RHO-associated protein kinase alpha potentiates insulin-induced MAP kinase activation in Xenopus oocytes.
We recently identified Xenopus Rho-associated protein kinase alpha (xROKalpha) as a Xenopus insulin receptor substrate-1 binding protein and demonstrated that the non-catalytic carboxyl terminus of xROKalpha 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 xROKalpha in insulin signal transduction in Xenopus oocytes. We demonstrate that injection of mRNA encoding the xROKalpha kinase domain or full length xROKalpha enhanced insulin-induced MAP kinase activation and germinal vesicle breakdown. In contrast, injection of a kinase-dead mutant of xROKalpha or pre-incubation of oocytes with an xROKalpha inhibitor significantly reduced insulin-induced MAP kinase activation. To further dissect the mechanism by which xROKalpha may participate in insulin signalling, we explored a potential function of xROKalpha 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 xROKalpha 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 xROKalpha in insulin signal transduction upstream of cellular Ras function.
Ohan
N
N
Loeb Health Research Institute, Ottawa Hospital, Department of Biochemistry, Microbiology & Immunology, University of Ottawa, Ottawa, K1Y 4E9, Canada.
Agazie
Y
Y
Cummings
C
C
Booth
R
R
Bayaa
M
M
Liu
X J
XJ
D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D066298:In Vitro Techniques; D007328:Insulin / Q000494:pharmacology*; D047908:Intracellular Signaling Peptides and Proteins; D009154:Mutation; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism; D010858:Pigmentation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000008:administration & dosage / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D014981:Xenopus; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism; D054460:rho-Associated Kinases
D007328:Insulin; D047908:Intracellular Signaling Peptides and Proteins; D012333:RNA, Messenger; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D018631:ras Proteins