Walsh A B, Bar-Sagi D
Department of Molecular Genetics and Microbiology and the Graduate Program in Physiology and Biophysics, State University of New York, Stony Brook, New York 11794, USA.
J Biol Chem. 2001 May 11;276(19):15609-15. doi: 10.1074/jbc.M010573200. Epub 2001 Feb 14.
Ras proteins are key regulators of cell growth and differentiation. Mammalian cells express three closely related Ras proteins: Ha-Ras, K-Ras, and N-Ras. We have compared the abilities of the Ha-Ras and K-Ras isoforms to activate the Rac effector pathway, using three Rac-dependent readouts: induction of membrane ruffling and pinocytosis, stimulation of cell motility, and Pak binding. The total surface area of membrane ruffles induced by K-RasV12 was 2-fold greater than that induced by Ha-RasV12. Likewise, the number of K-RasV12-induced pinocytic vesicles per cell was approximately 2-fold greater than that induced by Ha-RasV12. In a wound healing assay, K-RasV12-injected cells migrated twice as fast as Ha-RasV12-injected cells. Moreover, the Pak binding activity of Rac, which is indicative of the amount of GTP-bound Rac, was higher in K-RasV12-expressing cells than Ha-RasV12-expressing cells. These results suggest that K-Ras activates Rac more efficiently than Ha-Ras. The preferential activation of Rac by K-Ras is dependent on the mode of membrane anchoring and impacts on the ability of K-Ras to regulate cell survival.
Ras蛋白是细胞生长和分化的关键调节因子。哺乳动物细胞表达三种密切相关的Ras蛋白：Ha-Ras、K-Ras和N-Ras。我们使用三种Rac依赖性读数比较了Ha-Ras和K-Ras同工型激活Rac效应途径的能力：诱导膜皱襞和胞饮作用、刺激细胞运动以及Pak结合。由K-RasV12诱导的膜皱襞的总表面积比由Ha-RasV12诱导的大2倍。同样，每个细胞中由K-RasV12诱导的胞饮小泡数量比由Ha-RasV12诱导的大约多2倍。在伤口愈合试验中，注射K-RasV12的细胞迁移速度是注射Ha-RasV12的细胞的两倍。此外，Rac的Pak结合活性（指示结合GTP的Rac的量）在表达K-RasV12的细胞中比表达Ha-RasV12的细胞中更高。这些结果表明，K-Ras比Ha-Ras更有效地激活Rac。K-Ras对Rac的优先激活取决于膜锚定方式，并影响K-Ras调节细胞存活的能力。
