Identification and characterication of novel proteins involved in epithelial morphogenesis

• Project/Area Number: 15390090
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: General medical chemistry
• Research Institution: Kyoto University
• Principal Investigator: FURUSE Mikio Kyoto University, associate professor, 医学研究科, 助教授 (90281089)
• Project Period (FY): 2003 – 2004
• Project Status: Completed (Fiscal Year 2004)
• Budget Amount: ¥14,200,000 (Direct Cost: ¥14,200,000); Fiscal Year 2004: ¥5,900,000 (Direct Cost: ¥5,900,000); Fiscal Year 2003: ¥8,300,000 (Direct Cost: ¥8,300,000)
• Keywords: epithelial cells / cell polarity / cell adhesion / cytoskeletons / GFP / membrane domains

Research Abstract

1.Improvement of the fluorescence localization-based retrovirus-mediated expression cloning for identification of novel proteins localized at epithelial cell structures.
We constructed cDNA libraries of cultured epithelial cell lines MDCK and CSG using a mouse retrovirus expression vector. As host epithelial cells for retrovirus infection, MDCK cells overexpressing the retrovirus receptor were. In small-scale screenings, we confirmed that claudins, occludin, JAM, etc. can be cloned in this system. During several trials, we improved the protocol for effective screening.
2.Identification of novel proteins concentrated in epithelial cell-cell junctions.
In the middle-scale screening using the CSG cells-derived cDNA library, homer2, abLIM3, and a novel coiled-coil protein have been identified as novel proteins localizing at epithelial cell-cell junctions. We produced antibodies against these proteins and analyzed their tissue expression pattern. We also determined the domains required for junction allocalization for each protein. Among them, alLIM3 is a novel subtype of abLIM, which is known to function in neuron orretin a by interacting with actin filaments. We found that abLIM3 is localized at cell-cell junctions in endothelial cells in lung and muscle tissues, and interact with actin filaments in vitro. Tissue expression pattern of abLIM3 suggest some roles of this protein in cell-cell junctions in endothelial cells that receive extension and shrinkage.

Research Products

• [Journal Article] JACOP, a novel plaque protein localizing at the apical junctional complex with sequence similarity to cingulin (2004)
  • Author(s): Ohnishi H, Nakahara T, Furuse K, Sasaki H, Tsukita Sh, Furuse M
  • Journal Title: J.Biol.Chem. 279 Pages: 46014-46022
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Establishment and characterization of cultured epithelial cells lacking expression of ZO-1 (2004)
  • Author(s): Umeda K, Matsui T, Nakayama M, Furuse K, Sasaki H, Furuse M, Tsukita Sh
  • Journal Title: J.Biol.Chem. 279 Pages: 44785-44794
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Expression patterns of claudins, tight junction adhesion molecules, in the inner ear (2004)
  • Author(s): Kitajiri S, Furuse M, Morita K, Saishin-Kiuchi Y, Kido H, Ito J, Tsukita Sh
  • Journal Title: Hear.Res 187 Pages: 25-34
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] JACOP, a novel plaque protein localizing at the apical junctional complex with sequence similarity to cingulin. (2004)
  • Author(s): Ohnishi, H., Nakahara, T., Furuse, K., Sasaki, H., Tsukita, S., Furuse, M.
  • Journal Title: J.Biol.Chem. 279 Pages: 46014-46022
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Establishment and characterization of cultured epithelial cells lacking expression of ZO-1. (2004)
  • Author(s): Umeda, K., Matsui, T., Nakayama, M., Furuse, K., Sasaki, H., Furuse, M., Tsukita, Sh.
  • Journal Title: J.Biol.Chem. 279 Pages: 44785-44794
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Expression patterns of claudins, tight junction adhesion molecules, in the inner ear. (2004)
  • Author(s): Kitajiri, S., Furuse, M., Morita, K., Saishin-Kiuchi, Y., Kido, H., Ito, J., Tsukita, Sh.
  • Journal Title: Hear.Res. 187 Pages: 25-34
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] JACOP, a novel plaque protein localizing at the apical junctional complex with sequence similarity to cinaulin (2004)
  • Author(s): Ohnishi H., Nakahara T., Furuse K., Sasaki H., Tsukita S., Furuse M
  • Journal Title: J.Biol.Chem 279 Pages: 46014-46022
• [Journal Article] Expression patterns of claudin family of tight junction membrane proteins in developipa mouse submandibular gland (2004)
  • Author(s): Hashizume A., Ueno T., Furuse M., Tsukita S., Nakanishi Y., Hieda Y.
  • Journal Title: Dev.Dyn. 231 Pages: 425-431
• [Journal Article] Establishment and characterization of cultured epithelial cells lacking expression of ZO-1 (2004)
  • Author(s): Umeda K., Matsui T., Nakayama M., Furuse K., Sasaki H., Furuse M., Tsukita S.
  • Journal Title: J.Biol.Chem. 43 Pages: 44785-44794
• [Journal Article] A peculiar internalization of claudins, tight junction-apecific adhesion molecules, during the intercellular movement of epithelial cells (2004)
  • Author(s): Matsuda M., Kubo A., Furuse M., Tsukita S.
  • Journal Title: J.Cell Sci. 117 Pages: 1247-1257
• [Journal Article] Expression and distribution of ZO-3, a tight junction MAGUK protein, in mouse tissues. (2003)
  • Author(s): Inoko A, Itoh M, Tamura A, Matsuda M, Furuse M, Tsukita Sh
  • Journal Title: Genes Cells 8 Pages: 837-845
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Size-selective loosening of the blood-brain barrier in claudin-5-deficient mice. (2003)
  • Author(s): Nitta T, Hata M, Gotoh S, Seo Y, Sasaki H, Hashimoto N, Furuse M, Tsukita Sh
  • Journal Title: J.Cell Biol. 161 Pages: 653-660
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Dynamic behavior of paired claudin strands within apposing plasma membranes (2003)
  • Author(s): Sasaki H, Matsui C, Furuse K, Mimori-Kiyosue Y, Furuse M, Tsukita Sh
  • Journal Title: PNAS 100 Pages: 3971-3976
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Expression and distribution of ZO-3, a tight junction MAGUK protein, in mouse tissues. (2003)
  • Author(s): Inoko, A., Itoh, M., Tamura, A., Matsuda, M., Furuse, M., Tsukita, Sh.
  • Journal Title: Genes Cells 8 Pages: 837-845
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Size-selective loosening of the blood-brain barrier in claudin-5-deficient mice. (2003)
  • Author(s): Nitta, T., Hata, M., Gotoh, S., Seo, Y., Sasaki, H., Hashimoto, N., Furuse, M., Tsukita, Sh.
  • Journal Title: J.Cell Biol. 161 Pages: 653-660
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Dynamic behavior of paired claudin strands within apposing plasma membranes. (2003)
  • Author(s): Sasaki, H., Matsui, C., Furuse, K., Mimori-Kiyosue, Y., Furuse, M., Tsukita, Sh.
  • Journal Title: PNAS 100 Pages: 3971-3976
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Sasaki et al.: "Dynamic behavior of paired claudin strands within apposing plasma membranes,"PNAS. 100. 3971-3976 (2003)
• [Publications] Nitta et al.: "Size-selective loosening of the blood-brain barrier in claudin-5-deficient mice"J.Cell.Biol.. 161. 653-660 (2003)
• [Publications] Inoko et al.: "Expression and distribution of ZO-3, a tight junction MAGUK protein, in mouse tissues"Genes Cells. 8. 837-845 (2003)
• [Publications] Kitajiri et al.: "Expression patterns of claudins, tight junction adhesion molecules, in the inner ear"Hear Res.. 187. 25-34 (2004)