinase alpha to prevent and/or improve diabetic nephoropathy.

  Daiki Hayashi, Shuji Ueda, Minoru Yamanoue, Naoaki Saito, Hitoshi Ashida, SHIRAI YASUHITO

  17th international concerence of functional food center, Nov. 2014, San Diego


• 筋細胞におけるHSPB1の結合分子の探索とその機能解析

  KOKAJI YASUTO, UEDA SHUJI, YOSHINO KEN-ICHI, YAMANOUE MINORU, SHIRAI YASUHITO

  第87回日本生化学会大会, Oct. 2014, （公社）日本生化学会, 国立京都国際会館


• ガレート型カテキンだけでなくビタミンEも67kDaラミニンレセプターを介してジアシルグリセロールキナーゼαを活性化する

  HAYASHI Daiki, ASHIDA Hitoshi, UEDA Shuji, YAMANOUE Minoru, SHIRAI Yasuhito

  The 2014 Annual Conference of the Japan Society for Bioscience, Biotechnology and Agrochemistry in KANSAI, Sep. 2014


• Expression of diacylglycerol kinase theta during the organogenesis of mouse embryos

  UEDA Shuji, Tu-Sekine Becky, YAMANOUE Minoru, RABEN Daniel M, SHIRAI Yasuhito

  Lipid MAPS meeting, May 2014, LIPD MAPS, UCSD


• 鶏筋原線維のプロテアーゼ分解によるコネクチン20-kDa断片の生成

  YAMANOUE MINORU, SASAKI NODOKA, SASAKI KANA, UEDA SYUJI, SHIRAI YASUHITO

  第118回日本畜産学会大会, Mar. 2014, つくば国際会議場


• ジアシルグリセロールキナーゼｂ欠損マウスの異常行動に対するバルプロ酸の効果

  ISHISAKA MITSUE, TSURUMA KAZUHIRO, SHIMAZAWA MASAMITSU, SHIRAI YASUHITO, HARA HIDEAKI

  第87回 日本薬理学会年会, Mar. 2014, 仙台


• カテキン類によるジアシルグリセロールキナーゼα活性化機構の解明

  HAYASHI Daiki, ASHIDA Hitoshi, UEDA Shuji, YAMANOUE Minoru, SHIRAI Yasuhito

  The 2014 Annual Conference of the Japan Society for Bioscience, Biotechnology and Agrochemistry, Mar. 2014


• mTORはDGKβが活性依存的及び活性非依存的に誘起する神経突起伸長に関与する

  KANO TAKUYA, NAKAI HIROKO, NAKASHIMA AKIO, KIKKAWA USHIO, SAITO NAOAKI, TAKEI NOBUYUKI, UEDA SHUJI, YAMANOUE MINORU, SHIRAI YASUHITO

  第87回 日本薬理学会年会, Mar. 2014, 仙台


• カテキン類及びビタミンEは67kDaラミニンレセプターを介してジアシルグリセロールキナーゼαを活性化する (Chatechins and vitamin E activate diacylglycerol kinaseα through 67kDa laminin receptor)

  HAYASHI Daiki, ASHIDA Hitoshi, UEDA Shuji, YAMANOUE Minoru, SHIRAI Yasuhito

  The 87th annual meeting of the Japanese biochemical society, 2014


• 筋肉における低分子量ヒートショック蛋白質HSPB1の結合分子の探索

  小鍛冶 泰斗, 上田 修司, 吉野 健一, 山之上 稔, SHIRAI YASUHITO

  若手フロンティア研究会2013, Dec. 2013, 神戸大学


• 細胞周期及び核内ホスファチジン酸産生に及ぼすジアシルグリセロールキナーゼ阻害剤の影響

  山本 麻央, 丸山 量子, 木曽 裕子, 上田 修司, 山之上 稔, 齋藤 尚亮, SHIRAI YASUHITO

  日本分子生物学会年会, Nov. 2013, 神戸国際会議場


• クロマン環構造に着目した新規ジアシルグリセロールキナーゼα活性化剤の探索

  林 大輝, ASHIDA HITOSHI, UEDA SHUJI, YAMANOUE MINORU, SAITO NAOAKI, SHIRAI YASUHITO

  第124回日本薬理学会近畿部会, Nov. 2013, 日本薬理学会近畿部会, 京都ガーデンパレス


• Both C1 domain and a basic amino acid cluster at C-terminus are important for branching and neurite induction ability of DGKb

  SHIRAI YASUHITO, Takuya Kano, Takeshi Kouzuki, Shuji Ueda, Minoru Yamanoue, Fumio Sakane, Naoaki Saito

  neuroscience meeting 2013, Nov. 2013, San Diego


• The expression of diacylglycerol kinase theta during the organogenesis of mouse embryos

  UEDA Shuji, Tu-Sekine Becky, YAMANOUE Minoru, RABEN Daniel M, SHIRAI Yasuhito

  第86回日本生化学会大会, Sep. 2013, 日本生化学会, パシフィコ横浜


• Role of protein kinase C in histamine release

  SHIRAI YASUHITO

  第８６回 日本生化学会, Sep. 2013, 横浜パシフィコ


• Ｃ１ドメインとＣ末塩基性アミノ酸クラスターがDGKbによる神経突起伸長に重要である

  加野 拓也, 上月 健, 上田 修司, 山之上 稔, 坂根 郁夫, 齋藤 尚亮, SHIRAI YASUHITO

  ２０１３日本神経化学会, Jun. 2013, 京都国際会館


• ジアシルグリセロールキナーゼβによる特徴的な神経突起伸長にはcPKCも関与する

  NAKAI Hiroko, KANO Takuya, UEDA Shuji, YAMANOUE Minoru, SAITO Naoaki, SHIRAI Yasuhito

  第55回日本脂質生化学会大会, Jun. 2013, 日本脂質生化学会, 宮城県松島町


• ジアシルグリセロールキナーゼβ欠損マウスにおけるけいれん感受性の増加

  石坂光絵, 鶴間一寛, 嶋澤雅光, SHIRAI YASUHITO, 齋藤尚亮, 原英彰

  第86回 日本薬理学会年会, Mar. 2013, 福岡


• C2C12筋管のIGF-1刺激による筋肥大におけるRhoAの活性化機構の解析

  桐村悠祐, 上田修司, 山之上稔, 佐藤孝哉, SHIRAI YASUHITO

  第86回 日本薬理学会年会, Mar. 2013, 福岡


• 乾燥肌モデルマウス試験におけるパイナップル果実由来グルコシルセラミド経口摂取の皮膚機能改善効果及びin vitro腸管モデルへの影響

  湯浅弘樹, 西谷洋輔, SHIRAI YASUHITO, 馬場健史, 野嶋潤, 大戸信明, 桒原浩誠, 水野雅史

  第478回 日本農芸化学会関西支部, Feb. 2013, 京都


• The expression of diacylglycerol kinase theta during mouse embryonic development

  UEDA Shuji, Tu-Sekine Becky, YAMANOUE Minoru, RABEN Daniel M, SHIRAI Yasuhito

  The ASCB 2012 Annual meeting, Dec. 2012, American Society for Cell Biology, Moscone Center, USA


• DGKbetaによる神経突起伸張機構の解析

  KANO Takuya, UEDA Shuji, YAMANOUE Minoru, SAITO Naoaki, SHIRAI Yasuhito

  第85回日本生化学会大会, Dec. 2012, 日本生化学会, 福岡国際会議場


• DGKbetaによる神経突起伸張機構の解析

  加野拓也, 上田修司, 山之上稔, 齋藤尚亮, SHIRAI YASUHITO

  第85回 日本生化学会, Dec. 2012, 福岡


• Functional correlation between PKC and DGK in neurons

  Yasuhito Shirai

  KAIST&KIST, Nov. 2012, Soul (Korea)


• Role of PKCalpha-mediated cofilin phosphorylation in the degranulation from RBL-2H3 mast cells

  Megumi Sakuma, Yasuhito Shirai, Ken-ichiYoshino, Maho Kuramasu, Tomofumi Nakamura, Kensaku Mizuno, Izumi Hide, Yoshihiro Nakata, Naoaki Saito

  22ndIUBMB&37thFEBS, Sep. 2012, Iberia(Spain)


• Molecular mechanism of DGKβ- mediated neurite blanching and spine formation

  NAKAI Hiroko, TAKEI Nobuyuki, NAKASHIMA Akio, KIKKAWA USHIO, SAITO Naoaki, UEDA Shuji, YAMANOUE Minoru, SHIRAI Yasuhito

  The 35th Annual Meeting of the Japan Neuroscience Society, Sep. 2012, Japan Neuroscience Society, Nagoya Congress Center


• Molecular mechanism of DGKβ-mediated neurite blanching and spine formation

  Hiroko Nakai, Nobuyuki Takei, Akio Nakashima, Ushio Kikkawa, Naoaki Saito, Shuji Ueda, Minoru Yamanoue, Yasuhito Shirai

  Neuro2102, Sep. 2012, Nagoya


• Involvement of mTOR in Diacylglycerol kinase β (DGKβ)-induced neurite branching and spine formation.

  NAKAI Hiroko, NAKASHIMA Akio, KIKKAWA USHIO, SAITO Naoaki, TAKEI Nobuyuki, UEDA Shuji, YAMANOUE Minoru, SHIRAI Yasuhito

  The 22nd IUBMB & the 37th FEBS, Sep. 2012, IUBMB & FEBS, Sevilla, Spain


• Involvement of mTOR in Diacylglycerol kinase β (DGKβ)-induced neurite branching and spine formation

  Hiroko Nakai, Akio Nakashima, Ushio Kikkawa, Naoaki Saito, Nobuyuki Takei, Shuji Ueda, Minoru Yamanoue, Yasuhito Shirai

  22ndIUBMB&37thFEBS, Sep. 2012, Iberia(Spain)


• Functional correlation between PKC and DGK in neuronal morphological change including branching and spine formation

  Yasuhito Shirai

  Neuro2102, Sep. 2012, Nagoya


• Analysis of tissue distribution of diacylglycerol kinase theta

  UEDA Shuji, KIRIMURA Yuji, YAMANOUE Minoru, SHIRAI Yasuhito

  The 14th International Congress of Histochemistry and Cytochemistry, Aug. 2012, International Federation of Societies for Histochemistry and Cytochemisty, Kyoto International Conference Center


• Analysis of tissue distribution of diacylglycerol kinase theta

  Shuji Ueda, Yusuke Kirimura, Minoru Yamanoue, Yasuhito Shirai

  14th International Congress of Histochemistry and Cytochemistry, Aug. 2012, kyoto


• ジアシルグリセロールキナーゼβによる特徴的な神経突起伸にはmTORが関与する

  SHIRAI YASUHITO, 中井寛子, 中西広樹, 中嶋昭雄, 吉川潮, 上田修司, 山之上稔, 斎藤尚亮

  第54回日本脂質生化学会, Jun. 2012, 福岡


• ジアシルグリセロールキナーゼ_による特徴的な神経突起伸にはmTORが関与する

  SHIRAI Yasuhito, NAKAI Hiroko, NAKANISHI Hiroki, NAKASHIMA Akio, KIKKAWA Ushio, UEDA Shuji, YAMANOUE Minoru, SAITO Naoaki

  第54回日本脂質生化学会大会, Jun. 2012, 日本脂質生化学会, 九州大学


• 食肉の日齢評価に利用可能なタンパク質マーカーの探索

  清水俊作, 上田修司, 桐村悠佑, 本田和久, 上曽山博, SHIRAI YASUHITO, 山之上稔

  日本畜産学会, Mar. 2012, 名古屋


• 筋肉細胞におけるEGCgの糖代謝に及ぼす影響について

  上田学, 上田修司, SHIRAI YASUHITO, 芦田均

  日本農芸化学会2012年度大会, Mar. 2012, 京都


• ジアシルグリセロールキナーゼθの機能解析

  UEDA Shuji, KIRIMURA Yuji, YAMANOUE Minoru, SHIRAI Yasuhito

  日本農芸化学会2012年度大会, Mar. 2012, 日本農芸化学会, 京都女子大学


• ジアシルグリセロールキナーゼthetaの機能解析

  上田修司, 桐村悠佑, 山之上稔, SHIRAI YASUHITO

  日本農芸化学会2012年度大会, Mar. 2012, 京都


• The role of diacylglycerol kinase in the degranulation from RBL-2H3 mast cells

  M. Sakuma, Shirai Yasuhito, T. Ueyama, N. Saito

  第85回 日本薬理学会年会, Mar. 2012, 京都


• PKCetaによるケラチノサイト分化機構の解明

  SHIRAI YASUHITO, 盛岡祥子, 佐久間恵, 千田和広, 斎藤尚亮

  日本農芸化学会2012年度大会, Mar. 2012, 京都


• Physiological function of nuclear DGKα under serum depleted conditions

  Y. Kiso, Shirai Yasuhito, M. Sakuma, M. Ikeda, N. Saito

  第85回 日本薬理学会年会, Mar. 2012, 京都


• Diacylglycerol kinase as a possible drug target

  Shirai Yasuhito, N. Saito

  第85回 日本薬理学会年会, Mar. 2012, 京都


• 筋肉細胞におけるEGCgの糖代謝に及ぼす影響について

  上田 学UEDA Manabu, UEDA Syuji, SHIRAI Yasuhito, ASHIDA Hitoshi

  日本農芸化学会2012年度大会, 2012, 京都


• Novel. phosphorylation site(s) of cofilinby PKCα and its negative role in the degranulation from RBL-2H3 mast cells

  M. Sakuma, Shirai Yasuhito, K. Yoshino, M .Kuramasu, T. Nakamura, K. Mizuno, I Hide, Y. Nakata, N. Saito

  2011 ASCB Annual Meeting, Dec. 2011, アメリカ・ニューオリンズ


• Function of PKC and DGK in neurons

  SHIRAI YASUHITO

  2011 Taiwan-Japan Joint Symposium on Cell Signaling and Gene Regulation, Nov. 2011, 台湾・台南


• ジアシルグリセロールキナーゼβ（DGKβ）による神経特異的突起伸長機構の解明

  中井寛子, 加野拓也, 上月健, 斉藤尚亮, 中嶋昭雄, 吉川潮, 上田修司, 山之上稔, SHIRAI YASUHITO

  第3回神戸大学バイオサイエンス研究会・若手研究者交流会, Sep. 2011, 神戸


• Regulation of diacylglycero kinase by phosphorylation

  SHIRAI YASUHITO

  Fifty-Second International Symposium on “Regulation of Enzyme Activity and Synthesis in Normal and Neoplastic Tissues”, Sep. 2011, イタリア・ボローニャ


• Novel PKCαphosphorylation site(s) contribute to proper termination of histamine release

  M Sakuma, Shirai Yasuhito, K Yoshino, M Kuramasu, T Nakamura, K Mizuno, I Hide, Y Nakata, N Saito

  第８４回日本生化学会, Sep. 2011, 京都


• 異なるチロシンリン酸化によるジアシルグリセロールキナーゼalphaの局在調節機構

  SHIRAI YASUHITO, 池田 もも, 佐久間 恵, 木曽 裕子, 斎藤 尚亮

  第119回 薬理学会近畿支部大会, Jul. 2011, 名古屋


• ジアシルグリセロールキナーゼαの核移行と細胞周期制御に関する研究２－c-Ablによるチロシンリン酸化サイトの同定－

  SHIRAI YASUHITO, 池田 もも, 木曽 裕子, 佐久間 恵, 齋藤 尚亮

  第53回日本脂質生化学会, May 2011, 東京


• 血清によるDGKαの核外移行におけるc-Ablによるリン酸化サイトの同定

  池田もも, SHIRAI YASUHITO, 木曽 裕子, 佐久間 恵, 斎藤 尚亮

  第84回 日本薬理学会年会（但し地震のため紙上開催）, Mar. 2011, 横浜


• PKCetaによるケラチノサイト扁平化機構～PKCeta とRalAの結合とケラチノサイト扁平化における役割り～

  SHIRAI YASUHITO, 斎藤 尚亮

  第84回 日本薬理学会年会（但し地震のため紙上開催）, Mar. 2011, 横浜


• Direct binding of RalA to PKCeta and its crucial role in morphological change during keratinocytes differentiation

  SHIRAI YASUHITO

  神戸大学ブリュッセルオフィイスオープニング記念シンポジウム, Mar. 2011, ベルギー・ブリュッセル


• Impairment of cognitive function and lithium-sensitive behavior of knockout mice of neuron-enriched subtype of DG kinase, DGKｂ

  Shirai Yasuhito, T Kouzuki, K Kakefuda, A. Ohyagi, K. Horie, S. Moriguchi, S. Morita, M. Shimazawa, K. Fukunaga, J. Takeda, N. Saito, H. Hara

  First annual meeting of International Society of Radiation Neurobiology., Jan. 2011, Maebashi


• Nuclear diacylglycerol signaling regulated by nuclear-cytoplasmic shuttling of diacylglycerol kinaseα and its correlation to cell cycle control

  Shirai Yasuhito, N Saito

  BMB2010 (兼 第83回日本生化学会大会), Dec. 2010, English, Kobe


• Correlation between abnormal behavior and lipids contents of DGKβ KO mice. The 5th international mini-symposium on diacylglycerol kinase (DGK)

  Shirai Yasuhito

  5th mini-symposium on DGK, Dec. 2010, English, Kobe


• Visualization of protein kinase C and diacylglycerol kinase in living cells

  Shirai Yasuhito, N. Saito

  28th JES summer seminar on Endocrinology & Metabolism, Jul. 2010, English, nagasaki


• Membrane lipids control by neuron-enriched diacylglycerol kinaseb is important for spine formation and cognitive function

  Shirai Yasuhito, T. Kouzuki, K. Kakefuda, S. Moriguchi, A. Oyagi, K. Horie, S. Morita, M. Shimazawa, K. Fukunaga, J. Takeda, N. Saito, H. Hara

  Keystone symposia “Bioactive lipids: biochemistry and diseases", Jun. 2010, English, kyoto


• 神経特異的ジアシルグリセロールキナーゼDGK betaの機能解析

  上月 健, Shirai Yasuhito, 掛札 賢一, 大八木 篤, 堀江 恭二, 嶋澤 雅光, 福永 浩司, 竹田 潤二, Saito Naoaki, 原 英彰

  第82回日本薬理学会年会, Mar. 2009, Japanese, パシフィコ横浜


• 気分障害と神経特異的ジアシルグリセロール

  Shirai Yasuhito

  第82回日本薬理学会年会, Mar. 2009, Japanese, パシフィコ横浜, Domestic conference

  Invited oral presentation


• 神経特異的ジアシルグリセロールキナーゼDGKの機能解析

  Shirai Yasuhito, 上月 健, 掛札 賢一, 大八木 篤, 堀江 恭二, 嶋澤 雅光, Saito Naoaki

  第31回日本分子生物学会年会・第81回日本生化学大会合同大会, Dec. 2008, Japanese, 神戸, Domestic conference

  Poster presentation


• RalAはeta PKCによるケラチノサイトの扁平化に必須である

  Shirai Yasuhito, 盛岡 祥子, 大辻 千裕, Yoshino Ken-ichi, 酒井 規雄, Ueyama Takehiko, Saito Naoaki

  第113回日本薬理学会近畿部会, Jun. 2008, Japanese, 岡山, Domestic conference

  Oral presentation


• Differential regulation of dual oxidase 1 by dual activator 1 splicing variants.

  Tsujibe Satoshi, Ueyama Takehiko, Shirai Yasuhito, Leto L. Thomas, Saito Naoaki

  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


• Clarification of regulatory mechanism of Orail(SOCs)by PKC.

  Kawasaki Takumi, Ueyama Takehiko, Shirai Yasuhito

  14th Congress of Ca2+ binding proteins & Ca2+ function in health & disease, Oct. 2007, English, La Palma Spain, International conference

  Poster presentation


• c-Ab1, a tyrosine kinase involved in chronic myeloid leukemia, regulates nucleo-cytoplasmic shuttling of DGK alpha and cell cycle.

  Shirai Yasuhito, Matsubara Takehiro, Saito Naoaki

  FASEB Summer Research Conferences, Aug. 2007, English, Indian Wells CA, International conference

  Poster presentation


• 脂質代謝酵素DGKの核―細胞質問shuttling機構と細胞周期制御への関与

  松原 岳大, 白井 康仁, 齋藤 尚亮

  第49回日本脂質生化学会, Jun. 2007, Japanese, 北海道大学, Domestic conference

  Oral presentation


• 脂質代謝酵素DGK alphaの核―細胞質問shuttling機構と細胞周期制御への関与

  Matsubara Takehiro, Shirai Yasuhito, Saito Naoaki

  第49回日本脂質生化学会, Jun. 2007, Japanese, 第49回日本脂質生化学会, Domestic conference

  Oral presentation


• 核内脂質シグナル及び核移行蛋白質の可視化システムを用いた細胞周期機構の解明－DGKの核－細胞質間移行機構と核内における機能に着目して－

  Shirai Yasuhito, Matsubara Takehiro, Saitou Naoaki

  第49回日本脂質生化学会, Jun. 2007, Japanese, 北海道大学, Domestic conference

  Oral presentation


• ヒトケラチノサイトにおけるTNF-によるNGF分泌促進作用とそのシグナル伝達機能

  高岡 幸二, 白井 康仁, 齋藤 尚亮

  第111回日本薬理学会近畿部会, Jun. 2007, Japanese, 名古屋, Domestic conference

  Oral presentation


• DG-PKCシグナル系の可視化とその異常による疾患

  Saito Naoaki, Shirai Yasuhito, Matsubara Takehiro, Adachi Naoko, Takahashi Hideyuki, Sakai Norio

  第49回日本脂質生化学会, Jun. 2007, Japanese, 北海道大学, Domestic conference

  Oral presentation


• 表皮細胞ケラチノサイトの分化におけるPKCの機能解析

  Morioka Shoko, Shirai Yasuhito, Otsuji Chihiro, Kashiwagi Kaori, Sakai Norio, 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

  Poster presentation


• 表皮細胞ケラチノサイトの分化における eta PKCの機能解析

  Morioka Shoko, Shirai Yasuhito, 大辻 千裕, Kashiwagi Kaori, Sakai Norio, 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

  Poster presentation


• 新規蛍光蛋白システムを用いたphox蛋白内の特異的結合の解明：p40phoxの分子内結合に注目して

  Kusakabe Tomoko, Shimizu Aya, Ueyama Takehiko, Kawasaki Takumi, Shirai Yasuhito, 唐澤 智司, Miyawaki Atsushi, 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


• 新規蛍光蛋白システムを用いたphox蛋白内の特異的結合の解明：p40phoxの分子内結合に注目して

  Kusakabe Tomoko, Shimizu Aya, Ueyama Takehiko, Kawasaki Takumi, Shirai Yasuhito, Karasawa Tomoshi, 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


• RBL-2H3細胞の脱顆粒におけるPKCとアクチンの機能協関解析

  Kuramasu Maho, Shirai Yasuhito, Nakamura Tomofumi, Hide Izumi, Nakata Yoshihiro, 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

  Poster presentation


• The mechanism of nucleo-cytoplasmic shuttling of diacylglycerol kinaseα.

  Shirai Y, Matsubara T, Saito N

  International Symposium Functional organization of the nucleus, Jan. 2007, English, 淡路夢舞台（兵庫）, International conference

  Oral presentation


• The mechanism of nucleo-cytoplasmic shuttling of diacylglycerol kinase alpha.

  Shirai Yasuhito, Matsubara Takehiro, Saito Naoaki

  International Symposium Functional organization of the nucleus, Jan. 2007, English, Hyougo, Japan, International conference

  Oral presentation


• ジアシルグリセロールキナーゼによるクラスリン依存性エンドサイトーシスの制御

  Kawasaki Takumi, Kobayashi Tsuyoshi, 上山 健彦, Shirai Yasuhito, Saito Naoaki

  第110回日本薬理学会近畿部会, Nov. 2006, Japanese, ぱるるプラザ京都, Domestic conference

  Oral presentation


• Spatial and temporal controls of PKC isoform activation.

  Shirai Yasuhito, Saito Naoaki

  International Union of Pharmacology, Jul. 2006, English, 北京五洲大酒店（中国）, International conference

  Invited oral presentation


• プロテインキナーゼCεのフォスファチジルイノシトール２リン酸結合能と神経突起伸長能との相関

  Shirai Yasuhito, Murakami Takuya, Kuramasu Maho, Saito Naoaki

  第48回日本脂質生化学会, Jun. 2006, Japanese, 東京大学医科学研究所, Domestic conference

  Oral presentation


• The Interaction between the Domains of DGK & gamma.

  Sanse Koichi, Shirai Yasuhito, Matsubara Takehiro, Yamaguchi Yasuto, Saito Naoaki

  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


• 糖尿病性腎症改善薬のターゲットとしてのジアシルグリセロールキナーゼ(DGK)

  SHIRAI Yasuhito, SAITO Naoaki

  第79回日本薬理学会年会, Mar. 2006, Japanese, Domestic conference

  Oral presentation


• NADPHオキシダーゼ複合体の形成メカニズムの解析

  TATSUNO Toshihiko, UYAMA Takehiko, TUJIBE Satoshi, SHIRAI Yasuhitp, SAITO Naoaki

  第79回日本薬理学会年会, Mar. 2006, Japanese, Domestic conference

  Oral presentation


• クラスリン依存性エンドサイトーシスにおけるDGキナーゼの役割

  Kawasaki Takumi, Kakehi Tomoko, Shirai Yasuhito, SAITO Naoaki

  第28回日本分子生物学会, Dec. 2005, Japanese, Domestic conference

  Oral presentation


• 発達期のラットの脳におけるジアシルグリセロールキナーゼ、サブタイプの発現変化.

  ADACHI Naoko, OKUBO Miho, TANIGUCHI Taizo, YAMAGUCHI Yasuhito, TAKENAKA Rika, SHIRAI Yasuhito, SAITO Naoaki

  第４６回日本組織細胞化学会, Oct. 2005, Japanese, Domestic conference

  Oral presentation


• DGKa の核局在制御と細胞周期

  MATSUBARA Takehiro, SHIRAI Yasuhito, SATANI Hideyuki, SAITO Naoaki

  第78回日本生化学会大会, Oct. 2005, Japanese, Domestic conference

  Oral presentation

• 北米神経科学会


• 日本脂質生化学会


• 日本神経科学会


• 日本薬理学会


• 日本生化学会

• DGキナーゼを介した食品成分による糖尿病性腎症改善機構の解明とその応用

  白井 康仁

  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, 01 Apr. 2020 - 31 Mar. 2023


• Intracellular lipid signaling analysis for the development of novel diabetes therapeutic agents targeting neogenesis and proliferation of pancreatic beta-cells

  Ishikawa Tomohisa

  Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (B), University of Shizuoka, 01 Apr. 2017 - 31 Mar. 2020

  Our previous study showed that an increased number of small islets was observed in the pancreas of β-cell-specific diacylglycerol kinase δ (DGKδ) knockout (βDGKδ KO) mice. Based on these findings, we investigated the mechanism for the increase in β-cell mass due to DGKδ deficiency and the possibility whether the suppression of DGKδ improves hyperglycemia. We found that that DGKδ knockdown in MIN6 β-cells showed a significant increase in proliferation, accompanied with an increased expression of cell-cycle related genes. In addition, we confirmed that streptozotocin-induced hyperglycemia and β-cell loss were alleviated in βDGKδ KO mice. These results suggest that the suppression of DGKδ in β-cells leads to an increased β-cell mass via proliferation, thereby improving hyperglycemia.


• 小腸上皮多糖受容体を介した自然免疫修飾を利用した健康増強に資する高機能性食品開発

  水野 雅史

  科学研究費補助金／基盤研究(B), Apr. 2015 - Mar. 2018

  Competitive research funding


• 酵素の細胞内局在調節機構の包括的解明－ＤＧキナーゼを例として－

  白井 康仁

  学術研究助成基金助成金／基盤研究(C), Apr. 2015 - Mar. 2018, Principal investigator

  Competitive research funding


• Dynamical study of multi-domain kinases reconstructed by domain ligation

  Mishima Masaki

  Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Challenging Exploratory Research, Tokyo Metropolitan University, 01 Apr. 2015 - 31 Mar. 2017

  It is generally recognized that kinases consist of multiple domain (mutli-domain proteins), which have flexible linker(s). This flexibility may hamper crystallization. In this study, our purpose is visualization of structural rearrangements of multi-domain kinases (transition from inhibited inactive states to activated states) using solution techniques at molecular level to understand the regulation mechanism. We have succeeded in domain ligation of C1B domain with kinase domain of protein kinase C using sortase. Further, we synthesized DOTA-M8, a rigid chelator for lanthanide ions, to measure PCS to obtain long-range distance information. It contains maleimide group which is useful to attach to proteins. Using this chelator, we actually measured PCS.


• 生命現象を形作る生体組織の巧みな構造－瀬戸内の豊かな食材を例に学ぶ生物の「超」ミクロの世界－

  白井 康仁

  独立行政法人日本学術振興会, 研究成果の社会還元・普及事業 ひらめき☆ときめきサイエンス, 2017, Principal investigator

  Competitive research funding


• 細胞・組織が作り出す生命の多様な構造－顕微鏡観察で学ぶミクロの世界－

  白井 康仁

  ひらめき☆ときめきサイエンス, 2015, Principal investigator

  Competitive research funding


• 生命現象を形作る生体組織の巧みな構造-地元食材から学ぶ細胞・組織のミクロの世界-

  白井 康仁

  ひらめき☆ときめきサイエンス, 2014, Principal investigator

  Competitive research funding


• A-STEP「ＰＫＣ及びＤＧキナーゼをターゲットとする新規抗ヒスタミン薬の開発と分子標的機能性食品への応用」

  白井 康仁

  研究成果最適展開支援プログラム フィージビリティスタディステージ 探索タイプ, 2013, Principal investigator

  Competitive research funding


• A-STEP「ＰＫＣ及びＤＧキナーゼをターゲットとする新規抗ヒスタミン薬の開発と分子標的機能性食品への応用」

  白井 康仁

  研究成果最適展開支援プログラム フィージビリティスタディステージ 探索タイプ, 2012, Principal investigator

  Competitive research funding


• ＰＫＣの機能破綻により発症する神経変性疾患モデル動物作製と解析および創薬への応用

  齋藤 尚亮

  科学研究費補助金／基盤研究(B), 2009

  Competitive research funding


• ＰＫＣとの機能協関に着目したＤＧキナーゼの機能解析とその応用

  白井 康仁

  科学研究費補助金／基盤研究(C), 2009, Principal investigator

  Competitive research funding


• プロテインキナーゼＣシグナル系路の異常による疾患の発症メカニズムの解明とその応用

  齋藤 尚亮

  科学研究費補助金／基盤研究(B), 2007

  Competitive research funding


• 蛋白質のシグナル伝達機能

  吉川 潮

  2006

  Competitive research funding


• ＤＮＡ Ｖａｃｃｉｎａｔｉｏｎによる膜機能蛋白に対する抗体作成

  谷口 泰造

  科学研究費補助金／萌芽研究, 2005

  Competitive research funding


• 蛋白質のシグナル伝達機能

  吉川 潮

  2005

  Competitive research funding


• 細胞膜リン脂質の動的代謝による細胞応答誘導の分子機構

  齋藤 尚亮

  科学研究費補助金／基盤研究(B), 2005

  Competitive research funding


• 活性酸素産生機構における脂質シグナルのダイナミックイメージング解析

  齋藤 尚亮

  科学研究費補助金／特定領域研究, 2005

  Competitive research funding


• 核内脂質シグナル及び核移行蛋白質の可視化システムを用いた細胞周期制御機構の解明

  白井 康仁

  科学研究費補助金／特定領域研究, 2005, Principal investigator

  Competitive research funding


• 小脳長期抑圧現象におけるプロテインキナーゼCおよびDGキナーゼの時空間解析

  齋藤 尚亮, 白井 康仁, 柏木 香保里

  日本学術振興会, 科学研究費助成事業, 特定領域研究, 神戸大学, 2004 - 2004

  本研究では、PKCとDGK分子機能相関を明らかにした上で、小脳スライスのLTDに対してPKCとDGKの結合および相互機能修飾がどのように働くかを、電気生理学的現象と同時にライブイメージングを用いて神経可塑性における分子メカニズムを明らかにすることを目的とした。 その結果、(1)GFP標識PKCを発現する遺伝子操作動物を作製した。この動物においてはGFP標識PKCの脳内の発現部位および発現時期がともに制御可能であり、各脳部位でのPKC分子のライブイメージングが可能であるだけでなく、各脳部位でのPKC発現による行動変化の解析も可能である。今回、この動物の小脳スライスを用いて、Purkinje細胞におけるgammaPKC-GFPのイメージングを行ったところ、平行線維刺激によっては、gammaPKC-GFPの細胞膜へのトランスロケーションが、樹状突起遠位から近位へと伝播する様子が認められた。この事実は、シナプスを介した刺激によりPKCの活性化がニューロン内で伝播されることを示しており、LTDの分子メカニズムを理解する上で非常に興味深い。(2)小脳Purkinje細胞に共存する2種類のリン酸化酵素PKCおよびDGK分子間の機能的協関を解析した。PKCとDGKは、PKCの活性化とともに結合し、DGKはPKCによりリン酸化されることが明らかとなった。Mass spectrometryにより、DGKのリン酸化部位を決定し、変異体を作製した結果、PKCによるDGKのリン酸化は、DGK活性の上昇を導くことが明らかになった。これらの結果は、受容体刺激によるDG産生およびCa2+上昇を介したPKCの活性化は、DGKの細胞膜上でのPKCとの結合を引き起こし、そこでのDGKのPKCによるリン酸化による活性化を導く。このDGKの活性化は細胞膜上のDG量を減少させ、PKCの活性化を終了させると推測され、LTDの分子機構にも重要なメカニズムであると考えられる。


• PKCターゲティングのFRETによる抗癌剤のハイスループットスクリーニングの開発

  齋藤 尚亮, 白井 康仁, 柏木 香保里

  日本学術振興会, 科学研究費助成事業, 萌芽研究, 神戸大学, 2003 - 2004

  PKCの多彩な機能は、10種類以上のサブタイプが、種々の細胞害外刺激に応じて、細胞内の目的とする部位に移動し、特定の基質をリン酸化するという機能(ターゲティング機能)を有するためであると考えられてきている。 本年度の研究では、種々の合成ホルボールエステルによるPKC各サブタイプのターゲティングをスクリーニングすることにより、特定のPKC機能の阻害剤(活性化剤)を見出すことを目的とした。また、種々の病態における各PKCサブタイプの役割についても検討した。その結果、(1)特定のPKCサブタイプを特異的に活性化する合成ホルボールエステルを同定した。このホルボールエステルはin vitroでは特異性を示さないものの、in vivoではbetaPKCを特異的に活性化する働きを有した。(2)脳虚血による脳神経細胞死に対して予防作用を持つエストロゲンは、PKC特にgammaPKCを介して、その効果を発揮することを示した。また、その神経保護作用は従来の核型エストロゲン受容体ではなく、細胞膜に存在する新しいエストリロゲン受容体を介することが明らかとなった。(3)VEGF受容体を介したERKの活性化機構にはdeltaPKCの活性化が必須であることを示した。(4)ミクログリア細胞によるドン食作用には2種のPKC、epsilonおよびbetaが関与しているが、それぞれ、異なる機構に関与することを示した。つまり、活性酸素産生に関与するbetaPKCと、ドン食運動に関与するepsilonである。 以上の結果から、細胞種、細胞機能により機能するPKCが異なっており、各生理作用に関与するPKCサブタイプを決定し、さらにサブタイプ特異的なPKC作用薬の開発を行うことにより、副作用の少ない薬物の開発が可能となることを示した。


• Analysis of higher structure of fatty aid -responsible domains in PKC and DGK

  SHIRAI Yasuhito, SAITO Naoaki, TANIGUCHI Taizo

  Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), Kobe University, 2003 - 2004

  In the series of the experiments to study higher structure of domains responsible for fatty acids in PKC and DGK we first tried to identify the domains responsible for arachidonic acid (AA) in PKC and DGK. The C1B, but not C1A, domain was necessary for AA-induced plasma membrane targeting of DGKγ. Similarly, the C1B domain was involved in the AA-induced targeting of εPKC to the Golgi complex. On the other hand, either C1A or C1B was enough for arachidonic acid-mediated retention of γPKC on the plasma membrane. To use NMR, we made fusion protein of DGKγ C1A or C1B peptide to GST or MBP. Finally, we obtained about 0.5 mg of MBP-DGKγ C1A or MBP-DGKγ C1B but got only small amount of GST-DGKγ C1A and GST-DGKγ C1B. Therefore, MBP-DGKγ C1A and MBP- DGKγ C1B were treated with a protease, Factor Xa, and then applied to gel filtration using Superose 6HR. However, we could not obtain enough amount of both peptides. To monitor small amount of the peptides by anti-FLAG antibody, we produced MBP-DGKγ C1A-FLAG and MBP-DGKγ C1B-FLAG, and then we found both DGKγ C1A-FLAG and DGKγC1B-FLAG were eluted in the void volume, suggesting the peptides aggregated. The aggregation was seen in the cases of DGKγC1-FLAG, εPKCC1B-FLAG and εPKCC1-FLAG. In addition, we could not detect the binding of MBP-εPKCC1B and MBP-DGKγ C1A to phorbol ester. These results suggest that higher structure of the MBP-fusion proteins are not intact. Another approaches to the C1 peptides for NMR, i.e., HA tag or Baculo virus system, should be considered.


• CキナーゼとDGキナーゼの神経情報伝達におけるクロストーク機構のイメージング解析

  齋藤 尚亮, 白井 康仁

  日本学術振興会, 科学研究費助成事業, 特定領域研究, 神戸大学, 2002 - 2002

  【緒言】プロテインキナーゼC(PKC)は、神経可塑性を調節していることが報告されているが、神経可塑性に関与する情報伝達経路の中でどのPKCサブタイプが、いつ、どこで、どのような役割を果たしているかという時空間レべルの解明は、未だ行われていない。我々は、GFP融合PKCを用いた検討により、PKCには種々に刺激に応じて異なる蛋白質へターゲットする機構(ターゲティング機構)があることも報告し、このターゲティング機構が神経可塑性においても重要であることを示唆してきた。本年度の研究において、神経におけるPKCターゲティング機構の解明、および、PKCターゲティングを制御する因子であるDGKとの関連の解明を目的として研究を行った。 【結果と考察】1)脂質メッセンジャーであるアラキドン酸およびセラミドはPKCepsilon(前シナプス型サブタイプ)およびPKCdelta(痛覚路発現サブタイプ)のC1Bドメインに作用し、サブタイプ特異的にゴルジ体にトランスロケーションさせること、2)PKCの活性化をともなわないターゲティングによっても、細胞死などの現象を引き起こすこと、3)PKCgammaとDGKgammaは、カルシウム依存性に細胞内で直接結合し、お互いの機能を制御しうること、を見出した。これらの結果より、酵素の活性化とは関係なくPKCターゲティングが生理機能をもつこと、および、このPKCのターゲティングは調節する薬物のターゲットとして、PKC epsiolonのC1Bドメイン、DGKgammaなどが候補となることが示唆された。さらに、GFP標識した各PKCサブタイプを脳部位特異的、時期特異的に発現可能なトランスジェニックマウスを作成し、各脳部位でのPKCのターゲティングと電気生理現象の関連を解析を可能とした。脳スライスにおけるPKCターゲティングは培養細胞において観察されたものと異なっていることから、神経可塑性におけるPKCおよびDGKの機能を解析するためには、脳スライス内を用いたより生理的な条件下でのPKCターゲティングの検討が必要であることが示唆された。


• CキナーゼとDGキナーゼの脂質性シグナルによる標的認識機構の解明

  白井 康仁

  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, 2001 - 2002

  プロテインキナーゼC(PKC)及びジアシルグリセロールキナーゼ(DGK)の脂質性シグナルによるサブタイプ特異的標的認識(ターゲティング)機構を解明する目的で、εPKC及びDGKγの変異体とGFPとの融合蛋白質を作製し、脂質性シグナルによる動態をコンフォーカルレーザー顕微鏡で観察した。その結果、1,C1Bドメインを変異(欠損または点変異)させたεPKCは、アラキドン酸(AA)及びセラミドによるゴルジ体へのターゲティングが消失したことから、これらの脂質性シグナルによるターゲティングにはC1Bドメインが関与していることが明らかになった。 2,δPKCは、セラミドによりゴルジ体へターゲティングされるが、AAには応答しない。そこで、εPKCのC1BドメインをδPKCのC1Bに入れ替えたキメラを作製したところ、AAに応答しなくなった。反対に、εPKCのC1BドメインをもつキメラδPKCは、AAに応答するようになった。これらのことから、εPKCとδPKCのAAに対する応答能の違いは、それぞれのC1Bが決定づけていることが明らかになった。 3,DGKγはAA及びTPAにより細胞質膜へターゲティングされる。C1Bドメインを欠損させるとAAによるターゲティングは消失したが、C1Aドメインを欠損させてもAAによるターゲティング能は有していた。一方、C1Aドメインを欠損させるとAAに対する応答能は保持していたが、TPAには応答しなくなった。このことから、DGKγのAA応答にはC1Bドメインが、TPA応答にはC1Aドメインが重要でり、TPAとAAに応答するモチーフはそれぞれ独立していることが明らかになった。


• Spatio-temporal signaling mechanism of protein kinase C

  SAITO Naoaki, SAKAI Norio, SHIRAI Yasuhito

  Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (B), KOBE UNIVERSITY, 2001 - 2002

  PKC consists of more than 10 isoforms and the existence of multiple isoforms strongly suggests that each isoform has individual functional role in signal transduction. We have been investigated "When, Where, Which isoform of PKC is activated and How the cellular function is regulated by the activation" by using green fluorescent protein(GFP)-tagged PKC isoforms. We have found that PKC translocation is dynamic and reversible in living cells and that PKC targeting is isoform-specific, stimulus-specific and cell-specific. In the present study, we have demonstrated that 1) arachidonic acid and ceramide act on CIB domains of epsilon and delta PKC and translocate these PKC isoforms to Golgi complex in isoform-specific manner, 2) ceramide induces translocation of deltaPKC to Golgi complex and activates this PKC isoform through tyrosine-phosphorylation., 3) Activation of deltaPKC by ceramide results in apoptosis which is distinctly different cell response from that induced by phorbol ester, another activator of PKC., 4) endogenous PKC can be knock-down by siRNA in isoform and species specifically., 5) Multiple PKC isoforms are involved in phagocytosis of microglial cells. Finally, we produced transgenic mice expressing GFP-tagged PKC isoforms using tetracyclin-regulated system. The expression of PKC-GFP can be regulated spatially and temporally. These transgenic mice enable us to monitor PKC targeting in brain slices by physiological condition and we observed PKC translocation in living brain which was different from that observed in cultured cells.


• 神経可塑性制御に対するPKCの役割-トランスジェニックマウスを用いた研究-

  酒井 規雄, 天野 託, 松林 弘明, 白井 康仁, 斎藤 尚亮

  日本学術振興会, 科学研究費助成事業, 特定領域研究, 2001 - 2002

  (1)Tet制御システムを用いたγPKC-GFP,εPKC-GFPトランスジェニックマウスの解析 neuron specific enolase promoterの下流にtTAを発現するマウス(NSE-tTA)、calcium calmodulin kinaseII promoterの支配下にtTAを発現するマウス(CamKII-tTA)と掛け合わした際のPKC-GFPの発現部位の特徴についての解析はほぼ終了した。現在、NSE-tTA x γPKC-GFPバイトランスジェニックマウスから小脳生スライスを作製し、γPKC-GFP発現小脳プルキンエ細胞において、2光子励起レーザー顕微鏡や高感度CCDカメラを用いて、γPKCのトランスロケーションの観察を試みた。各種グルタミン酸受容体作用薬、あるいは電気刺激により、γKCのトランスロケーションが観察可能で刺激に応じてPKC-GFPのトランスロケーションが伝播していくライブイメージをとらえることが可能になった。また、CamKII-tTAマウスとγPKC-GFPマウスを掛け合わせ、海馬を含む前脳全体にγPKC-GFPが発現するマウスにおいてもγPKCのトランスロケーションの観察を試みを行っている。このように、我々の作成したマウスはターゲティングを指標にPKCの神経系での機能を解析するツールとなるだけでなく、神経の興奮の伝播を蛋白質レベルで明瞭にイメージングすることが出来る新しい神経機能解析のための遺伝子操作動物であると考えられる。 (2)PKCターゲティングを脳部位特異的に減弱させたトランスジェニックマウスを作製し神経特異的機能とPKCとの関連を明らかにする。 δPKC γPKCのPDK1リン酸化部位をアラニンに変異させた変異体を作成し、培養細胞に発現させ受容体刺激に伴うトランスロケーションを観察したところ、野生型に比し、細胞膜に長くとどまることが明らかになった。このように、PDK1はPKCをリン酸化することによりターゲティングを調節しうることが明らかになった。


• CキナーゼとDGキナーゼの神経情報伝達におけるクロストーク機構の解析

  齋藤 尚亮, 白井 康仁

  日本学術振興会, 科学研究費助成事業, 特定領域研究(C), 神戸大学, 2001 - 2001

  神経可塑性における蛋白質リン酸化酵素を介した細胞内情報伝達の時・空間ネットワークを解明することを目的とし、GFP標識PKC各サブタイプを脳部位特異的に発現するTet制御システム遺伝子導不動物を用いて、神経可塑性(特にLTP、LTD)におけるこれらリン酸化酵素各サブタイプの時間的・空間的動態を電気生理学的現象と同時に解析することを試みた。同時に、PKC, DGキナーゼそれぞれ複数のサブタイプに焦点をあて、それらを波長の異なる2種類の蛍光タンパク質で標識し、発現させ、生細胞で同時に可視化し、リン酸化酵素群のクロストークを解析した。 【結果】PKCおよびDGKの神経における働き、特に神経可塑性におけるこれらリン酸化酵素のクロストークをライブイメージングし、解析を行った。その成果として、1)PKCによる基質タンパク質のリン酸化にはPKCのターゲティングが必須であること、2)DGKがPKCのターゲティングのスィッチオフに深く関与すること、3)FRETを用いた解析により、DGKとPKCが直接結合すること、を明らかにし、また、4)tTA・TetOPシステムを用いて、小脳プルキンエ細胞、海馬錐体細胞などにGFP標識各PKCサブタイプを発現誘導可能なマウスの作製に成功した。作製したGFP標識PKC発現トランスジェニックマウスの脳スライスを用いて、PKCターゲティングを観察した結果、脳スライスにおけるPKCターゲティングは、培養細胞あるいは初代培養神経細胞におけるターゲティングと異なっている事が明らかとなった。 また、PKCターゲティング機構の分子メカニズムおよびその生理的意義を明らかにするために種々の脂質シグナルによるPKCターゲティングを解析した。その結果、deltaPKCは、セラミドの刺激によって特異的にゴルジ体にターゲティングし、同時にチロシンリン酸化によって活性化されることを示した。このセラミドによるdeltaPKCの活性化機構は、ホルボールエステルにより細胞膜へのトランスロケーションを伴う活性化機構とは全く異なっており、それにより導かれる細胞応答ち異なっていた。つまり、HeLa細胞においては、セラミドは細胞のアポトーシスを引き起こすのに対して、ホルボールエステルはG2/M期での停止を起こした。このように、同一のPKCサブタイプであっても、ターゲティング部位の違いによって、明らかに異なる細胞反応を引き起こすことから、PKCターゲティング機構の重要性が明らかにされた。


• 可視化システムを用いたプロテインキナーゼCのターゲティング機構の解析

  白井 康仁

  日本学術振興会, 科学研究費助成事業, 奨励研究(A), 神戸大学, 1999 - 2000

  本研究は、「PKCの多様なタ一ゲッティングを調節する機構を分子レベルで解明すること」を目的としている。本年度は、計画1(ターゲッティングに関与するepsilon PKC分子内ドメインの決定)に焦点を絞り実験を行った。実際には、蛍光蛋白質で標識したepsilon PKCの変異体を作製し、ホルボールエステルと、各脂肪酸によるトランスロケーション様式を解析した。その結果、ステアリン酸による細胞質膜ヘのトランスロケーションには、疑似基質領域付近とClBドメインが重要であるのに対して、アラキドン酸やリノール酸によるゴルジ体へのトランスロケーションにはClBドメインが必須であることを明らかにした。また、このClBドメインをCHO-K1細胞内に発現させるとゴルジ体に集積することから、このドメイン内にゴルジ体と相互作用する部位が存在することも明らかにした。一方、TPAによる細胞質膜への不可逆的なトランスロケーションには、C1BドメインとClAドメインが関与しており、このうち主にClBドメインが重要であることが明らかになった。さらに、脂肪酸などの脂質シグナルとそれを産生するホスホリパーゼA2が、受容体を介したPKCのトランスロケーションに関与していることを明らかにし、Journal of Cell Scienceに報告した。今後、このC1Bドメインを中心として、計画2(脂質シグナルとPKCとの結合様式の解析)と計画3(ゴルジ体において、epsilon PKCと結合する分子の同定)を進めていく予定である。


• プロテインキナーゼCサブタイプの活性化とターゲッティング機構の関連に関する研究

  斎藤 尚亮, 白井 康仁

  日本学術振興会, 科学研究費助成事業, 特定領域研究(A), 神戸大学, 1998 - 1998

  プロテインキナーゼC(PKC)のtranslocation(ターゲッティング)機構を解析することにより、PKC各分子種の生理的な役割について解析した。我々はGFP(Green fluorescent protein)とラット各PKC分子種(γ-,.ε-,δ-PKC)の融合蛋白質を各種培養細胞に発現させ、細胞外刺激により活性化された生細胞内のPKCの細胞内の動態の観察を行った。その結果、 1)γ-,δ-,ε-PKCすべでが、G蛋白質共役型ATP受容体刺激により、20sec以内に細胞膜にtranslocationし、3min以内に元の状態に戻る(Retranslocation)。しかし、ホルボールエステル刺激によっては、すべての分子種が比較的ゆっりくりと(3-5min)ranslocationし、そのまま膜にとどまる。つまり、同一の分子種であっても、刺激により異なるターゲッティング機構を持つことが示された。 2)アラキドン酸刺激により、γ-PKCは素早く細胞膜に、ε-PKCはややゆっくりとGolgi体周辺にtranslocationするのに対して、δ-PKCはtranslocationしない。つまり、同一の刺激によっても分子種によってtargeting部位(translocation)は異なることが示された。 3)δ-PKCを活性化する3種の刺激(ATP,TPA,H202)は、それぞれ異なるターゲッティング機構によるものであり、細胞の機能に対する調節機構も異なると推測される。以上の結果から、PKCのターゲッティング機構は、分子種特異的、刺激特異的でありしかも時間的、空間的にも異なっていることが示され、ターゲッティング機構の解析はPKC分子種独自の機能の解明に有力な方法と考えられた。


• GFP標識プロテインキナーゼC遺伝子改変マウスの作製とその神経可塑性研究への応用

  斎藤 尚亮, 白井 康仁

  日本学術振興会, 科学研究費助成事業, 特定領域研究(A), 神戸大学, 1998 - 1998

  本研究では、培養細胞系を用いて、各PKCサブタイプ特異的なターゲッティング機構を生細胞内で明らかにするとともに、長期増強や長期抑圧現象におけるPKCの細胞内動態を解析することにより、PKCのシナプス可塑性における働きを明らかにすることを目的とした。そのために、1)培養細胞系におけるGFP標識PKC各分子種のトランスロケーション現象と活性化機構の関連についての解析、2)GFP標識PKCトランスジェニックマウスの作製の試みを行った。その結果、1)γ-,δ-,ε-PKCすべてが、G蛋白質共役型ATP受容体刺激により、20sec以内に細胞膜にtranslocationし、3min以内に元の状態に戻る(Retranslocation)。しかし、ホルボールエステル刺激によっては、すべての分子種が比較的ゆっくりと(3-5min)ranslocationし、そのまま膜にとどまる。つまり、同一の分子種であっても、刺激により異なるターゲッティング機構を持つことが示された。2)アラキドン酸刺激により、γ-PKCは素早く細胞膜に、ε-PKCはややゆっくりとGolgi体周辺にtranslocationするのに対して、δ-PKCはtranslocationしない。つまり、同一の刺激によっても分子種によってtargeting部位(translocation)は異なることが示された。3)δ-PKCを活性化する3種の刺激(ATP,TPA,H202)は、それぞれ異なるターゲッティング機構によるものであり、細胞の機能に対する調節機構も異なると推測される。 以上の結果から、PKCのターゲッティング機構は、分子種特異的、刺激特異的でありしかも時間的、空間的にも異なっていることが示され、ターゲッティング機構の解析はPKC分子種独自の機能の解明に有力な方法と考えられた。


• Research for modulation of expression of serotonin transporter and its involvement in mental disorder

  SAITO Naoaki, SAKAI Norio, SHIRAI Yasuhito

  Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research, Grant-in-Aid for Scientific Research (C), KOBE UNIVERSITY, 1997 - 1998

  We examined the regulatory mechanism of serotonin transporter and the involvement of serotonin transporter (SET) in depression to elucidate the function role of SET in mental disorder. 1. Regulation of SET by phosphorylation. The regulation of SET expressed in COS7 cells by the activation of protein kinase C with phorbol ester or by inhibition of protein phosphatases with calyculin A was examined. Both treatment decreased the activity of SET with a reduction in Vmax without affecting Km. Site-directed mutagenesis of five putative PKC phosphorylation sites in SET revealed that these inhibitory modulation of SET activity did not act via direct phosphorylation of SET by PKC. 2. Immunocytochemical localization of SET.The cellular and intracellular localization of SET was examined by immunocytochemistry. SET was localized in the varicose nerve fibers and nerve terminals but not in the cell bodies. Under electron microscopy, SET was seen around the synaptic vesicles as well as presynaptic membrane. 3. The effects of interferon-alpha and -gamma on the transcription of SET.Both SET mRNA and SET activity were increased by treatment with interferon-alpha and -gamma for 3 h. Treatment with dibutyryl cAMP also increased SET mRNA and SET activity. The level of SET mRNA was increased both in the midbrain and adrenal glands of mice which were treated with interferons for 3 h. These results suggest that the interferon-induced psychiatric side effects arise through regulation of serotonin transporter transcription and that the transcriptional regulation of the serotonin transporter is a possible neurochemical mechanism of affective disorders.


• 蛍光標識プロテインキナーゼCの遺伝子改変マウスの作製とその神経可塑性研究への応用

  斎藤 尚亮, 白井 康仁, 酒井 規雄

  日本学術振興会, 科学研究費助成事業, 重点領域研究, 神戸大学, 1997 - 1997

  本研究は、長期増強、長期抑圧現象におけるプロテインキナーゼC(PKC)分子種の役割を明らかにすることを目的とし、PKC各分子種のトランスロケーション現象の解析を行い、活性化型PKCとリン酸化基質の空間的関係を形態学的に解析することを目的とした。本年度は神経系に多く存在するPKC分子種のうち、シナプス後部に存在するγ-分子種に焦点を当て研究を行った。γ-PKCにGFP融合した蛋白質を細胞に発現させ、コンフォーカル顕微鏡下でGFP蛍光を指標として種々の細胞外刺激によるPKCの細胞内動態を経時的に記録した。刺激後のGFP蛍光の局在は、免疫蛍光染色によるそれぞれのPKC分子種の局在と一致しており、また、Immunoblotによってもγ-PKC-GFPの分解産物が認められなかったことから、経時的に観察したGFP蛍光はPKC自体の動態をあらわすものと考えられた。また、酵素学的な解析によりγ-PKC-GFPもγ-PKCと同様の酵素学的特性を持つことが明らかとなり、GFPの融合によってもγ-PKCの特性には影響が少ないと考えられた。このGFP融合PKC分子種を用いて1)TPAによるtranslocationは比較的遅い時間経過で起こり、irreversibleであった、2)CaイオンによるtranslocationおよびG蛋白質結合型受容体の刺激によるtranslocationも20sec以内に細胞膜にtranslocationし、3min以内に元の状態に戻る速くreversibleなものであった。また、site-directed mutagenesisを用いた実験により、受容体の刺激によるtranslocationには、Caイオン、C2 domainは必須であるが、TPAによるtranslocationには、Caion,C2 domainは必須ではないことが明らかになった。


• Na^+/Cl^-依存性トランスポーターの分子間相互作用と機能協関する蛋白質の検索

  斎藤 尚亮, 白井 康仁, 酒井 規雄

  日本学術振興会, 科学研究費助成事業, 重点領域研究, 神戸大学, 1997 - 1997

  本研究では、Na^+/Cl^-依存性トランスポーターの一種であり、感情障害の原因とも考えられているセロトニントランスポーター(SET)に注目し、SETと結合し機能協関する蛋白質の検索と機能解析を目的とした。SETの生理機能を解析するために、まず、リン酸化、脱リン酸化によってSETの機能がどのように修飾されるかを検討した。SET活性は、PKCの活性化により有意に抑制され、その抑制はリン酸化酵素阻害薬によって完全に阻害された。また、不活性型のホルボールエステルはこのような抑制効果を示さなかった。また、フォスファターゼ1および2Aの阻害剤であるカリクリンAも同様にSET活性を抑制し、そのカリクリンによる抑制もリン酸化酵素阻害薬によって阻害された。これらの事実により、SET活性はリン酸化、脱リン酸化のバランスによって制御されていることが示された。Eadie-Hofstee解析により、このTPAやCLAによる制御はSETのセロトニンに対する親和性には影響を与えず、最大取り込み速度の抑制によることが示された。このプロテインキナーゼCによるリン酸化が、直接のリン酸化によるものであるかどうかを検討するために、site-directed mutagenesisを用いて、5カ所のリン酸化されうると推定されるそれぞれのセリンあるいはトレオニン残基を変異させた変異体を作製し、リン酸化、脱リン酸化の影響を検討した。その結果、すべてのリン酸化されうる部位を変異させても、TPA、CLAの効果が認められ、少なくともこの5カ所は直接リン酸化されないことが明らかになった。これらの結果から、SETはリン酸化・脱リン酸化酵素によって、間接的に制御されておいることが示唆された。今後、その間接的な制御機構を分子レベレで解明するためにSET結合蛋白の単離を行う予定である。


• カイコ前胸腺刺激ホルモン分泌におけるリン酸化タンパク質の解析

  白井 康仁

  日本学術振興会, 科学研究費助成事業, 奨励研究(A), 神戸大学, 1995 - 1995

  アセチルコリンが誘起する前胸腺刺激ホルモン分泌(PTTH)において、特異的にリン酸化される3種(32k,25k21k)の蛋白質の機能を解明する研究の一環として、カイコ脳-側心体-アラタ体複合器官(Br-CC-CA)における低分子量G蛋白質を検索し、その分子特性並びに細胞内局在性をリン酸化蛋白質のそれと比較した。 その結果、Br-CC-CA中に3種(28k,25k,21k)の低分子量G蛋白質を見いだした。このうち、カイコ25kG蛋白質は、哺乳類の低分子量G蛋白質であるRab3Aに対する抗体に交叉性を示したが、他のカイコ低分子量G蛋白質は、哺乳類の低分子量G蛋白質に対するどの抗体とも交叉性を示さなかった。ついで、Rab3A抗体を用いてカイコ25kG蛋白質の組織局在性を調べたところ、脳及び食道下神経節に多量に存在している事が明らかになった。この知見は、Rab3Aが哺乳類において神経特異的に発現している事実と類似していた。一方、PTTH分泌においてリン酸化される蛋白質とカイコ低分子量G蛋白質の細胞内局在性を調べたところ、共に側心体一アラタ体より調製した粗シナプトソーム画分に存在した。 以上、分子量並びにその細胞内局在性が一致することから、PTTH分泌においてリン酸化される25kリン酸化蛋白質はRab3Aと相同性を有する低分子量G蛋白質であり、21kリン酸化蛋白質は新規な低分子量G蛋白質であることが示唆された。現在、2次元電気泳動を駆使して、両リン酸化蛋白質が真に低分子量G蛋白質であるか否かを詳細に検討している。また、32kリン酸化蛋白質に機能については、今後の研究課題として残された。