抗がん遺伝子産物ＳＴＡＲＴ－ＧＡＰ／ＤＬＣファミリータンパク質の細胞内局在化機構

• Project/Area Number: 22501016
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Tumor biology
• Research Institution: University of Hyogo
• Principal Investigator: 八木澤 仁 兵庫県立大学, 生命理学研究科, 准教授 (40192380)
• Project Period (FY): 2010-04-01 – 2013-03-31
• Project Status: Completed (Fiscal Year 2012)
• Budget Amount: ¥4,290,000 (Direct Cost: ¥3,300,000、Indirect Cost: ¥990,000); Fiscal Year 2012: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000); Fiscal Year 2011: ¥1,300,000 (Direct Cost: ¥1,000,000、Indirect Cost: ¥300,000); Fiscal Year 2010: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000)
• Keywords: ホスホリパーゼC / 生体分子 / 細胞組織 / がん抑制 / 細胞接着斑 / 酵素 / 細胞・組織 / 癌 / シグナル伝達 / アクチン骨格

Research Abstract

遺伝子の欠失やプロモーター領域の不活化により、腫瘍形成の原因と なっていることが報告されているSTART-GAP(START domain-containing Rho GTPase-activating protein、別名 DLC: deleted in liver cancer)ファミリー遺伝子産物の3種類のアイソフォームのうち、START-GAP1/DLC1について、(1)細胞内局在化調節機構を解析し、このタンパク質の細胞内分布と生理機能との関係を解明する。( 2)機能ドメインであるSTARTドメイン、および、GAPドメイン、ならびに、全長タンパク質の大量発現系と精製 系を確立し、タンパク質結晶化とX線結晶構造解析を行うことによって、その構造を明らかにする ことを目的 とする。その結果に基づき、冠攣縮性狭心症 (CSA) の原因解明を目指す。というのが本年度の目標であった。このうち前者については、HeLa細胞において、EGF刺激で一過性のSTART-GAP1のリン酸化亢進がおこることをかを確認した。また、PP2以外のリン酸化阻害剤をいくつかスクリーニングしたが、focal adhesion kinase (FAK)阻害剤で同様の影響が見られることが示された。(2)については目的の機能ドメインに対応するSTART-GAP1のGST-融合発現系を構築し、これらを用いて、正常型およびCSAの組換えタンパクをいずれもmg単位で得ることができた。しかし、いずれも精製に困難をともなっており、結晶化スクリーニングの段階に至っていない。また関連研究としてSTART-GAP1に結合して活性化し、細胞の運動機構に関与していると思われる新規ホスホリパーゼCをクローニングした。

Current Status of Research Progress

Reason

24年度が最終年度であるため、記入しない。

Strategy for Future Research Activity

24年度が最終年度であるため、記入しない。

Research Products

• [Journal Article] DGKζ is degraded through the cytoplasmic ubiquitin-proteasome system under excitotoxic conditions, which causes neuronal apoptosis because of aberrant cell cycle reentry (2012)
  • Author(s): Okada M, Hozumi Y, Tanaka T, Suzuki Y, Yanagida M, Araki Y, Evangelisti C, Yagisawa H, Topham MK, Martelli AM, Goto K.
  • Journal Title: Cell Signal Volume: 24 Issue: 8 Pages: 1573-82
  • DOI: 10.1016/j.cellsig.2012.03.021
  • Peer Reviewed
• [Journal Article] Membrane-induced alteration of the secondary structure in the SWAP-70 pleckstrin homology domain (2012)
  • Author(s): Tokuda N, Kawai K, Lee YH, Ikegami T, Yamaguchi S, Yagisawa H, Fukui Y, Tuzi S
  • Journal Title: J Biochem Volume: 151 Issue: 4 Pages: 391-401
  • DOI: 10.1093/jb/mvr146
  • NAID: 10031166078
  • Peer Reviewed
• [Journal Article] DGKzeta is involved in LPS-activated phagocytosis through IQGAP1/Rac1 pathway (2012)
  • Author(s): Okada, M.
  • Journal Title: Biochem Biophys Res Commun Volume: 420 Issue: 2 Pages: 479-84
  • DOI: 10.1016/j.bbrc.2012.03.057
  • Peer Reviewed
• [Journal Article] NAP Interaction of nucleosome assembly proteins abolishes nuclear localization of DGKζ by attenuating its association with importins (2011)
  • Author(s): Okada M, Hozumi Y, Ichimura T, Tanaka T, Hasegawa H, Yamamoto M, Takahashi N, Iseki K, Yagisawa H, Shinkawa T, Isobe T, Goto K.
  • Journal Title: Exp Cell Res Volume: 317 Issue: 20 Pages: 2853-63
  • DOI: 10.1016/j.yexcr.2011.09.014
  • Peer Reviewed
• [Journal Article] START-GAP1/DLC1 is localized in focal adhesions through interaction with the PTB domain of tensin2. (2010)
  • Author(s): Kawai, K., et al.
  • Journal Title: Advances in Enzyme Regulation Volume: 50 Pages: 202-215
  • Peer Reviewed
• [Presentation] Interaction between START-GAP/DLC ; a family of novel antioncogene products, and tensin, a component of a focal adhesion complex (2012)
  • Author(s): H Yagisawa, et al.
  • Organizer: 神戸大学先端融合科学シンポジウム
  • Place of Presentation: 神戸大学理学部(兵庫県)
  • Year and Date: 2012-01-31
• [Presentation] Cloning and characterization of phosphoinositide-specific phospholipase C of a giant amoeba, Amoeba proteus. (2012)
  • Author(s): Yagisawa, H
  • Organizer: 45回日本発生生物学会・64回日本細胞生物学会合同大会
  • Place of Presentation: 神戸国際会議場（兵庫県）
• [Presentation] The START-GAP/DLC family: antioncogene products that act as RhoGAPs and activators of phospholipase C (2012)
  • Author(s): Yagisawa, H
  • Organizer: FASEB Science Research Conference “Lipid Signaling Pathways in Cancer”
  • Place of Presentation: コロラド、アメリカ合衆国
• [Presentation] Amoeba proteusイノシトールリン脂質特異的ホスホリパーゼC (PI-PLC) の構造と性質 (2012)
  • Author(s): Yagisawa H.
  • Organizer: 45回日本原生動物学会
  • Place of Presentation: 兵庫県立大学（兵庫県）
• [Presentation] A new prospect of START-GAP1/DLC1 (deleted in liver cancer-1) as a pathogen for coronary spastic angina (CSA). (2012)
  • Author(s): Yagisawa H.
  • Organizer: The 17th Japan-Korea Cancer Research Workshop
  • Place of Presentation: 釜山（韓国）
• [Presentation] Cloning of a gene encoding a novel type of phosphoinositide-specific phospholipase C from a giant amoeba, Amoeba proteus. (2012)
  • Author(s): Yagisawa H.
  • Organizer: 35回日本分子生物学会年会
  • Place of Presentation: 福岡国際会館（福岡県）
• [Presentation] START-GAP1 (DLC1), a novel antioncogene product, transiently delocalizes from focal adhesions under EGF stimulus (2012)
  • Author(s): H Yagisawa
  • Organizer: The 7th Korea-Japan Conference on Cellular Signalling for Young Scientist
  • Place of Presentation: Ulsan National Institute of Science and Technology,(ウルサン、大韓民国)
• [Presentation] EGF regulates localization of START-GAP1 (DLC1), a novel antioncogene product, at focal adhesions (2011)
  • Author(s): M Ikita, H Nishitani, H Yagisawa
  • Organizer: 第34回分子生物学会年会
  • Place of Presentation: パシフィコ横浜(神奈川県)
  • Year and Date: 2011-12-14
• [Presentation] Changes in the secondary structure of the SWAP-70 PH domain induced at the lipid bilayer surface (2011)
  • Author(s): S Tuzi, et al., H Yagisawa
  • Organizer: The 10th JBS Biofrontier Symposium commemorating Kyushu University Centennial Anniversary International Symposium : New Aspects of Phospholipid Biology and Medicine 2011
  • Place of Presentation: ホテルルイガンス(福岡県)
  • Year and Date: 2011-11-16
• [Presentation] Calcium fluxes cause nuclear shrinkage and the translocation of phospholipase C-δ1 into the nucleus (2011)
  • Author(s): H Yagisawa, M Okada
  • Organizer: Symposium : "The diversity of inositide signaling" The 54th Annual Meeting of the Japanese Society for Neurochemistry
  • Place of Presentation: 山代温泉郷(石川県)(招待講演)
  • Year and Date: 2011-09-26
• [Presentation] Measurement of inositol polyphosphates by ion chromatography and their physiological status in higher plant (2011)
  • Author(s): T Mimura, et al., H Yagisawa
  • Organizer: Symposium : "The diversity of inositide signaling" The 54th Annual Meeting of the Japanese Society for Neurochemistry
  • Place of Presentation: 山代温泉郷(石川県)(招待講演)
  • Year and Date: 2011-09-26
• [Presentation] 抗がん遺伝子産物START-GAP1(DLC1)は接着斑構成因子vinculinのC末端領域を介して結合する (2011)
  • Author(s): 八木澤仁、四方佑斉、生田桃子
  • Organizer: 第84回日本生化学会大会
  • Place of Presentation: 京都国際会館(京都府)
  • Year and Date: 2011-09-23
• [Presentation] Mechanisms of nucleocytoplasmic transport of phospholipase C-δ1 and its role in cellular function (2011)
  • Author(s): YAGISAWA, Hiotoshi
  • Organizer: GORDON RESEARCH CONFERENCE : Signal Transduction in the Nucleus
  • Place of Presentation: Four Points Sheraton Ventura(U.S.A.)
  • Year and Date: 2011-03-01
• [Presentation] Nuclear-cytoplasmic shuttling of PLCδ1 and its implication in cell proliferation (2010)
  • Author(s): YAGISAWA, Hitoshi
  • Organizer: BMB2010第33回日本生化学会・第83回日本分子生物学会合同大会
  • Place of Presentation: 神戸国際会議場(兵庫県)
  • Year and Date: 2010-12-08
• [Presentation] がん抑制遺伝子産物START-GAP1(DLC1)は接着斑構成分子vinculinとLDモチーフを介して結合する (2010)
  • Author(s): 生田桃子、四方佑斉, 他、八木澤仁
  • Organizer: BMB2010第33回日本生化学会・第83回日本分子生物学会合同大会
  • Place of Presentation: 神戸国際会議場(兵庫県)
  • Year and Date: 2010-12-07
• [Presentation] Interaction with tensin2 PTB domain regulates tarteting of START-GAP1/DLC1 to punctatedautophagosome-like structures. (2010)
  • Author(s): MAEHIRA, K, et al.
  • Organizer: 第62回日本細胞生物学会大会
  • Place of Presentation: 大阪国際会議場(大阪府)
• [Presentation] Characterization of the START domain of p122RhoGAP/START-GAP/DLC-1 (2010)
  • Author(s): YAGISAWA, H, BANIK, G, YAMAGA, M
  • Organizer: The 27th Naito Conference on Membrane Dyanmics and Lipid Biology[I]
  • Place of Presentation: シャトレーゼ・ガトーキングダム札幌(北海道)
• [Presentation] Conformational alterations of the pleckstrin homology domains at the pid bilayer surface (2010)
  • Author(s): TUZI, S, et al.
  • Organizer: The 2010 International Chemical Congress of Pacific Basin Societies(PACIFICHEM 2010)
  • Place of Presentation: Hawaii Convention Center(U.S.A.)
• [Book] Chapter: START-GAP/DLC family proteins are novel antioncogene products. In “Future aspects of tumor suppressor genes” ISBN 978-953-51-1063-7 (2013)
  • Author(s): Yagisawa H.
  • Publisher: InTech
• [Remarks]
  • URL: http://www.sci.u-hyogo.ac.jp/life/biosig/japanese/rinku_1.html
• [Remarks]
  • URL: http://www.sci.u-hyogo.ac.jp/life/biosig/index.html