Analysis of protein kinases relating to glucotoxicity

• Project/Area Number: 21770144
• Research Category: Grant-in-Aid for Young Scientists (B)
• Allocation Type: Single-year Grants
• Research Field: Functional biochemistry
• Research Institution: Kagawa University
• Principal Investigator: SUEYOSHI Noriyuki Kagawa University, 農学部, 准教授 (90346635)
• Project Period (FY): 2009 – 2010
• Project Status: Completed (Fiscal Year 2010)
• Budget Amount: ¥4,680,000 (Direct Cost: ¥3,600,000、Indirect Cost: ¥1,080,000); Fiscal Year 2010: ¥1,170,000 (Direct Cost: ¥900,000、Indirect Cost: ¥270,000); Fiscal Year 2009: ¥3,510,000 (Direct Cost: ¥2,700,000、Indirect Cost: ¥810,000)
• Keywords: 糖尿病 / 糖毒性 / プロテインキナーゼ / CaMKIV / マルチPK抗体 / 2型糖尿病 / Calpain

Research Abstract

Glucotoxicity is a critical component of the pathophysiology of type 2 diabetes mellitus ; however, the molecular mechanisms of glucotoxicity are still not fully understood. In this study, we analyzed the protein kinase that correlated with insulin synthesis in INS-1 cells under glucotoxic conditions. When expression patterns of protein kinases in INS-1 cells were analyzed by Western blotting with Multi-PK antibodies, a kinase of 63 kd was significantly reduced concomitant with the decrease of insulin secretion under glucotoxic conditions. Judging from the molecular size of a native kinase/cyanogen bromide fragment and pI value, the 63-kd protein kinase was deduced to be CaMKIV. The decreased CaMKIV levels under glucotoxic conditions recovered to original levels after changing the medium to a normal glucose concentration. Recombinant CaMKIV was degraded in a Ca^<2+>-dependent manner by incubation with cell lysates from INS-1 cells under glucotoxic conditions, and degradation was protected by calpain inhibitor. Furthermore, CaMKIV was reduced in the pancreatic islets of diabetic Otsuka Long-Evans Tokushima fatty rats, whereas that of nondiabetic Long-Evans Tokushima Otsuka rats was not. This study suggests that the abnormal regulation of CaMKIV is a component of β-cell dysfunction caused by high glucose.

Research Products

• [Journal Article] CoPK32 is a novel stress-responsive protein kinase in the mushroom Coprinopsis cinerea. (2011)
  • Author(s): Kaneko K., Sugiyama Y., Yamada Y., Sueyoshi N., Watanabe A., Asada Y., Ishida A., Kameshita I.
  • Journal Title: Biochimica et Biophysica Acta in press
  • Peer Reviewed
• [Journal Article] Influence of Ser/Pro-rich domain and kinase domain of doublecortin-like protein kinase on microtubule-binding activity. (2011)
  • Author(s): Nagamine T., Shimomura S., Sueyoshi N., Kameshita I.
  • Journal Title: The Journal of Biochemistry in press
  • Peer Reviewed
• [Journal Article] プロテインキナーゼ解析用ツールとしての網羅的抗体 (2011)
  • Author(s): 亀下勇、末吉紀行
  • Journal Title: 生化学 83巻 Pages: 330-335
  • NAID: 10029068281
  • Peer Reviewed
• [Journal Article] Detection of protein kinase substrates in tissue extracts after separation by isoelectric focusing. (2011)
  • Author(s): Senga Y., Nagamine T., Sekiguchi M., Kaneko K., Sueyoshi N., Kameshita I.
  • Journal Title: Analytical Biochemistry 408(2) Pages: 345-347
  • Peer Reviewed
• [Journal Article] Calcium/calmodulin-dependent protein kinase IV involvement in the pathophysiology of glucotoxicity in rat pancreatic β-cells. (2011)
  • Author(s): Sugiyama Y., Murao K., Imachi H., Sueyoshi N., Ishida T., Kameshit I.
  • Journal Title: Metabolism 60(1) Pages: 145-153
  • Peer Reviewed
• [Journal Article] Calcium/calmodulin-dependent protein kinase IV involvement in the pathophysiology of glucotoxicity in rat pancreatic β-cells. (2011)
  • Author(s): Yasunori Sugiyama, et al.
  • Journal Title: Metabolism Volume: 60(1) Pages: 145-153
  • Peer Reviewed
• [Journal Article] Detection of protein kinase substrates in tissue extracts after separation by isoelectric focusing. (2011)
  • Author(s): Yukako Senga, et al.
  • Journal Title: Analytical Biochemistry Volume: 408(2) Pages: 345-347
  • Peer Reviewed
• [Journal Article] Influence of Ser/Pro-rich domain and kinase domain of doublecortin-like protein kinase on microtubule-binding activity. (2011)
  • Author(s): Tadashi Nagamine, et al.
  • Journal Title: The Journal of Biochemistry Volume: (掲載確定)
  • Peer Reviewed
• [Journal Article] CoPK32 is a novel stress-responsive protein kinase in the mushroom Coprinopsis cinerea. (2011)
  • Author(s): Keisuke Kaneko, et al.
  • Journal Title: Biochimica et Biophysica Acta Volume: (掲載確定)
  • Peer Reviewed
• [Journal Article] The DNA-binding activity of mouse DNA methyltransferase 1 is regulated by phosphorylation with casein kinase 1delta/epsilon. (2010)
  • Author(s): Sugiyama Y., Hatano N., Sueyoshi N., Suui N., Su N., SuN., Suuuuu N., Suetake I., Tajima S., Kinoshita E., Kinoshita-Kikuta E., Koike T., Kameshita I.
  • Journal Title: Biochemical Journal 427(3) Pages: 489-497
  • Peer Reviewed
• [Journal Article] A minimum size homologue of Ca^2+/calmodulin-dependent protein kinase IV naturally occurring in zebrafish (2010)
  • Author(s): Nimura T., Sugiyama Y., Sueyoshi N., Shigeri Y., Ishida A., Kameshita I.
  • Journal Title: The Journal of Biochemistry 147(6) Pages: 857-865
  • NAID: 10027876530
  • Peer Reviewed
• [Journal Article] Expression and characterization of PKL01, an Ndr kinase homolog in Lotus japonicus. (2010)
  • Author(s): Kameshita I., Shimomura S., Nishio K., Sueyoshi N., Nishida T., Nomura M., Tajima S.
  • Journal Title: The Journal of Biochemistry 147(6) Pages: 799-807
  • NAID: 10027876295
  • Peer Reviewed
• [Journal Article] Identification of an endogenous substrate of zebrafish doublecortin-like protein kinase using a highly active truncation mutant. (2010)
  • Author(s): Shimomura S., Nagamine T., Hatano N., Sueyoshi N., Kameshita I.
  • Journal Title: The Journal of Biochemistry 147(5) Pages: 711-722
  • NAID: 10027875877
  • Peer Reviewed
• [Journal Article] In-gel protein phosphatase assay using fluorogenic substrates (2010)
  • Author(s): Kameshita I., Baba H., Umeda Y., Sueyoshi N.
  • Journal Title: Analytical Biochemistry 400(1) Pages: 118-122
  • NAID: 120007015426
  • Peer Reviewed
• [Journal Article] Identification of an endogenous substrate of zebrafish doublecortin-like protein kinase using a highly active truncation mutant. (2010)
  • Author(s): Sachiko Shimomura, et al.
  • Journal Title: The Journal of Biochemistry Volume: 147(5) Pages: 711-722
  • Peer Reviewed
• [Journal Article] Expression and characterization of PKL01, an Ndr kinase homolog in Lotusjaponicus. (2010)
  • Author(s): Isamu Kameshita, et al.
  • Journal Title: The Journal of Biochemistry Volume: 147(6) Pages: 799-807
  • Peer Reviewed
• [Journal Article] A minimum size homologue of Ca^<2+>/calmodulin-dependent protein kinase IV naturally occurring in zebrafish. (2010)
  • Author(s): Takaki Nimura, et al.
  • Journal Title: The Journal of Biochemistry Volume: 147(6) Pages: 857-865
  • Peer Reviewed
• [Journal Article] The DNA-binding activity of mouse DNA methyltransferase 1 is regulated by phosphorylation with casein kinase 1delta/epsilon. (2010)
  • Author(s): Yasunori Sugiyama, et al.
  • Journal Title: Biochemical Journal Volume: 427(3) Pages: 489-497
  • Peer Reviewed
• [Journal Article] Calcium/calmodulin-dependent protein kinase IV involvement in the pathophysiology of glucotoxicity in rat pancreatic beta cells (2010)
  • Author(s): Sugiyama, Y.
  • Journal Title: Metabolism (掲載確定)
  • Peer Reviewed
• [Journal Article] In-gel protein phosphatase assay using fluorogenic substrates (2010)
  • Author(s): Kameshita, I.
  • Journal Title: Anal.Biochem. 400 Pages: 118-122
  • Peer Reviewed
• [Journal Article] Ca^2+/calmodulin-dependent protein kinase phosphatase (CaMKP) is indispensable for normal embryogenesis in zebrafish, Danio rerio. (2009)
  • Author(s): Sueyoshi, N., Nimura, T., Ishida, A., Taniguchi, T., Yoshimura, Y., Ito, M., Shigeri, Y., Kameshita, I.
  • Journal Title: Arch.Biochem.Biophys. 488 Pages: 48-59
  • Peer Reviewed
• [Journal Article] Novel Ca^2+/calmodulin-dependent protein kinase expressed in actively growing mycelia of the basidiomycetous mushroom Coprinus cinereus. (2009)
  • Author(s): Kaneko, K., Yamada, Y., Sueyoshi, N., Watanabe, A., Asada, Y., Kameshita, I.
  • Journal Title: Biochim.Biophys.Acta 1790 Pages: 71-79
  • Peer Reviewed
• [Journal Article] Mechanistic insights into the hydrolysis and synthesis of ceramide by neutral ceramidase. (2009)
  • Author(s): Inoue, T., Okino, N., Kakuta, Y., Hijikata, A., Okano, H., Goda, H.M., Tani, M., Sueyoshi, N., Kambayashi, K., Matsumura, H., Kai, Y., Ito, M.
  • Journal Title: J.Biol.Chem. 284 Pages: 9566-9577
  • Peer Reviewed
• [Journal Article] Calcium/calmodulin-dependent protein kinase phosphatase (CaMKP) is indispensable for normal embryogenesis in zebrafish, Danio rerio (2009)
  • Author(s): Sueyoshi, N.
  • Journal Title: Arch.Biochem.Biophys 488 Pages: 48-59
  • Peer Reviewed
• [Presentation] Functional Processing of Nuclear Ca^2+/Calmodulin-dependent Protein Kinase Phosphatase (CaMKP-N): EVIDENCE FOR A CRITICAL ROLE OF PROTEOLYTIC PROCESSING IN THE REGULATION OF ITS CATALYTIC ACTIVITY, SUBCELLULAR LOCALIZATION AND SUBSTRATE TARGETING IN VIVO. (2011)
  • Author(s): Noriyuki Sueyoshi, Takaki Nimura, Takashi Onouchi, Hiromi Baba, Shinobu Takenaka, Atsuhiko Ishida, Isamu Kameshita
  • Organizer: 1^st International Symposium on Carcinogenic Spiral & 9th International Conference on Protein Phosphatase
  • Place of Presentation: Tokyo
  • Year and Date: 2011-02-03
• [Presentation] Functional Processing of Nuclear Ca^<2+>/Calmodulin-dependent Protein Kinase Phosphatase (CaMKP-N) : EVIDENCE FOR A CRITICAL ROLE OF PROTEOLYTIC PROCESSING IN THE REGULATION OF ITS CATALYTIC ACTIVITY, SUBCELLULAR LOCALIZATION AND SUBSTRATE TARGETING IN VIVO. (2011)
  • Author(s): Noriyuki Sueyoshi, et al.
  • Organizer: 1^<st> International Symposium on Carcinogenic Spiral & 9^<th> International Conference on Protein Phosphatase
  • Place of Presentation: The University of Tokyo (Tokyo, Japan)
  • Year and Date: 2011-02-03
• [Presentation] Ca^2+/CaM依存性プロテインキナーゼI (CaMKI)-delta はゼブラフィッシュの正常な胚発生に重要である (2010)
  • Author(s): 千賀由佳子、永峰賢、二村貴樹、末吉紀行、亀下勇
  • Organizer: 第33回日本分子生物学会年会 第83回日本生化学会大会 合同大会
  • Place of Presentation: 神戸
• [Presentation] 植物Ndrキナーゼの自己チロシンリン酸化活性 (2010)
  • Author(s): 片山将一、杉山康憲、末吉紀行、寺地徹、亀下勇
  • Organizer: 第33回日本分子生物学会年会 第83回日本生化学会大会合同大会
  • Place of Presentation: 神戸
• [Presentation] 蛍光基質を用いたホスファターゼ検出法の新しい応用例 (2010)
  • Author(s): 馬場裕美、金子啓祐、末吉紀行、亀下勇
  • Organizer: 第33回日本分子生物学会年会 第83回日本生化学会大会合同大会
  • Place of Presentation: 神戸
• [Presentation] 核局在型Ca^2+/カルモジュリン依存性プロテインキナーゼホスファターゼ(CaMKP-N)の機能的な限定分解 (2010)
  • Author(s): 小野内貴士、二村貴樹、馬場裕美、竹中忍、石田敦彦、亀下勇、末吉紀行
  • Organizer: 第33回日本分子生物学会年会 第83回日本生化学会大会 合同大会
  • Place of Presentation: 神戸
• [Presentation] Ca^2+/calmodulin-dependentprotein kinase I (CaMKI)-delta isindispensable for normalembryogenesis in zebrafish (2010)
  • Author(s): Yukako Senga, Tadashi Nagamine, TakakiNimura, Noriyuki Sueyoshi, Isamu Kameshita
  • Organizer: Daniorerio.
  • Place of Presentation: ISD-Nara, Japan, Nara
• [Presentation] Identification and characterizationof protein kinases that bind toN-terminal domain of DNAmethyltransferase 1. (2010)
  • Author(s): Mari Sekiguchi, Yasunori Sugiyama,Noriyuki Sueyoshi, Isamu Kameshita
  • Organizer: 12th IUBMBConference
  • Place of Presentation: Melbourne
• [Presentation] Knockdown ofzebrafish Ca2+/calmodulin-dependentprotein kinase I・・・・・・ causesdevelopmental abnormalities inzebrafish, Danio rerio. (2010)
  • Author(s): Yukako Senga, Tadashi Nagamine, TakakiNimura, Eriko Nanto, Isamu Kameshita,Noriyuki Sueyoshi
  • Organizer: 12th IUBMBConference
  • Place of Presentation: Melbourne
• [Presentation] 担子菌Coprinopsiscinereaの子実体形成過程で活性が増大するホスファターゼ (2010)
  • Author(s): 金子啓祐、馬場裕美、渡邊彰、麻田恭彦、末吉紀行、亀下勇
  • Organizer: 日本農芸化学会四国支部大会
  • Place of Presentation: 香川
• [Presentation] Characterization ofnovel protein kinases in mushroomCoprinus cinereus (2010)
  • Author(s): Keisuke Kaneko, Noriyuki Sueyoshi,Isamu Kameshita
  • Organizer: The 9thInternational Mycological Congress
  • Place of Presentation: Edinburgh
• [Presentation] Detectionof protein kinases and proteinphosphatases expressed in mushroom (2010)
  • Author(s): Hiromi Baba, Keisuke Kaneko, NoriyukiSueyoshi, Isamu Kameshita
  • Organizer: The 9th International MycologicalCongress
  • Place of Presentation: Edinburgh
• [Presentation] ゼブラフィッシュの初期胚発生におけるCa^2+/CaM依存性プロテインキナーゼI delta (CaMKIdelta)の役割 (2010)
  • Author(s): 千賀由佳子、永峰賢、二村貴樹、末吉紀行、亀下勇
  • Organizer: 第51回日本生化学会中国・四国支部例会
  • Place of Presentation: 山口
• [Presentation] プロテインホスファターゼの新しい検出法の開発とその応用 (2010)
  • Author(s): 馬場裕美、梅田嘉徳、末吉紀行、亀下勇
  • Organizer: 第51回日本生化学会中国・四国支部例会
  • Place of Presentation: 山口
• [Presentation] Characterization of novel protein kinases in mushroom Coprinus cinereus. (2010)
  • Author(s): Keisuke Kaneko, et al.
  • Organizer: The 9th International Mycological Congress
  • Place of Presentation: Edinburgh International Conference Centre (Edinburgh, UK)
• [Presentation] Detection of protein kinases and protein phosphatases expressed in mushroom. (2010)
  • Author(s): Hiromi Baba, et al.
  • Organizer: The 9th International Mycological Congress
  • Place of Presentation: Edinburgh International Conference Centre (Edinburgh, UK)
• [Presentation] Identification and characterization of protein kinases that bind to N-terminal domain of DNA methyltransferase 1. (2010)
  • Author(s): Mari Sekiguchi, et al.
  • Organizer: 12th IUBMB Conference
  • Place of Presentation: Melbourne Exhibition & Convention Centre (Melbourne, Australia)
• [Presentation] Knockdown of zebrafish Ca^<2+>/calmodulin-dependent protein kinase I□□□□□ causes developmental abnormalities in zebrafish, Danio rerio (2010)
  • Author(s): Yukako Senga, et al.
  • Organizer: 12th IUBMB Conference
  • Place of Presentation: Melbourne Exhibition & Convention Centre (Melbourne, Australia)
• [Presentation] Ca^<2+>/calmodulin-dependent protein kinase I (CaMKI)-delta is indispensable for normal embryogenesis in zebrafish, Danio rerio. (2010)
  • Author(s): Yukako Senga, et al.
  • Organizer: ISD-Nara
  • Place of Presentation: Nara-ken New Public Hall (Nara, Japan)
• [Presentation] 担子菌Coprinopsis cinereaの子実体形成過程で活性が増大するホスファターゼ (2010)
  • Author(s): 金子啓祐, 他
  • Organizer: 2010年度日本農芸化学会四国支部大会
  • Place of Presentation: 香川大学(香川)
• [Presentation] ゼブラフィッシュの初期胚発生におけるCa^<2+>/CaM依存性プロテインキナーゼI delta(CaMKIdelta)の役割 (2010)
  • Author(s): 千賀由佳子, 他
  • Organizer: 第51回日本生化学会中国・四国支部例会
  • Place of Presentation: 山口大学(山口)
• [Presentation] プロテインホスファターゼの新しい検出法の開発とその応用 (2010)
  • Author(s): 馬場裕美, 他
  • Organizer: 第51回日本生化学会中国・四国支部例会
  • Place of Presentation: 山口大学(山口)
• [Presentation] Ca^<2+>/CaM依存性プロテインキナーゼI(CaMKI)-deltaはゼブラフィッシュの正常な胚発生に重要である (2010)
  • Author(s): 千賀由佳子, 他
  • Organizer: 第33回日本分子生物学会年会 第83回日本生化学会大会 合同大会
  • Place of Presentation: 神戸国際会議場(神戸)
• [Presentation] 植物Ndrキナーゼの自己チロシンリン酸化活性 (2010)
  • Author(s): 片山将一, 他
  • Organizer: 第33回日本分子生物学会年会 第83回日本生化学会大会 合同大会
  • Place of Presentation: 神戸国際会議場(神戸)
• [Presentation] 蛍光基質を用いたホスファターゼ検出法の新しい応用例 (2010)
  • Author(s): 馬場裕美, 他
  • Organizer: 第33回日本分子生物学会年会 第83回日本生化学会大会 合同大会
  • Place of Presentation: 神戸国際会議場(神戸)
• [Presentation] 核局在型Ca^<2+>/カルモジュリン依存性プロテインキナーゼホスファターゼ(CaMKP-N)の機能的な限定分解 (2010)
  • Author(s): 小野内貴士, 他
  • Organizer: 第33回日本分子生物学会年会 第83回日本生化学会大会 合同大会
  • Place of Presentation: 神戸国際会議場(神戸)
• [Presentation] 蛍光基質を用いたゲル内プロテインホスファターゼ検出法の開発 (2009)
  • Author(s): 馬場裕美、梅田嘉徳、末吉紀行、亀下勇
  • Organizer: 第32回日本分子生物学会年会
  • Place of Presentation: 横浜
• [Presentation] ゼブラフィッシュDoublecortin-like protein kinase(zDCLK)の活性型変異体を用いた内在性基質の探索ならびに同定 (2009)
  • Author(s): 下村幸子、永峰賢、波多野直哉、末吉紀行、亀下勇
  • Organizer: 第32回日本分子生物学会年会
  • Place of Presentation: 横浜
• [Presentation] CaMKIBはゼブラフィッシュの正常な胚発生に不可欠である. (2009)
  • Author(s): 千賀由佳子、永峰賢、二村貴樹、末吉紀行、亀下勇
  • Organizer: 第32回日本分子生物学会年会
  • Place of Presentation: 横浜
• [Presentation] 核局在型CaMキナーゼホスファターゼ(CaMKP-N)の細胞内分解機構の解析 (2009)
  • Author(s): 二村貴樹、岸野良美、末吉紀行、亀下勇
  • Organizer: 第32回日本分子生物学会年会
  • Place of Presentation: 横浜
• [Presentation] 担子菌キノコのストレス応答性プロテインキナーゼの解析 (2009)
  • Author(s): 金子啓祐、杉山康憲、末吉紀行、亀下勇
  • Organizer: 日本農芸化学会関西・中四国・西日本支部、日本栄養・食糧学会九州・沖縄支部、日本食品科学工学会西日本支部、2009年度合同沖縄大会
  • Place of Presentation: 沖縄
• [Presentation] Doublecortin-line protein kinase とDoublecortinの微小管結合力の違いはSP領域に起因する (2009)
  • Author(s): 永峰賢、下村幸子、末吉紀行、亀下勇
  • Organizer: 第82回日本生化学会大会
  • Place of Presentation: 神戸
• [Presentation] マウスDNAメチルトランスフェラーゼ1のDNA結合活性はカゼインキナーゼ1によるリン酸化によって制御される (2009)
  • Author(s): 杉山康憲、関口茉里、末吉紀行、波多野直哉、木下英司、木下恵美子、小池透、末武勲、田嶋正二、亀下勇
  • Organizer: 第82回日本生化学会大会
  • Place of Presentation: 神戸
• [Presentation] スフィンゴ脂質セラミドN-デアシラーゼ(SCDase)のX線結晶構造解析 (2009)
  • Author(s): 沖野望、角田佳充、鵜木(加藤)陽子、末吉紀行、伊東信
  • Organizer: 第82回日本生化学会大会
  • Place of Presentation: 神戸
• [Remarks] ホームページ等
  • URL: http://www.ag.kagawa-u.ac.jp/plantpathology/index.html