低分子量Gタンパク質Rabをゲノムワイドで網羅的に解析するツールの開発

• Project/Area Number: 17657067
• Research Category: Grant-in-Aid for Exploratory Research
• Allocation Type: Single-year Grants
• Research Field: Cell biology
• Research Institution: Tohoku University (2006); The Institute of Physical and Chemical Research (2005)
• Principal Investigator: 福田 光則 東北大学, 大学院生命科学研究科, 教授 (50311361)
• Project Period (FY): 2005 – 2006
• Project Status: Completed (Fiscal Year 2006)
• Budget Amount: ¥3,500,000 (Direct Cost: ¥3,500,000); Fiscal Year 2006: ¥1,700,000 (Direct Cost: ¥1,700,000); Fiscal Year 2005: ¥1,800,000 (Direct Cost: ¥1,800,000)
• Keywords: 低分子量G蛋白質 / Rab / Rabエフェクター / 膜輸送 / dominant active / dominant negative / ゲノムワイド / 網羅的解析

Research Abstract

真核細胞生物の細胞内はさらに膜で包まれたオルガネラにより満たされており、主に膜輸送を介してオルガネラ同士で頻繁に物質のやり取りを行っている。低分子量Gタンパク質Rabファミリーは膜輸送に関与する分子群としては最大のもめで、哺乳動物においては約60種類ものアイソフォームが存在している。各々のRabは特定の膜輸送を制御すると考えられているが、12番以降のRabアイソフォームになるといつどこで何を運ぶのかに関してはほとんど解明されていない。Rabは活性化型固定化(domihant active, DA)変異体あるいは不活性化型固定化(dominant negative, DN)変異体を作成することが出来るため、特走の膜輸送に対する影響を検討することが可能であるが、ある種の受容体が未知のRabにより細胞膜に輸送される場合には、60種類全てのRabクローンに対してDA/DN変異体を作成する必要があり、60種類のうちどのRabが関与するのかを調べることはこれまで不可能であった。このため世界的にRabを綢羅的に解析するツールのニーズが高まってきている。そこで本研究ではマウス及びヒトに存在する60種類のRabを全てクローニングし、これらのRabの機能を網羅的に解析するツーノレ(「Rab bank」)を作成することを目指した。本年度は、これまでに作成したRabの網羅的廨析ツールを利用し、実際に内分泌細胞(PC12細胞)からのホルモン分泌に関わるRabのスクリーニングを行った。その結果、60種類のRabのうちPC12細胞のホルモン顆粒上にはRab3AとRab27Aの二種類のRabが存在し、お互いに協調的にホルモン顆粒の細胞膜へめ繋ぎ止めやホルモンの放出に関与することを明らかにすることができた(J. Cell Scl.(2006)119:2196-2203)。

Research Products

• [Journal Article] The polybasic sequence in the C2B domain of rabphilin is required for the vesicle docking step in PC12 cells. (2007)
  • Author(s): Tsuboi, T., Kanno, E., Fukuda, M.
  • Journal Title: J. Neurochem. 100 Pages: 770-779
• [Journal Article] Dvl regulates endo- and exocytotic processes through binding to synaptotagmin. (2007)
  • Author(s): Kishida, S., Hamao, K., Inoue, M., Hasegawa, M., Matsuura, Y., Mikoshiba, K., Fukuda, M., Kikuchi, A.
  • Journal Title: Genes Cells 12 Pages: 49-61
• [Journal Article] Versatile roles of R-Ras GAP in neurite formation of PC12 cells and embryonic vascular development. (2007)
  • Author(s): Iwashita, S., Kobayashi, K., Kubo, Y., Hinohara, Y., Sezaki, M., Nakamura, K., Suzuki-Migishima, R., Yokoyama, M., Satoh, S., Fukuda.M., Ohba, M., Kato, C., Adachi, E., Song, S.-Y.
  • Journal Title: J. Biol. Chem. 282 Pages: 3413-3417
• [Journal Article] Live cell imagingによる細胞活動の可視化解析 (2007)
  • Author(s): 坪井貴司, 福田光則
  • Journal Title: 日本比較内分泌学会ニュース 124(2) Pages: 45-53
  • NAID: 10018293579
• [Journal Article] Distinct Rab27A binding affinities of Slp2-a and Slac2-a/melanophilin : Hierarchy of Rab27A effectors. (2006)
  • Author(s): Fukuda, M.
  • Journal Title: Biochem. Biophys. Res. Commun. 343 Pages: 666-674
• [Journal Article] Rab27a regulates epithelial sodium channel (ENaC) activity through synaptotagmin-like protein (SLP-5) and Munc13-4 effector mechanism. (2006)
  • Author(s): Saxena, S.K., Horiuchi, H., Fukuda, M.
  • Journal Title: Biochem. Biophys. Res. Commun. 344 Pages: 651-657
• [Journal Article] The Slp4-a linker domain controls exocytosis through interaction with Munc18-1・syntaxin-la complex. (2006)
  • Author(s): Tsuboi, T., Fukuda, M.
  • Journal Title: Mol. Biol. Cell 17 Pages: 2101-2112
• [Journal Article] Rab3A and Rab27A cooperatively regulate the docking step of densecore vesicle exocytosis in PC12 cells. (2006)
  • Author(s): Tsuboi, T., Fukuda, M.
  • Journal Title: J. Cell Sci. 119 Pages: 2196-2203
• [Journal Article] Decreased basal mucus secretion by Slp2-a-deficient gastric surface mucous cells. (2006)
  • Author(s): Saegusa, C., Tanaka, T., Tani, S., Itohara, S., Mikoshiba, K., Fukuda, M.
  • Journal Title: Genes Cells 11 Pages: 623-631
• [Journal Article] Regulation of synaptic transmission by RAB-3 and RAB-27 in Caenorhabditis elegans. (2006)
  • Author(s): Mahoney, T.R., Liu, Q., Itoh, T., Luo, S., Hadwiger, G., Vincent, R., Wang, Z.-W., Fukuda, M., Nonet, M.L.
  • Journal Title: Mol. Biol. Cell 17 Pages: 2617-2625
• [Journal Article] Chronic olanzapine treatment causes differential expression of genes in frontal cortex of rats as revealed by DNA microarray technique. (2006)
  • Author(s): Fatemi, S.H., Reutiman, T.J., Folsom, T.D., Bell, C., Nos, L., Fried, P., Pearce, D.A., Singh, S., Siderovski, D.P., Willard, F.S., Fukuda, M.
  • Journal Title: Neuropsychopharmacol. 343 Pages: 666-674
• [Journal Article] Screening for target Rabs of TBC (Tre-2/Bub2/Cdc16) domain-containing proteins based on their Rab-binding activity. (2006)
  • Author(s): Itoh, T., Satoh, M., Kanno, E., Fukuda, M.
  • Journal Title: Genes Cells 11 Pages: 1023-1037
• [Journal Article] Neuronal calcium sensor-1 (NCS-1) and PI4Kβ stimulate ERK1/2 signaling by accelerating recycling through endocytic recycling compartment (ERC). (2006)
  • Author(s): Kapp-Barnea, Y., Ninio-Many, L., Hirschberg, K., Fukuda, M., Jeromin, A., Sagi-Eisenberg, R.
  • Journal Title: Mol. Biol. Cell 17 Pages: 4130-4141
• [Journal Article] Constitutive GDP/GTP exchange and secretion-dependent GTP hydrolysis activity for Rab27 in platelets. (2006)
  • Author(s): Kondo, H., Shirakawa, R., Higashi, T., Kawato, M., Fukuda, M., Kita, T., Horiuchi, H.
  • Journal Title: J. Biol. Chem. 281 Pages: 28657-28665
• [Journal Article] Identification of EPI64 as a GTPase-activating protein specific for Rab27A. (2006)
  • Author(s): Itoh, T., Fukuda, M.
  • Journal Title: J. Biol. Chem. 281 Pages: 31823-31831
• [Journal Article] Functional involvement of Noc2, a Rab27 effector, in rat parotid acinar cells. (2006)
  • Author(s): Imai, A., Yoshie, S., Nashida, T., Shimomura, H., Fukuda, M.
  • Journal Title: Arch. Biochem. Biophys. 455 Pages: 127-135
• [Journal Article] Rab27 and its effectors in secretory granule exocytosis : A novel docking machinery composed of a Rab27・effector complex. (2006)
  • Author(s): Fukuda, M.
  • Journal Title: Biochem. Soc. Trans. 11 Pages: 691-695
• [Journal Article] シナプトタグミンの分子進化 (2006)
  • Author(s): 福田光則
  • Journal Title: 生体の科学 57(5) Pages: 458-459
• [Journal Article] Distinct developmental expression of synaptotagmin I and IX in the mouse brain. (2006)
  • Author(s): Fukuda, M.
  • Journal Title: NeuroReport 17 Pages: 179-182
• [Journal Article] O-glycosylation is essential for intracellular targeting of synaptotagmins I and II in non-neuronal specialized secretory cells. (2005)
  • Author(s): Atiya-Nasagi, Y., Cohen, H., Medalia, O., Fukuda, M., Sagi-Eisenberg, R.
  • Journal Title: J.Cell Sci. 118 Pages: 1363-1372
• [Journal Article] Classical protein kinase C(s) regulates targeting of synaptotagmin IX to the endocytic recycling compartment. (2005)
  • Author(s): Haberman, Y, Ziv, I., Gorzalczany, Y., Fukuda, M., Sagi-Eisenberg, R.
  • Journal Title: J.Cell Sci. 118 Pages: 1641-1649
• [Journal Article] Functional analysis of Slac2-c/MyRIP as a linker protein between melanosomes and myosin VIIa. (2005)
  • Author(s): Kuroda, T.S., Fukuda, M.
  • Journal Title: J.Biol.Chem. 280 Pages: 28015-28022
• [Journal Article] AS160, the Akt substrate regulating GLUT4 translocation, has a functional Rab GTPase activating protein domain. (2005)
  • Author(s): Miinea, C.P., Sano, H., Kane, S., Sano, E., Fukuda, M., Peranen, J., Lane, W.S., Lienhard, G.E.
  • Journal Title: Biochem.J. 391 Pages: 87-93
• [Journal Article] Protein Kinase C-mediated phosphorylation of the two polybasic regions of synaptotagmin VI regulates their function in acrosomal exocytosis. (2005)
  • Author(s): Roggero, C.M., Tomes, C.N., De Blas, G.A., Castillo, J., Michaut, M.A., Fukuda, M., Mayorga, L.S.
  • Journal Title: Dev.Biol. 285 Pages: 422-435
• [Journal Article] Adenovirus-mediated silencing of Synaptotagmin 9 inhibits Ca^<2+>-dependent insulin secretion in islets. (2005)
  • Author(s): Iezzi, M., Eliasson, L., Fukuda, M., Wollheim, C.B.
  • Journal Title: FEBS Lett. 579 Pages: 5241-5246
• [Journal Article] Rab proteins regulate epithelial sodium channel activity in colonic epithelial HT-29 cells. (2005)
  • Author(s): Saxena, S., Singh, M., Engisch, K., Fukuda, M., Kaur, S.
  • Journal Title: Biochem.Biophys.Res.Commun. 337 Pages: 1219-1223
• [Journal Article] The short apical membrane half-life of rescued ΔF508-cystic fibrosis transmembrane conductance regulator (CFTR) CFTR results from accelerated endocytosis of ΔF508-CFTR in polarized human airway epithelial cells. (2005)
  • Author(s): Swiatecka-Urban, A., Brown, A., Moreau-Marquis, S., Renuka, J., Coutermarsh, B., Barnaby, R., Karison, K.H., Flotte, T.R., Fukuda, M., Langford, G.M., Stanton, B.A.
  • Journal Title: J.Biol.Chem. 280 Pages: 36762-36772
• [Journal Article] Slp4-a/granuphilin-a interacts with syntaxin-2/3 in a Munc18-2-dependent manner. (2005)
  • Author(s): Fukuda, M., Imai, A., Nashida, T., Shimomura, H.
  • Journal Title: J.Biol.Chem. 280 Pages: 39175-39184
• [Journal Article] The C2B domain of rabphilin directly interacts with SNAP-25 and regulates the docking step of dense core vesicle exocytosis in PC12 cells. (2005)
  • Author(s): Tsuboi, T., Fukuda, M.
  • Journal Title: J.Biol.Chem. 280 Pages: 39253-39259
• [Journal Article] Cloning and genomic characterization of sytdep, a new synaptotagmin XIV-related gene. (2005)
  • Author(s): Herrero-Turrion, M.J., Fukuda, M., Mollinedo, F.
  • Journal Title: Biocheim.Biophys.Res.Commun. 340 Pages: 386-394
• [Journal Article] Slp homology domain : A novel protein motif that specifically binds small GTPase Rab27. (2005)
  • Author(s): Fukuda, M.
  • Journal Title: Recent Res.Dev.Biochem. 6 Pages: 13-29
• [Journal Article] Versatile role of Rab27 in membrane trafficking : Focus on the Rab27 effector families. (2005)
  • Author(s): Fukuda, M.
  • Journal Title: J.Biochem. 137 Pages: 9-16
  • NAID: 10017342344
• [Journal Article] Functional analysis of Slac2-a/melaonophilin as a linker protein between Rab27A and myosin Va in melanosome transport. (2005)
  • Author(s): Kuroda, T.S., Itoh, T., Fukuda, M.
  • Journal Title: Methods Enzymol. 403 Pages: 419-431
• [Journal Article] Identification and biochemical analysis of Slac2-c/MyRIP as a Rab27-, myosin Va/VIIa-, and actin-binding protein. (2005)
  • Author(s): Kuroda, T.S., Fukuda, M.
  • Journal Title: Methods Enzymol. 403 Pages: 431-444
• [Journal Article] Analysis of the role of Rab27 effector Slp4-a/granuphilin-a in dense-core vesicle exocytosis. (2005)
  • Author(s): Fukuda, M., Kanno, E.
  • Journal Title: Methods Enzymol. 403 Pages: 445-457
• [Journal Article] Assay and functional interactions of Rim2 with Rab3. (2005)
  • Author(s): Fukuda, M.
  • Journal Title: Methods Enzymol. 403 Pages: 457-468
• [Journal Article] Assay of Rab binding specificity of rabphilin and Noc2 : Target molecules for Rab27. (2005)
  • Author(s): Fukuda, M., Yamamoto, A.
  • Journal Title: Methods Enzymol. 403 Pages: 469-481
• [Journal Article] Slac2-aとSlp2-a:メラニン色素輸送のしくみ (2005)
  • Author(s): 福田光則
  • Journal Title: 医学のあゆみ 212 Pages: 769-770
• [Journal Article] シナプトタグミンと類似タンパク質による膜輸送制御の分子メカニズム (2005)
  • Author(s): 福田光則
  • Journal Title: 生化学 77 Pages: 603-618
  • NAID: 10016603083
• [Journal Article] 「メラニン色素」輸送の分子メカニズム:肌や髮の毛が黒くなる仕組み (2005)
  • Author(s): 福田光則
  • Journal Title: 日皮協ジャーナル 54 Pages: 60-66
• [Journal Article] 美白への新たなアプローチ-メラニン輸送をストップさせる- (2005)
  • Author(s): 黒田垂歩, 福田光則
  • Journal Title: BIONICS 2(11) Pages: 68-71
  • NAID: 40007003528
• [Book] Molecular Mechanisms of Exocytosis (2006)
  • Author(s): Fukuda, M.
  • Publisher: The role of synaptotagmin and synaptotagmin-like protein (Slp) in regulated exocytosis.
• [Book] アンチ・エイジングシリーズ1「白髪・脱毛・育毛の実際」メラニン色素輸送メカニズムの解明 (2005)
  • Author(s): 福田光則, 黒田垂歩
  • Publisher: エヌ・ティー・エス