Takaishi K, Kikuchi A, Kuroda S, Kotani K, Sasaki T, Takai Y
Department of Biochemistry, Kobe University School of Medicine, Japan.
Mol Cell Biol. 1993 Jan;13(1):72-9. doi: 10.1128/mcb.13.1.72-79.1993.
Evidence is accumulating that rho p21, a ras p21-related small GTP-binding protein (G protein), regulates the actomyosin system. The actomyosin system is known to be essential for cell motility. In the present study, we examined the action of rho p21, its inhibitory GDP/GTP exchange protein (named rho GDI), its stimulatory GDP/GTP exchange protein (named smg GDS), and Clostridium botulinum ADP-ribosyltransferase C3, known to selectively ADP-ribosylate rho p21 and to impair its function, in cell motility (chemokinesis) of Swiss 3T3 cells. We quantitated the capacity of cell motility by measuring cell tracks by phagokinesis. Microinjection of the GTP gamma S-bound active form of rhoA p21 or smg GDS into Swiss 3T3 cells did not affect cell motility, but microinjection of rho GDI into the cells did inhibit cell motility. This rho GDI action was prevented by comicroinjection of rho GDI with the GTP gamma S-bound form of rhoA p21 but not with the same form of rhoA p21 lacking the C-terminal three amino acids which was not posttranslationally modified with lipids. The rho GDI action was not prevented by Ki-rasVal-12 p21 or any of the GTP gamma S-bound form of other small GTP-binding proteins including rac1 p21, G25K, and smg p21B. Among these small G proteins, rhoA p21, rac1 p21, and G25K are known to be substrates for rho GDI. The rho GDI action was not prevented by comicroinjection of rho GDI with smg GDS. Microinjection of C3 into Swiss 3T3 cells also inhibited cell motility. These results indicate that the rho GDI-rho p21 system regulates cell motility, presumably through the actomyosin system.
越来越多的证据表明，rho p21，一种与ras p21相关的小GTP结合蛋白（G蛋白），调节肌动球蛋白系统。已知肌动球蛋白系统对细胞运动至关重要。在本研究中，我们检测了rho p21、其抑制性GDP/GTP交换蛋白（称为rho GDI）、其刺激性GDP/GTP交换蛋白（称为smg GDS）以及肉毒杆菌ADP核糖基转移酶C3在瑞士3T3细胞运动（趋化运动）中的作用，已知后者可选择性地将ADP核糖基化到rho p21上并损害其功能。我们通过吞噬运动测量细胞轨迹来定量细胞运动能力。将GTPγS结合的活性形式的rhoA p21或smg GDS显微注射到瑞士3T3细胞中不影响细胞运动，但将rho GDI显微注射到细胞中确实会抑制细胞运动。这种rho GDI的作用可通过将rho GDI与GTPγS结合形式的rhoA p21共同显微注射来阻止，但与缺乏C末端三个氨基酸且未进行脂质翻译后修饰的相同形式的rhoA p21共同显微注射则不能阻止。rho GDI的作用不能被Ki-rasVal-12 p21或其他小GTP结合蛋白的任何GTPγS结合形式（包括rac1 p21、G25K和smg p21B）阻止。在这些小G蛋白中，rhoA p21、rac1 p21和G25K已知是rho GDI的底物。rho GDI与smg GDS共同显微注射不能阻止rho GDI的作用。将C3显微注射到瑞士3T3细胞中也会抑制细胞运动。这些结果表明，rho GDI-rho p21系统可能通过肌动球蛋白系统调节细胞运动。
