Oncogenic ras triggers the activation of 42-kDa mitogen-activated protein kinase in extracts of quiescent Xenopus oocytes.
Quiescent, full-grown Xenopus oocytes, which are arrested at the G2/M border of meiosis, contain an inactive 42-kDa mitogen-activated protein kinase (p42MAPK) that is activated when oocytes are stimulated to resume the meiotic cell cycle. We have made extracts from these oocytes that respond to four cell cycle activators: oncogenic [Val12]Ras protein, clam cyclins A delta 60 and B delta 97, and the phosphatase inhibitor okadaic acid. All four induce the tyrosine phosphorylation and activation of p42MAPK. Both cyclins and okadaic acid, but not [Val12]Ras, also lead to activation of the endogenous cyclin B/cdc2 kinase complexes in extracts of quiescent oocytes. Using extracts prepared from cycloheximide-arrested interphase cells, we show that although p42MAPK activation can occur in response to cyclin-activated cdc2, the Ras-induced activation of p42MAPK occurs without intervening cdc2 activation. Neither the nononcogenic [Gly12]Ras nor [Val12,Arg186]Ras, a mutant that lacks the C-terminal consensus sequence directing prenylation and subsequent membrane association, is an effective activator of p42MAPK in vitro.
Shibuya
E K
EK
Department of Anatomy and Cell Biology, Harvard Medical School, Boston, MA 02115.
Polverino
A J
AJ
Chang
E
E
Wigler
M
M
Ruderman
J V
JV
D000818:Animals; D001483:Base Sequence; D016213:Cyclins / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology; D004789:Enzyme Activation; D008540:Meiosis / Q000187:drug effects*; D008845:Microinjections; D019950:Mitogen-Activated Protein Kinase 1; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D009865:Oocytes / Q000201:enzymology*; D019000:Phosphotyrosine; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics / Q000494:pharmacology*; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism; D014982:Xenopus laevis / Q000196:embryology*
D016213:Cyclins; D009838:Oligodeoxyribonucleotides; D011993:Recombinant Fusion Proteins; D019000:Phosphotyrosine; D014443:Tyrosine; D003513:Cycloheximide; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D019950:Mitogen-Activated Protein Kinase 1; D016283:Proto-Oncogene Proteins p21(ras)