Study on the physiological function of a cell polarity protein, PAR-1 knase, for polarization process of mammalian epithelial cells.

• Project/Area Number: 15570163
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Cell biology
• Research Institution: Yokohama City University, Graduate School of Medical Science
• Principal Investigator: SUZUKI Atsushi Yokohama City University, Graduate School of Medical Science, Department of Molecular Cellular Biology, Lecturer, 大学院・医学研究科, 講師 (00264606)
• Project Period (FY): 2003 – 2004
• Project Status: Completed (Fiscal Year 2004)
• Budget Amount: ¥3,700,000 (Direct Cost: ¥3,700,000); Fiscal Year 2004: ¥1,800,000 (Direct Cost: ¥1,800,000); Fiscal Year 2003: ¥1,900,000 (Direct Cost: ¥1,900,000)
• Keywords: PAR / Epithelial cells / Cell polarity / PKC / プロテインキナーゼC

Research Abstract

aPKC and PAR-1 are required for cell polarity in various contexts. In mammalian epithelial cells, aPKC localizes at tight junctions (TJs) and plays an indispensable role in the development of asymmetric intracellular junctions essential for the establishment and the maintenance of apicobasal polarity. On the other hand, one of the mammalian PAR-1 kinases, PAR-1b/EMK1/MARK2, localizes to the lateral membrane in a complimentary manner with aPKC, but little is known about its role in apicobasal polarity of epithelial cells as well as its functional relationship with aPKC.
We demonstrate that PAR-1b is essential for the asymmetric development of membrane domains of polarized MDCK cells. Nonetheless, it is not required for the junctional localization of aPKC nor the formation of TJs, suggesting that PAR-1b works downstream of aPKC during epithelial cell polarization. On the other hand, we revealed that aPKC induces the PAR-1b binding with 14-3-3/PAR-5 by phosphorylates threonine 595 of PAR-1b. In polarized MDCK cells, T595 phosphorylation and 14-3-3 binding are only observed in the soluble form of PAR-1b, and okadaic acid treatment induces T595-dependent dissociation of PAR-1b from the lateral membrane. Furthermore, T595A mutation induces not only PAR-1b leakage into the apical membrane, but also abnormal development of membrane domains. These results suggest that in polarized epithelial cells aPKC phosphorylates PAR-1b at TJs, and in cooperation with 14-3-3, promotes the dissociation of PAR-1b from the lateral membrane to regulate PAR-1b activity for the membrane domain development.
These results suggest that mammalian aPKC functions upstream of PAR-1b both in the establishment and maintenance of epithelial cell polarity.

Research Products

• [Journal Article] aPKC acts upstream of PAR-1b in both the establishment and maintenance of mammalian epithelial polarity. (2004)
  • Author(s): Suzuki et al.
  • Journal Title: Curr.Biol. 14 Pages: 539-551
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Affixin interacts with a-actinin and mediates integrin signaling for reorganization of F-actin induced by initial cell-substrate interaction. (2004)
  • Author(s): Yamaji et al.
  • Journal Title: J.Cell Biol. 165 Pages: 539-551
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] The first CH domain of affixin activates Cdc42 and Rac1 through αPIX, a Cdc42/Rac1-specific guanine nucleotide exchanging factor. (2004)
  • Author(s): Mishima W. et al.
  • Journal Title: Genes to Cells 9 Pages: 193-204
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Junctional adhesion molecules (JAMs) : more molecules with dual functions? (2004)
  • Author(s): Ebnet K. et al.
  • Journal Title: J.Cell Science 117 Pages: 19-23
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] aPKC acts upstream of PAR-1b in both the establishment and maintenance of mammalian epithelial polarity (2004)
  • Author(s): Suzuki, A., Hirata, M, Kamimura, K., Maniwa, R., Yamanaka, T., Mizuno, K., Kishikawa, M., Hirose, H., Amano, Y., Izumi, N., Miwa, Y., Ohno, S.
  • Journal Title: Curr. Biol. 14 Pages: 1425-1435
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Affixin interacts with a-actinin and mediates integrin signaling for reorganization of F-actin induced by initial cell-substrate interaction (2004)
  • Author(s): Yamaji, S., Suzuki, A., Kanamori, H., Mishima, W., Yoshimi, R., Takasaki, H., Takabayashi, M., Fujimaki, K., Fujisawa, S., Ohno, S., Ishigatsubo, Y.
  • Journal Title: J. Cell. Biol. 165 Pages: 539-551
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] The first CH domain of affixin activates Cdc42 and Rac1 through aPIX, a Cdc42/Rac1-specific guanine nucleotide exchanging factor (2004)
  • Author(s): Mishima, W., Suzuki, A., Yamaji, S., Yoshimi, R., Ueda, A., Kaneko, T., Tanaka, J., Miwa, Y., Ohno, S., Ishigatsubo, Y.
  • Journal Title: Genes to Cells 9 Pages: 193-204
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Junctional adhesion molecules (JAMs): more molecules with dual functions? (2004)
  • Author(s): Ebnet, K., Suzuki, A., Ohno, S., Vestweber, D.
  • Journal Title: J. Cell Sci. 117 Pages: 19-29
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] The junctional adhesion molecule (JAM) family members JAM-2 and JAM-3 associate with the cell polarity protein PAR-3 : a possible role for JAMs in endothelial cell polarity. (2003)
  • Author(s): Ebnet K. et al.
  • Journal Title: J.Cell Science 116 Pages: 3879-3891
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Mammalian Lgl Forms a Protein Complex with PAR-6 and aPKC Independently of PAR-3 to Regulate Epithelial Cell Polarity. (2003)
  • Author(s): Yamanaka T. et al.
  • Journal Title: Curr.Biol. 13 Pages: 734-743
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Self-association of PAR-3 mediated by the conserved N-terminal domain contributes to the development of epithelial tight junction. (2003)
  • Author(s): Mizuno et al.
  • Journal Title: J.Biol.Chem. 278 Pages: 31240-31250
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] A PKC Isotype Essential for the Development of Multicellular Organisms. (2003)
  • Author(s): Suzuki A. et al.
  • Journal Title: J.Biochem. 133 Pages: 9-16
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] The junctional adhesion molecule (JAM) family members JAM-2 and JAM-3 associate with the cell polarity protein PAR-3: a possible role for JAMs in endothelial cell polarity (2003)
  • Author(s): Ebnet, K., Aurrand-Lions, M., Kuhn, A., Kiefer, F., Butz, S., Zander, K., Brickwedde, M.K., Suzuki, A., Imhof, B.A., Vestweber, D.
  • Journal Title: J. Cell Sci. 116 Pages: 3879-91
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Self-association of PAR-3 mediated by the conserved N-terminal domain contributes to the development of epithelial tight junction (2003)
  • Author(s): Mizuno, K., Suzuki, A., Hirose, T., Kitamura, K., Kutsuzawa, Y., Futaki, M., Amano, Y., Ohno, S.
  • Journal Title: J. Biol. Chem. 278 Pages: 31240-31250
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Mammalian Lgl Forms a Protein Complex with PAR-6 and aPKC Independently of PAR-3 to Regulate Epithelial Cell Polarity (2003)
  • Author(s): Yamanaka, T., Horikoshi, Y., Sugiyama, Y., Ishiyama, C., Suzuki, A., Hirose, T., Iwamatsu, A., Shinohara, A., Ohno, S.
  • Journal Title: Curr Biol 13 Pages: 734-743
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Protein kinase C I/i: A PKC Isotype Essential for the Development of Multicellular Organisms (2003)
  • Author(s): Suzuki, A., Akimoto, K.S., Ohno, S.
  • Journal Title: J. Biochem. 133 Pages: 9-16
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Mishima W. et al.: "The first CH domain of affixin activates Cdc42 and Rac1 through αPIX, a Cdc42/Rac1-specific guanine nucleotide exchanging factor"Genes to Cells. (In press). (2004)
• [Publications] Ebnet K. et al.: "Junctional adhesion molecules (JAMs) : more molecules with dual functions?"J.Cell Science. 117. 19-23 (2003)
• [Publications] Ebnet K. et al.: "The junctional adhesion molecule (JAM) family members JAM-2 and JAM-3 associate with the cell polarity protein PAR-3 a possible role for..."J.Cell Science. 116. 3879-3891 (2003)
• [Publications] Yamanaka T. et al.: "Mammalian Lg1 Forms a Protein Complex with PAR-6 and aPKC Independently of PAR-3 to Regulate Epithelial Cell Polarity"Curr.Biol.. 13. 734-743 (2003)
• [Publications] Mizuno et al.: "Self-association of PAR-3 mediated by the conserved N-terminal domain contributes to the development of epithelial tight junction"J.Biol.Chem.. 278. 31240-31250 (2003)
• [Publications] Suzuki A. et al.: "A PKC Isotype Essential for the Development of Multicellular Organisms"J.Biochem.. 133. 9-16 (2003)