Palmitoylation of Ha-Ras facilitates membrane binding, activation of downstream effectors, and meiotic maturation in Xenopus oocytes.
Ras proteins serve as critical relays in signal transduction pathways that control growth and differentiation and must undergo posttranslational modifications before they become functional. While it is established that farnesylation is necessary for membrane binding and cellular functions of all Ras proteins, the significance of palmitoylation is unclear. We have studied the contribution of Ha-Ras palmitoylation for biological activity in Xenopus oocytes. In contrast to wild-type Ha-Ras, which binds to membranes and induces meiosis when microinjected into oocytes, a nonpalmitoylated but farnesylated and methylated mutant mislocalizes to the cytosol and fails to promote maturation. This lack of responsiveness correlates with the inability of the mutant to induce phosphorylation and activation of mitogen-activated protein kinase and maturation promoting factor, which are both strongly activated by wild-type Ha-Ras. Costimulation of oocytes with insulin increases their responsiveness to Ras and partially rescues the biological activity of the palmitoylation-resistant mutant. However, 25-50 times higher doses of mutant were required to elicit responses equivalent to wild-type Ha-Ras. These results suggest that palmitoylation and membrane association of Ha-Ras is necessary for efficient activation of the mitogen-activated protein kinase cascade in vivo and are consistent with a biochemical function for Ras as a membrane targeting signal for downstream effectors in this pathway.
Dudler
T
T
Department of Chemistry, University of Washington, Seattle 98195-1700, USA.
Gelb
M H
MH
D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D005260:Female; D018698:Glutamic Acid; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000494:pharmacology; D007700:Kinetics; D008540:Meiosis; D009865:Oocytes / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D019308:Palmitic Acid; D010169:Palmitic Acids / Q000378:metabolism*; D011374:Progesterone / Q000494:pharmacology; D017368:Protein Prenylation; D011499:Protein Processing, Post-Translational; D016324:Sequence Tagged Sites; D014982:Xenopus laevis; D018631:ras Proteins / Q000096:biosynthesis / Q000378:metabolism*
D007328:Insulin; D010169:Palmitic Acids; D019308:Palmitic Acid; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D018698:Glutamic Acid; D011374:Progesterone; D006160:Guanosine Triphosphate; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D018631:ras Proteins