カルシウム感受性ミオシンをバイオチップとして利用する試み

• Project/Area Number: 19659059
• Research Category: Grant-in-Aid for Exploratory Research
• Allocation Type: Single-year Grants
• Research Field: General pharmacology
• Research Institution: Gunma University
• Principal Investigator: 小浜 一弘 Gunma University, 大学院・医学系研究科, 教授 (30101116)
• Co-Investigator(Kenkyū-buntansha): 石川 良樹 群馬大学, 大学院・医学系研究科, 師講 (20218663) ; 中村 彰男 群馬大学, 大学院・医学系研究科, 師講 (30282388) ; 吉山 伸司 群馬大学, 大学院・医学系研究科, 助教 (30396650) ; 田中 秀幸 群馬大学, 医学部, 助手 (70343085) ; 石川 良樹 群馬大学, 大学院・医学研究科, 講師 (20212863)
• Project Period (FY): 2007 – 2008
• Project Status: Completed (Fiscal Year 2008)
• Budget Amount: ¥3,200,000 (Direct Cost: ¥3,200,000); Fiscal Year 2008: ¥1,600,000 (Direct Cost: ¥1,600,000); Fiscal Year 2007: ¥1,600,000 (Direct Cost: ¥1,600,000)
• Keywords: アクチン / ミオシン / バイオチップ / カルシウム

Research Abstract

研究の目的ミオシンは筋肉を構成する主要蛋白質でモーター蛋白質の代表とされ、アクチンと呼ばれる1もうひとつの筋蛋白質を動かす性質がある。この性質によって、動物細胞では筋肉収縮や細胞運動、植物細胞では原形質流動が行われている。本研究ではこの「動き」を利用してバイオチップを作成する技術を開発することを目的とするものであるが、制御の効力、ないモーター機能だけでは利用が不可能で制御のかかることが要求される。本申請はこの様な背景の元に計画されたものである。1)運搬性の形成と荷物の積載:前年度にひきつづきファッシンによりアクチン繊維を束化させ、剛性をもたした運搬体につき、移動距離とその大きさの概要を求めた。前者は5μ皿程度で今後はそれを延、長する工夫が要求される。後者は0.5μ皿程度でサイズのばらつきが多く、大きさを揃える必要がある。積載については、昨年にポリスチレンビーズを、ビオチンとアビジンのアフィニティーを介して運搬体に結合させることができた。指定した箇所へ積載は不成功のままである。ポリスチレンの他に大腸菌も同じ方式で積載することができた。2)運搬体の動きの制御:ブセビスタチンはミオシン特異的阻害薬であるが、これより速度を調節することを試みた。具体的には、ブレビスタチン非存在下でアクチン束を動かじた。ブシビスタチン存在下で速度の低下をみた。青色光照射したブレビスタチンを与えて、速度回復を観察できた。3)運搬体の方向転換:イオン集中ビームでガラス板を加工し、土手をつくりこの土手にぶつけることにした。アクチン束の剛性により方向転換を観察することができた。

Research Products

• [Journal Article] Blebbistatin Inhibits the Chemotaxis of Vascular Smooth Muscle through Breaking the Interaction between Myosi hand Actin. (2008)
  • Author(s): Wang H., Tanaka H., Qin X., Zhao T., Ye L., Okagaki T., Katayama T. Nakamura A., Isshikawa R., Thatcher S., Wright G., and Kohama k.
  • Journal Title: J. Physiol. Heart Circ. Physiol 294
  • Peer Reviewed
• [Journal Article] Plectin 1 links intermediate filaments to costameric sarcolemma through β-synemin,α-dystrobrevin and actin. (2008)
  • Author(s): Hijikata T., Nakamura A., Isokawa Imamura M., Yuasa K., Ishikawa R., Kohama K., Takeda S., Yorifuji H.
  • Journal Title: J, Cell Sci. 121 Pages: 20062-2074
  • Peer Reviewed
• [Journal Article] Assembly and Function of Myosin II on Ultraviolet/Ozone Patterned Trimethylcholorosilane Substrates. (2008)
  • Author(s): Takatsuki H., Kolli M., Rice K. M., Day B. S., Asano S., Rahman M., Zhamg Y., Ishikawa R., Kohama K., and Blough E. R.
  • Journal Title: J.Bionnanoscience 2 Pages: 1-7
  • Peer Reviewed
• [Journal Article] Role of non-kinase activity of myosin light-chain kinase in regulating smooth muscle contraction, a review dedicated to Dr. Setsuro Ebashi. (2008)
  • Author(s): Nakamura A., Xie C., Zhang Y, Gao Y, Wang H-H., YeL-H., Kishi H., Okagaki T., Yoshiyama S., Hayakawa K., Ishikawa R., Kohama K.
  • Journal Title: Bioehem. Biophys. Res. Commun. 369 Pages: 135-143
  • Peer Reviewed
• [Journal Article] Drebrin attenuates the interaction between actin and myosin-V. (2007)
  • Author(s): Ishikawa R., Katoh K. Takahashi A., Xie C., Oseki K., Watanabe M., Igarashi M., Nakamura A., Kohama K.
  • Journal Title: Biochem. Biophys. Res. Commun 359 Pages: 398-401
  • Peer Reviewed
• [Journal Article] Actin-binding Proteins in Nerve Cell Growth Cones (2007)
  • Author(s): Ishikawa R., Kohama K.
  • Journal Title: J. Pharmacol. Sci 105 Pages: 6-11
  • NAID: 10024315489
  • Peer Reviewed
• [Presentation] The non-kinase activity of myosin light chain kinase in regulating smooth muscle contraction. (2008)
  • Author(s): Nakamura A., Xie C., Matsumoto A., Zhang Y., Gao Y., Kohama K.
  • Organizer: 10th Meeting of the European Calcium Society
  • Place of Presentation: Leuven, Belgium
• [Presentation] 質量分析装置を用いたミオシン軽鎖キナーゼのリン酸化部位の決定 (2008)
  • Author(s): 中村彰男、岸博子、松本篤、石川良樹、吉山伸司、早川晃一、小林誠、小濱一弘
  • Organizer: 第118回薬理学会関東部会
  • Place of Presentation: 東京(星薬科大)
• [Presentation] 平滑筋ミオシン軽鎖キナーゼの非キナーゼ活性 (2008)
  • Author(s): 中村彰男、松本篤、謝策、張月、吉山伸司、小浜一弘
  • Organizer: 日本動物学会第79回大会
  • Place of Presentation: 福岡(福岡大学)
• [Presentation] Sliding of F-actin on myosin II under the conditions fully saturated with variety of actin-binding proteins. (2008)
  • Author(s): Ishikawa R., Takahashi A., Nakamura A., Kohama K.
  • Organizer: The 82nd Annual Meeting of the Japanese Pharn acological Society
  • Place of Presentation: 横浜
• [Presentation] Cell motility of vascular smooth muscle cells exposed to the cigarette smoke components. (2008)
  • Author(s): Yoshiyama S., Wang H. -H., Nakamura A., Kohama K.
  • Organizer: The 82nd Annual Meeting of the Japanese Pharn acological Society
  • Place of Presentation: 横浜
• [Presentation] Non-kinase activity of MLCK in inducing filopodia and chemotaxis of vascular smooth muscle cells toward sphingosylphosphorylcholine. (2008)
  • Author(s): Wang H. -H., Nakamura A., Matsumoto A., Yoshiyama S., Zin X., Ye L.-H., Xie C., Zhang Y., Gao Y., Ishikawa R., Kohama K.
  • Organizer: The 82nd Annual Meeting of the Japanese Pharn acological Society
  • Place of Presentation: 横浜
• [Presentation] Characterization of the kinase-dead mutants of smooth muscle myosin light chain kinase. (2008)
  • Author(s): Xie C., Nakamura A., Zhang Y., Gao Y., Ishikawa R., Yoshiyama S., Kawano K., Kohama K.
  • Organizer: The 82nd Annual Meeting of the Japanese Pharn acological Society
  • Place of Presentation: 横浜
• [Presentation] The analysis of calcium inhibition by constructing hybrid myosins between physarum and smooth muscle myosins. (2008)
  • Author(s): Zhang Y., Kawamichi H., Nakamura A., Nyitray L., Kohama K
  • Organizer: The 82nd Annual Meeting of the Japanese Pharn acological Society
  • Place of Presentation: 横浜
• [Presentation] In vitro sliding on myosin II of F-actin under the conditions fully saturated with variety of actin-binding proteins. (2008)
  • Author(s): Ishikawa R., Takahashi A., Nakamura A., and Kohama K.
  • Organizer: 2nd International Symposium on Bio-nanosystems
  • Place of Presentation: 東京
• [Presentation] Actin-bundles as a transporter of nano-sized cargoes. (2008)
  • Author(s): Yoshiyama S., Ishikawa R., Nakamura A., Higashi-Fujime S., Takatsuki H., Rice K., Blough E.R., and Kohama K.
  • Organizer: 2nd International Symposium on Bio-nanosystems
  • Place of Presentation: 東京
• [Presentation] A novel phosphatase from Physarum polycephalum III. (2008)
  • Author(s): Okada C., Nakamura A., Yoshiyama S., Kohama K., Kaneko T.
  • Organizer: 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
  • Place of Presentation: 神戸
• [Presentation] Characterization of the non-kinase activity of smooth muscle myosin light chain kinase. (2008)
  • Author(s): Xie C., Nakamura A., Zhang Y., Gao Y., Ishikawa R., Yoshiyama S., Kohama K.
  • Organizer: 48th annual meeting of the American Society for Cell Biology
  • Place of Presentation: San Francisco, USA
• [Presentation] Effect of a high-fat diet on diabetic mother rats and their offsprings and insulin signaling. (2008)
  • Author(s): Nasu R., Nakamura A., Kakarla S., Blough E.R., Kohama K., Kohama T.
  • Organizer: 48th annual meeting of the American Society for Cell Biology
  • Place of Presentation: San Francisco, USA
• [Presentation] The non-kinase activity of myosin light chaina kinase in regulating smooth muscle contraction (2008)
  • Author(s): Nakamura A.
  • Organizer: The 81^st Annual Meeting of The Japanese Pharmacological Society
  • Place of Presentation: Yokohama
• [Presentation] Screening of phenotype-specific genes of vascular smooth muscle cell(VSMC)and their characterization (2008)
  • Author(s): Miyamoto H., Nagahama R, Nakamura A., Kohama K., Okagaki T.
  • Organizer: The 81^stAnnual Meeting of The Japanese Pharmacological Society
  • Place of Presentation: Yokohama
• [Presentation] Brain specific acin-binding protein Drebrin inhibits the cofilin-dependent actin dynamics (2008)
  • Author(s): Ishikawa R., Katoh K., Ebihara S., Takahashi A., Igarashi M., Nakamura A., Kohama K
  • Organizer: The 81^stAnnual Meeting of The Japanese Pharmacological Society
  • Place of Presentation: Yokohama
• [Presentation] Ultrastructural change of myosin filaments in smooth muscle cell with blebbistatin (2008)
  • Author(s): Tanaka H., Watanabe M., Wang H-H., Kohama K.
  • Organizer: The 81st Annual Meeting of The Japanese Phamlacological Society
  • Place of Presentation: Yokohama
• [Presentation] Arachidonic acid binds to heavy chain of smooth muscle myosin and stimulates ATPase activity (2008)
  • Author(s): Katayama T., Miyakawa T., Yoshiyama S., Nakamura A., Tanaka H., Tanokura M., Kohama K
  • Organizer: The 81st Annual Meeting of The Japanese Pharmacological Society
  • Place of Presentation: Yokohama
• [Presentation] Non-kinase activity of MLCK in elongated filopodia formation and chemotaxis of vascular smooth muscle cells in response to sphingosylphosphorylcholcholine (2008)
  • Author(s): Wang H-H., Nakamura A., Matsumoto A., Hayakawa K, Yoshiyama S., Qin X., Tanaka H., Ye L., Li S., Ishikawa R., Kohama K
  • Organizer: The 81st Annual Meeting of The Japanese Pharmacological Society
  • Place of Presentation: Yokohama
• [Presentation] Effect of a high-fat diet on diabetic mother rats and their offsprings and insulin signaling (2008)
  • Author(s): Nasu R., Kohama T., Kohama K., Kakarla S., Blough E. R
  • Organizer: The 81st Annual Meeting of The Japanese Phamiacological Society
  • Place of Presentation: Yokohama
• [Presentation] Dynamics of actomyosin and intracellular Ca^<2+>during cell movement in case of Phsarum polycephalum (2008)
  • Author(s): Yoshiyama S., Ishigami M., Kohama K
  • Organizer: The 81st Annual Meeting of The Japanese Pharmacological Society
  • Place of Presentation: Yokohama
• [Presentation] Expression and characterization of the kinase-dead mutants of myosin light chain kinase (2008)
  • Author(s): Xie C., Nakamura A., Zhang Y, Gao Y., Kawano K., Ishikawa R, Matsumoto A., Tanaka H., Kohama K.
  • Organizer: The 81st Annual Meeting of The Japanese Pharmacological Society
  • Place of Presentation: Yokohama
• [Presentation] The regulatory effect of Ca^<2+>/calmodulin on the actin-binding acticity of smooth muscle myosin lingt chain kinase (2008)
  • Author(s): Zhang Y., Xie C., Nakamura A., Gao Y., Ishikawa R., Yoshiyama S., Matsumoto A., Kohama K
  • Organizer: The 81st Annual Meeting of The Japanese Phammacological Society
  • Place of Presentation: Yokohama
• [Book] Peptide Science 2007 (2008)
  • Author(s): Arai K., Oku H., YamadA K., Gao Y., Ishiokawa R., Yoshiyama S., Kohama K.
  • Publisher: The Japanese Peptide Society.
• [Patent(Industrial Property Rights)] 相関観察法(蛍光抗体染色像・電子顕微鏡像重ね合わせ技法) (2009)
  • Inventor(s): 田中秀幸・小浜一弘・太田直哉・朝岡峻一郎・茂木健司
  • Industrial Property Rights Holder: 国立大学法人群馬大学
  • Filing Date: 2009-03-17