Meller Julia, Vidali Luis, Schwartz Martin Alexander
Department of Microbiology, University of Virginia, Charlottesville, VA 22908, USA.
J Cell Sci. 2008 Jun 15;121(Pt 12):1981-9. doi: 10.1242/jcs.025130. Epub 2008 May 27.
Rac activation by integrins is essential for cell spreading, migration, growth and survival. Based mainly on overexpression of dominant-negative mutants, RhoG has been proposed to mediate integrin-dependent Rac activation upstream of ELMO and Dock180. RhoG-knockout mice, however, display no significant developmental or functional abnormalities. To clarify the role of RhoG in integrin-mediated signaling, we developed a RhoG-specific antibody, which, together with shRNA-mediated knockdown, allowed analysis of the endogenous protein. Despite dramatic effects of dominant-negative constructs, nearly complete RhoG depletion did not substantially inhibit cell adhesion, spreading, migration or Rac activation. Additionally, RhoG was not detectably activated by adhesion to fibronectin. Using Rac1(-/-) cells, we found that constitutively active RhoG induced membrane ruffling via both Rac-dependent and -independent pathways. Additionally, endogenous RhoG was important for Rac-independent cell migration. However, RhoG did not significantly contribute to cell spreading even in these cells. These data therefore clarify the role of RhoG in integrin signaling and cell motility.
整合素激活Rac对细胞铺展、迁移、生长及存活至关重要。主要基于显性负性突变体的过表达，有人提出RhoG在ELMO和Dock180上游介导整合素依赖性Rac激活。然而，RhoG基因敲除小鼠未表现出明显的发育或功能异常。为阐明RhoG在整合素介导信号传导中的作用，我们制备了一种RhoG特异性抗体，该抗体与shRNA介导的敲低技术一起，可用于分析内源性蛋白。尽管显性负性构建体有显著作用，但几乎完全耗尽RhoG并未实质性抑制细胞黏附、铺展、迁移或Rac激活。此外，黏附于纤连蛋白并未检测到RhoG被激活。利用Rac1(-/-)细胞，我们发现组成型激活的RhoG通过Rac依赖性和非依赖性途径诱导膜皱褶形成。此外，内源性RhoG对不依赖Rac的细胞迁移很重要。然而，即使在这些细胞中，RhoG对细胞铺展的作用也不显著。因此，这些数据阐明了RhoG在整合素信号传导和细胞运动中的作用。
