| Project/Area Number |
13670115
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General medical chemistry
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| Research Institution | RIKEN |
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Principal Investigator |
YONEMURA Shigenobu RIKEN Center for Developmental Biology, Laboratory for Cellular Morphogenesis, Team Leader, 細胞形態形成研究チーム, チームリーダー (60192811)
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| Project Period (FY) |
2001 – 2002
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| Project Status |
Completed (Fiscal Year 2002)
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| Budget Amount *help |
¥4,100,000 (Direct Cost: ¥4,100,000)
Fiscal Year 2002: ¥1,600,000 (Direct Cost: ¥1,600,000)
Fiscal Year 2001: ¥2,500,000 (Direct Cost: ¥2,500,000)
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| Keywords | Rho / Microvilli / TCA / LPA / EGF / cleavage furrow / 可視化 / 低分子量Gタンパク質 / 抗体 / 局在 / TCA固定 |
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| Research Abstract |
We screened combinations of anti-Rho antibodies and fixation protocols for precise localization of Rho within cells. Several commercial antibodies and antibodies produced by ourselves were found to be reliable with appropriate fixation protocols such as TCA fixation. For this screening, we doubly stained Myc-RhoA-expressing cultured cells with an anti-myc antibody and an anti-Rho antibody. Reliable anti-Rbo antibodies should stain the cells as anti-Myc antibody does. Using valid antibodies, we localized Rho within cells and tissues. In cultured epithelial cells, Rho was accumulated on lateral membranes. In fibroblastic cells, Rho was distributed almost evenly throughout the cytoplasm. During cytokinesis, Rho was generally concentrated at the cleavage furrow. In tissues, although Rho concentration at microvilli was obvious, Rho localization within cells differed depending on cell type, probably reflecting the functional difference of the cells in tissues. Biochemical data indicated that Rho translocates from the cytoplasm to the plasma membrane on its activation. We visualized this translocation for the first time using cultured cells stimulated with LPA or EGF. On stimulatioh with these factors, Rho was rapidly recruited from the cytoplasm to the plasma membrane within 30 seconds.
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