Reconstitution of p21ras-dependent and -independent mitogen-activated protein kinase activation in a cell-free system.
In Xenopus oocytes, mitogen-activated protein (MAP) kinase can be activated by progesterone treatment or by microinjection of cyclin A, both of which lead to activation of the cdc2 protein kinase. The tyrosine kinase pp60v-src has previously been shown to accelerate progesterone-induced oocyte maturation and to increase the phosphorylation of ribosomal protein S6 by pp90rsk, most likely by activating MAP kinase. In extracts of resting oocytes, MAP kinase kinase and MAP kinase were activated by addition of pp60v-src or cyclin A. Activation by pp60v-src was blocked by a dominant-negative p21ras protein (RAST), but activation by cyclin A/cdc2 was unaffected. Thus these two pathways that converge at MAP kinase kinase but are clearly divergent upstream of a p21ras-dependent step can be studied in a cell-free system.
Howard Hughes Medical Institute, University of Colorado School of Medicine, Denver 80262.
B
B
VanRenterghem
J B
JB
Gibbs
J L
JL
Maller
D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002474:Cell-Free System; D016213:Cyclins / Q000378:metabolism; D004789:Enzyme Activation; D005260:Female; D007700:Kinetics; D016200:Maturation-Promoting Factor / Q000378:metabolism; D020929:Mitogen-Activated Protein Kinase Kinases; D015688:Oncogene Protein pp60(v-src) / Q000378:metabolism; D009865:Oocytes / Q000187:drug effects / Q000201:enzymology; D011374:Progesterone / Q000494:pharmacology; D011494:Protein Kinases / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins / Q000378:metabolism; D014162:Transfection; D014982:Xenopus laevis
D016213:Cyclins; D011994:Recombinant Proteins; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins; D011374:Progesterone; D011494:Protein Kinases; D015688:Oncogene Protein pp60(v-src); D017871:Calcium-Calmodulin-Dependent Protein Kinases; D016200:Maturation-Promoting Factor; D020929:Mitogen-Activated Protein Kinase Kinases; D016283:Proto-Oncogene Proteins p21(ras)