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ADAMファミリーメンバーのクローニングと機能解析

Research Project

Project/Area Number 11240205
Research Category

Grant-in-Aid for Scientific Research on Priority Areas

Allocation TypeSingle-year Grants
Review Section Biological Sciences
Research InstitutionBeppu University (2003)
大分医科大学 (1999-2002)

Principal Investigator

山本 俊輔  別府大学, 食物栄養学部, 教授 (90040188)

Co-Investigator(Kenkyū-buntansha) 松浦 恵子  大分大学, 医学部, 助手 (00291542)
泥谷 直樹  大分医科大学, 医学部, 助手 (80305036)
樋口 安典  大分医科大学, 医学部, 助教授 (60040284)
Project Period (FY) 1999 – 2003
Project Status Completed (Fiscal Year 2003)
Budget Amount *help
¥42,000,000 (Direct Cost: ¥42,000,000)
Fiscal Year 2003: ¥7,000,000 (Direct Cost: ¥7,000,000)
Fiscal Year 2002: ¥7,400,000 (Direct Cost: ¥7,400,000)
Fiscal Year 2001: ¥9,800,000 (Direct Cost: ¥9,800,000)
Fiscal Year 2000: ¥8,000,000 (Direct Cost: ¥8,000,000)
Fiscal Year 1999: ¥9,800,000 (Direct Cost: ¥9,800,000)
KeywordsADAM15 / バリアント蛋白 / Lck / Hck / Src / T細胞活性化 / ADAM6 / shedding / ADAMファミリー / スプライスバリアント / Srcファミリー / ADAM8 / CD156 / アポトーシス / メタロプロテアーゼ / ジスインテグリン / トランスジェニック / ADAM17 / TACE / ADAM / 単クローン抗体 / AD56
Research Abstract

ADAM15:ヒトADAM15のバリアント:細胞内ドメインに25個とそれに続く24個計49個のアミノ酸の挿入配列を持つADAM15v1とADAM15v2を見出した。この細胞内配列はプロリンに富むクラスIIおよびクラスIのSH3結合配列から成り、ADAM12の細胞内ドメインのそれに酷似するが、ADAM15v2はこの配列をタンデムに持つことからより強力な構造である。このバリアントmRNAはT細胞およびマクロファージに強く発現している。ADAM15v2のSH3結合配列に対する単クローン抗体(mAb)を作成し、免疫沈降法でバリアント蛋白の発現を確認した。マウスのADAM15遺伝子の全塩基配列を決定し、イントロン19内に同様のバリアントが存在することを確認した。マウスのADAM15v2は24個のSH3結合配列はクラスIIの1個のコンセンサス配列の違いから、クラスIのみの配列である。ヒトおよびマウスのADAM15v2とSrcファミリー蛋白との結合性や結合度をADAM15のそれと比較検討するために、Lck、Hck-GST蛋白、非燐酸化および燐酸化ADAM15およびADAM15v2HIS蛋白を作成しプルダウン法および免疫沈降法でADAM15v2がより強く、また、燐酸化蛋白がより強く反応することを確認した。さらに、非燐酸化および燐酸化ADAM15およびADAM15v2GST蛋白を作成し,抗LckあるいはHck抗体を用いてJurkat細胞やHL60細胞のLckあるいはHckが燐酸化ADAM15v2により強く反応することを確認した。また、マウスのEL4細胞では類似の方法で燐酸化LckおよびSrcが強く反応することを確認した。また、JurkatやEL4細胞をミリスチン酸や抗CD3抗体刺激で燐酸化ADAM15v2により強く反応することを観察した。Jurkat細胞では抗CD3抗体刺激でLFA-1とICAM-1の結合が亢進するが、この反応がBioporterを用いた抗ADAM15v2mAbの細胞内注入により阻害されることを確認した。(2)ADAM8(CD156):ADAM8のstabl transfectantを293T細胞に作成し、これに、TNF-α,HB-EGF,L-selectin,FasL,CD44などをtransient transfectし、免疫沈降法でTNF-αおよびHB-EGFが遊離されること(shedding)を確認した。TNF-αでは遊離をELISA法でも確認した。細胞内ドメインにはクラスIIおよびクラスI配列が存在し、これらにHckやLckが強く反応することを観察した。

Report

(5 results)
  • 2003 Annual Research Report
  • 2002 Annual Research Report
  • 2001 Annual Research Report
  • 2000 Annual Research Report
  • 1999 Annual Research Report
  • Research Products

    (48 results)

All Other

All Publications (48 results)

  • [Publications] Higuchi, Y.: "CD156 transgenic mice : different responses between inflammatory types"Pathobiology. 7・1. 47-54 (2002)

    • Related Report
      2003 Annual Research Report
  • [Publications] Shimizu, E: "Structure and expression of the murine ADAM 15 gene and its splice variants"Biochim.Biophys.Res.Commun.. 309・4. 779-785 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Yasui, A.: "Expression of splice variants of the human ADAM 15 gene, and strong interaction between the cytoplasmic domain"Pathobiology. (印刷中).

    • Related Report
      2003 Annual Research Report
  • [Publications] 山本俊輔: "ADAMファミリー蛋白"炎症と免疫. 11・4. 516-518 (2003)

    • Related Report
      2003 Annual Research Report
  • [Publications] Yasui, A.: "Leukocyte Typing VII White Cell Differentiation Antigens"Oxford University Press. 366-367 (2002)

    • Related Report
      2003 Annual Research Report
  • [Publications] Higuchi, Y. 他: "CD156 transgenic mice : different responses between inflammatory types"Pathobiology. 70(1). 47-54 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Hijiya, N. 他: "Possible involvement of Toll-like receptor 4 in endothelial cell activation of larger vessels in response to lipopolysaccharide"Pathobiology. 70(1). 18-25 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Higuchi, Y. 他: "Mouse soluble CD14 truncated at amino acid 71 in transgenic mice : Preventive effect on endotoxin-mediated toxic shock"Pathobiology. 70(2). 103-114 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Liu, S.-Y. 他: "Expression of murine N-myc by insertion of retrovirus sequences in a murine macrophage cell line, RAW264"Pathobiology. (In press). (2003)

    • Related Report
      2002 Annual Research Report
  • [Publications] Higuchi, Y. 他: "The roles of soluble osteopontin using osteopontin-transgenic mice in vivo : proliferation of CD4^+ T lymphocytes and the enhancement of cell-mediated immune responses"Pathobiology. (In press). (2003)

    • Related Report
      2002 Annual Research Report
  • [Publications] Yasui, A. 他: "Leukocyte Typing VII White Cell Differentiation Antigens"Oxford University Press. 2 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Matsuno, O.: "Leukocyte Typing VII White Cell Differentiation"Oxford University Press(in press).

    • Related Report
      2001 Annual Research Report
  • [Publications] Yasui, A.: "Leukocyte Typing VII White Cell Differentiation"Oxford University Press(in press).

    • Related Report
      2001 Annual Research Report
  • [Publications] Matsuno, O.: "Leukocyte Typing VII White Cell Differentiation"Leukocyte Typing VII(in press).

    • Related Report
      2001 Annual Research Report
  • [Publications] Schlomann,U.: "Tumor necrosis factor (TNF) alpha induces"J.Neurosci.. 20(21). 7964 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Wang,D.: "Transcriptional regulation of the human osteopontin"Oncogene. 19(50). 5801 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Matsuura,K.: "Involvement of CD14 in LPS-induced liver injury"J.Enotox.Res.. (In press).

    • Related Report
      2000 Annual Research Report
  • [Publications] 山本俊輔: "ADAMファミリーと炎症"感染・炎症・免疫. 30(1). 12 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] 泥谷直樹: "エンドトキシシ研究シリーズ2 新しい展開"菜根出版. 4 (2000)

    • Related Report
      2000 Annual Research Report
  • [Publications] Matsuno,O.: "Leukocyte Typing VII White Cell Differentiation"Oxford University Press (in press).

    • Related Report
      2000 Annual Research Report
  • [Publications] Matsuno,O.: "Leukocyte Typing VII White Cell Differentiation"Oxford Unviersity Press (in press).

    • Related Report
      2000 Annual Research Report
  • [Publications] Yasui,A.: "Leukocyte Typing VII White Cell Differentiation"Oxford University Press (in press).

    • Related Report
      2000 Annual Research Report
  • [Publications] Tamura, Y.他: "CD14 transgenic mice expressing membrane and soluble forms"Int. Immunol.. 11(3). 333-339 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Hisaka, H.他: "Close localization of mouse CD14 and CD32/16 in the cell surface of"Pathobiology. 67(2). 92-97 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Yamamoto, S.他: "ADAM family proteins in immune system"Immunol. Today. 20(5). 278-284 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Ohki, K.他: "Suppressive effects of serum on the LPS-induced production of nitric oxide"Immunol. Cell. Biol.. 77(2). 143-152 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Yamamoto, T.他: "Expression of scavenger receptor class A and CD14 in lipopolysaccharide-"Pathol. Int.. 49(11). 983-992 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 山本俊輔他: "エンドトキシンショック"病理と臨床(臨時増刊号). 17. 320 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 山本俊輔: "自然免疫鳥瞰ー一次防御、および獲得免疫の支持と利用"Molecular Medicine. 36(5). 468-474 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] 山本俊輔: "LPS臓器障害とマクロファージ"病理と臨床. 169,(). 1100-1111 (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Yoshiyama, K.他: "CD156(human ADAM8): expression, primary amino acid sequence, and"Genomics.. 41(1). 56-62 (1997)

    • Related Report
      1999 Annual Research Report
  • [Publications] Takai, N.他: "Primary structure of rat CD14 and characteristics of rat CD14, cytokine"J. Leuko. Biol.. 61(6). 736-744 (1997)

    • Related Report
      1999 Annual Research Report
  • [Publications] Shinji, H.他: "Lipopolysaccharide-induced biphasic inositol 1, 4, 5-triphosphate response"J. Immunol.. 158(2). 1370-1376 (1997)

    • Related Report
      1999 Annual Research Report
  • [Publications] Kataoka, M.他: "Structure of the murine CD156 gene, characterization of its promotor"J. Biol. Chem.. 272(29). 18209-18215 (1997)

    • Related Report
      1999 Annual Research Report
  • [Publications] Shinji, H.他: "Lipopolysaccaride-induced biphasic inositol 1. 4, 5-triphosphate response"J. Immunol.. 158(2). 1370-1376 (1996)

    • Related Report
      1999 Annual Research Report
  • [Publications] Onozuka, K.他: "Participation of CD14 in the phagocytosis of smooth-type Salmonella"Microbiol. Immunol.. 41(10). 765-772 (1997)

    • Related Report
      1999 Annual Research Report
  • [Publications] Kirikae T.他: "Rhodobacter spheroides diphosphoryl lipid A inhibits interleukin-6"J. Endotox. Res.. 4(2). 115-122 (1997)

    • Related Report
      1999 Annual Research Report
  • [Publications] Morita, H.他: "Difference in clearance of exogenously administered smooth-form LPS"J. Endotox. Res.. 4(6). 415-423 (1997)

    • Related Report
      1999 Annual Research Report
  • [Publications] 樋口安典他: "CD14トランスジェニックマウス"臨床免疫. 29(11). 1345-1350 (1997)

    • Related Report
      1999 Annual Research Report
  • [Publications] Haziot, A.他: "Resistance to endotoxin shock and reduced dissemination of Gram-negative"Immunity. 4(4). 407-414 (1996)

    • Related Report
      1999 Annual Research Report
  • [Publications] 山本俊輔他: "CD156(ADAM 8)の構造、機能およびファミリー"Molecular Medicine34(臨時増刊号)免疫1997-98. 148-162 (1997)

    • Related Report
      1999 Annual Research Report
  • [Publications] Matsuura, K.他: "Involvement of CD14 in LPS-induced liver"Wiley-Liss. in press (1999)

    • Related Report
      1999 Annual Research Report
  • [Publications] Yammoto, S.他: "Roles of CD14 in LPS signaling and scavenging: studies using CD14-"Weiley-Liss. 8 (1998)

    • Related Report
      1999 Annual Research Report
  • [Publications] Higuchi, Y.: "CD156 (ADAM 8) workshop panel report. Leukocyte Typing VI."Garland Publ Inc. 3 (1998)

    • Related Report
      1999 Annual Research Report
  • [Publications] Kataoka, M.: "Generation and analysis of CD14-transgenic mice. Leukocyte Typing VI."Garland Publ Inc. 2 (1998)

    • Related Report
      1999 Annual Research Report
  • [Publications] Takai, N.他: "Structure of rat CD14, and expression of rat CD14 and cytokines"Oxford Univ. Press. 3 (1998)

    • Related Report
      1999 Annual Research Report
  • [Publications] Kataoka, M.他: "Generation and analysis of CD14-transgenic mice"Oxford Univ. Press. 2 (1998)

    • Related Report
      1999 Annual Research Report
  • [Publications] 片岡晶志: "エンドキシン感受性亢進および低下マウス"菜根出版. 12 (1998)

    • Related Report
      1999 Annual Research Report

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Published: 1999-04-01   Modified: 2018-03-28  

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