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Elucidation of the Mechanism for the cell cycle regulation by a novel isoform of p27^<Kip1> in the vascular cells

Research Project

Project/Area Number 13670723
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Circulatory organs internal medicine
Research InstitutionKYUSHU UNIVERSITY

Principal Investigator

HIRANO Katsuya  KYUSHU UNIV, GRAD SCH MED SCI, LECTURER, 大学院・医学研究院, 講師 (80291516)

Project Period (FY) 2001 – 2002
Project Status Completed (Fiscal Year 2002)
Budget Amount *help
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2002: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 2001: ¥2,600,000 (Direct Cost: ¥2,600,000)
KeywordsVASCULAR SMOOTH MUSCLE / VASCULAR ENDOTHELIAL CELL / CYCLIN-DEPENDENT KINASE INHIBITOR / ISOFORM / NUCLEAR LOCALIZATION SIGNAL / ATHEROSCLEROSIS / TRANSCRITION / 細胞周期 / サイクリン依存性キナーゼ / p27^<Kip1> / 蛋白質分解 / プロテアソーム / 血管平滑筋細胞
Research Abstract

A novel isoform of p27^<Kip1>, a cyclin-dependent kinase inhibitor, has been isolated from a cDNA library of porcine aortic endothelial cells. The N-terminal 162 amino acid of the new isoform was identical to those of p27^<Kip1>, while it contained a unique 18 amino acid C-terminus. The novel isoform was found to be resistant toward protease-mediated degradation, thus naming it p27^<Kip1R>, a degradation resistant sioform of p27^<Kip1>. The region 153-168 was determined to be necessary for its significant nuclear localization. However, this region contains only one basic amino acid, and an aliphatic amino acid was found to play a functional role in the nuclear localization signal. Namely, p27^<Kip1R> contains an atypical bipartite nuclear localization signal. Since functional substitution by an aliphatic amino acid was incomplete, p27^<Kip1R> also demonstrated a weak but significant localization in the cytosol, in addition to the strong nuclear localization.
Using the green fluorescence … More protein expression system, the effect of p27^<Kip1R> on the cell growth was investigated. It was found to strongly inhibit the cell growth of vascular smooth muscle cells and HeLa cells, as in the case with p27^<Kip1>. The growth inhibitory effect of both p27^<Kip1R> and p27^<Kip1> was completely abolished by removing the N-terminal region that bind to cyclin and cyclin-dependent kinase, while these truncated mutants demonstrated a significant nuclear localization. As a result, the mechanism for growth inhibition by p27^<Kip1R> was similar to that of p27^<Kip1>. The structural difference between two isforms was thus not linked to the growth inhibition.
The aorta of 6-week old spontaneously hypertensive rat (SHR) dominantly expressed p27^<Kip1>, while the aorta of normal rat (WKY) dominantly expressed p27^<Kip1R>. On the other hand, the aorta of 8 and 13-week old SHR dominantly expressed p27^<Kip1R>. When the aortic smooth muscle cells of normal rat were cultured, they dominanly expressed p27^<Kip1>. The expression of p27^<Kip1R> appeared to inversely correlate with proliferative state of the smooth muscle.
p27^<Kip1> was found to be upregulated by transcriptional upregulation in the vascular endothelial cells, when the cells formed a tight cell-to-cell contact. The promoter assay with a cloned Kip1 gene demonstrated an increase in the promoter activity upon formation of cell contract, thus suggesting that the promoter region contains a element that respond to cell contact. Less

Report

(3 results)
  • 2002 Annual Research Report   Final Research Report Summary
  • 2001 Annual Research Report
  • Research Products

    (18 results)

All Other

All Publications (18 results)

  • [Publications] Hirano K, Zeng Y, Hirano M, Nishismura J, Kanaide H: "Sequence requirement for nuclear localization and growth inhibition of p27^<Kip1R>, a degradation-resistant isoform of p27^<Kip1>"J Cell Biochem. (in press).

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Nakayama T, Hirano K, Shintani Y, Nishimura J, Nakatsuka A, Kuga H, Takahashi S, Kariaide H: "Unproductive cleavage and inactivation of protease-activated receptor-1 by trypsin in vascular endotheial cells"Br J Pharmacol. 138. 121-130 (2003)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Ihara E, Hirano K, Hirano M, Nishimura J, Nawata H, Kanaide H: "The mechanism of down-regulation of L-type Ca^<2+> channel in the proliferating smooth muscle cells of rat aorta"J Cell Biochem. 87. 242-251 (2002)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Hirano M, Hirano K, Nishimura J, Kanaide H: "Transcriptional up-regulation of p27^<Kip1> during contact-induced growth arrest in vascular endothelial cells"Exp Cell Res. 271. 356-367 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Hirano K, Hirano M, Zeng Y, Nishimura J, Hara K, Muta K, Nawata H, Kanaide H: "Cloning and functional expression of a degradation-resistant novel isoform of p27^<Kip1>"Biochem J. 353. 51-57 (2001)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Hirano K, Zeng Y, Hirano M, Nishimura J, Kanaide H:: "Sequence requirement for nuclear localization and growth inhibition of p27 p27^<Kip1R>, a degradation-resistant isoform of p27 p27^<Kip1>"J Cell Biochem (in press).

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Nakayama T, Hirano K, Shintani Y, Nishimura J, Nakatsuka A, Kuga H, Takahashi S, Kanaide H:: "Unproductive cleavage and inactivation of protease-activated receptor-1 by trypsin in vascular endotheial cells"Br J Pharmacol. 138. 121-130 (2003)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Ihara E, Hirano K, Hirano M, Nishimura J, Nawata H, Kanaide H:: "The mechanism of down-regulation of L-type Ca^<2+> channel in the proliferating smooth muscle cells of rat aorta"J Cell Biochem. 87. 242-251 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Hirano M, Hirano K, Nishimura J, Kanaide H:: "Transcriptional up-regulation of p27^<Kip1> during contact-induced growth arrest in vascular endothelial cells"Exp Cell Res. 271. 356-367 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Hirano K, Hirano M, Zeng Y, Nishimura J, Hara K, Muta K, Nawata H, Kanaide H:: "Cloning and functional expression of a degradation-resistant novel isoform of p27^<Kip1>"Biochem J. 353. 51-57 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2002 Final Research Report Summary
  • [Publications] Hirano K, Zeng Y, Hirano M, Nishismura J, Kanaide H: "Sequence requirement for nuclear localization and growth inhibition of p27^<Kip1R>, a degradation-resistant isoform of p27^<Kip1>"J Cell Biochem. (in press).

    • Related Report
      2002 Annual Research Report
  • [Publications] Nakayama T, Hirano K, Shintani Y, Nishimura J, Nakatsuka A, Kuga H, Takahashi S, Kanaide H: "Unproductive cleavage and inactivation of protease-activated receptor-1 by trypsin in vascular endotheial cells"Br J Pharmacol. 138. 121-130 (2003)

    • Related Report
      2002 Annual Research Report
  • [Publications] Takahashi R, Nishimura J, Hirano K, Naito S, Kanaide H: "The mechanisms for tachykinin-induced contractions of the rabbit corpus cavernosum"Br J Pharmacol. 137. 845-854 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Ihara E, Hirano K, Hirano M, Nishimura J, Nawata H, Kanaide H: "The mechanism of down-regulation of L-type Ca^<2+> in the proliferating smooth muscle cells of rat aorta"J Cell Biochem. 87. 242-251 (2002)

    • Related Report
      2002 Annual Research Report
  • [Publications] Hirano M, Hirano K, Nishimura J, Kanaide H: "Transcriptional Up-regulation of p27^<Kip1> during Contact-Induced Growth Arrest in Vascular Endothelial Cells"Experimental Cell Research. 271. 356-367 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Hirano K, Hirano M, Zeng Y, Hishimura J, Hara K, Muta K, Nawata H, Kanaide H: "Cloning and functional expression of a degradation-resistant novel isoform of p27^<Kip1>"Biochemical Journal. 353. 51-57 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Muranyi A, Zhang R, Liu F, Hirano K, (他3名): "Myotonic dystrophy protein kinase phosphorylates the myosin phosphatase targeting subunit and inhibits myosin phosphatase activity"FEBS Letter. 493. 80-84 (2001)

    • Related Report
      2001 Annual Research Report
  • [Publications] Ichiki T, Hirano K, Kanaide H, Takashita A (他5名): "Downregulation of angiotensin II type 1 receptor by hydrophobic 3-hydroxy-3-methylgiutaryl coenzyme A reductase inhibitors in vascular smooth muscle cells"Arteriosclerosis, Thrombosis and Vascular Biology. 21. 1896-1901 (2001)

    • Related Report
      2001 Annual Research Report

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Published: 2001-04-01   Modified: 2016-04-21  

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