RHO-associated protein kinase α potentiates insulin-induced MAP kinase activation in Xenopus oocytes

Abstract

We recently identified Xenopus Rho-associated protein kinase α (xROKα) as a Xenopus insulin receptor substrate-1 binding protein and demonstrated that the non-catalytic carboxyl terminus of xROKα binds Xenopus insulin receptor substrate-1 and blocks insulin-induced MAP kinase activation and germinal vesicle breakdown in Xenopus oocytes. In the current study we further examined the role of xROKα in insulin signal transduction in Xenopus oocytes. We demonstrate that injection of mRNA encoding the xROKα kinase domain or full length xROKα enhanced insulin-induced MAP kinase activation and germinal vesicle breakdown. In contrast, injection of a kinase-dead mutant of xROKα or pre-incubation of oocytes with an xROKα inhibitor significantly reduced insulin-induced MAP kinase activation. To further dissect the mechanism by which xROKα may participate in insulin signalling, we explored a potential function of xROKα in regulating cellular Ras function, since insulin-induced MAP kinase activation and germinal vesicle breakdown is known to be a Ras-dependent process. We demonstrate that whereas injection of mRNA encoding c-H-Ras alone induced xMAP kinase activation and GVBD in a very low percentage (about 10%) of injected oocytes, co-injection of mRNA encoding xROKα and c-H-Ras induced xMAP kinase activation and germinal vesicle breakdown in a significantly higher percentage (50-60%) of injected oocytes. These results suggest a novel function for xROKα in insulin signal transduction upstream of cellular Ras function.