细胞黏附和胚胎发育对依赖C3G的Rap1激活的需求。
Requirement for C3G-dependent Rap1 activation for cell adhesion and embryogenesis.
作者信息
Ohba Y, Ikuta K, Ogura A, Matsuda J, Mochizuki N, Nagashima K, Kurokawa K, Mayer B J, Maki K, Miyazaki J, Matsuda M
机构信息
Department of Tumor Virology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan.
出版信息
EMBO J. 2001 Jul 2;20(13):3333-41. doi: 10.1093/emboj/20.13.3333.
C3G is a guanine nucleotide exchange factor (GEF) for Rap1, and is activated via Crk adaptor protein. To understand the physiological role of C3G, we generated C3G knockout mice. C3G(-/-) homozygous mice died before embryonic day 7.5. The lethality was rescued by the expression of the human C3G transgene, which could be excised upon the expression of Cre recombinase. From the embryo of this mouse, we prepared fibroblast cell lines, MEF-hC3G. Expression of Cre abolished the expression of C3G in MEF-hC3G and inhibited cell adhesion-induced activation of Rap1. The Cre-expressing MEF-hC3G showed impaired cell adhesion, delayed cell spreading and accelerated cell migration. The accelerated cell migration was suppressed by the expression of active Rap1, Rap2 and R-Ras. Expression of Epac and CalDAG-GEFI, GEFs for Rap1, also suppressed the accelerated migration of the C3G-deficient cells. This observation indicated that Rap1 activation was sufficient to complement the C3G deficiency. In conclusion, C3G-dependent activation of Rap1 is required for adhesion and spreading of embryonic fibroblasts and for the early embryogenesis of the mouse.
C3G是一种针对Rap1的鸟嘌呤核苷酸交换因子（GEF），并通过Crk衔接蛋白被激活。为了了解C3G的生理作用，我们构建了C3G基因敲除小鼠。C3G（-/-）纯合小鼠在胚胎第7.5天之前死亡。通过人C3G转基因的表达挽救了致死性，该转基因可在Cre重组酶表达时被切除。从这种小鼠的胚胎中，我们制备了成纤维细胞系，即MEF-hC3G。Cre的表达消除了MEF-hC3G中C3G的表达，并抑制了细胞黏附诱导的Rap1激活。表达Cre的MEF-hC3G表现出细胞黏附受损、细胞铺展延迟和细胞迁移加速。活性Rap1、Rap2和R-Ras的表达抑制了加速的细胞迁移。Rap1的GEF，即Epac和CalDAG-GEFI的表达也抑制了C3G缺陷细胞的加速迁移。这一观察结果表明，Rap1激活足以弥补C3G缺陷。总之，胚胎成纤维细胞的黏附和铺展以及小鼠的早期胚胎发育需要C3G依赖的Rap1激活。
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