Inactivation of raf-1 by a protein-tyrosine phosphatase stimulated by GTP and reconstituted by Galphai/o subunits.
A membrane-associated form of Raf-1 in v-Ras transformed NIH 3T3 cells can be inactivated by protein phosphatases regulated by GTP. Herein, a distinct protein-tyrosine phosphatase (PTPase) in membrane preparations from v-Ras transformed NIH 3T3 cells was found to be activated by guanyl-5'-yl imidodiphosphate (GMPPNP) and was identified as an effector for pertussis toxin (PTx)-sensitive G-protein alpha subunits. PTPase activation was blocked by prior treatment of cells with PTx. PTPase activation by GTP, but not GMPPNP, was transient. A GMPPNP-stimulated PTPase (PTPase-G) co-purified with Galphai/o subunits during Superose 6 and Mono Q chromatography. PTPase-G activity in Superose 6 fractions from GDP-treated membranes was reconstituted by activated Galphai/o, but not G beta gamma, subunits. PTPase-G may contribute to GMPPNP-stimulated inactivation of Raf-1 in v-Ras cell membranes because Raf-1 inactivation was PTx-sensitive and PTPase-G inactivated exogenous Raf-1.
Dent
P
P
Howard Hughes Medical Institute, the Markey Center for Cell Signaling, University of Virginia, Charlottesville, Virginia 22908, USA.
Reardon
D B
DB
Wood
S L
SL
Lindorfer
M A
MA
Graber
S G
SG
Garrison
J C
JC
Brautigan
D L
DL
Sturgill
T W
TW
D016475:3T3 Cells; D000818:Animals; D002461:Cell Line, Transformed; D002462:Cell Membrane / Q000378:metabolism; D002850:Chromatography, Gel; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D011905:Genes, ras; D006153:Guanosine Diphosphate / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000494:pharmacology*; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D007700:Kinetics; D051379:Mice; D037342:Pertussis Toxin; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D019908:Proto-Oncogene Proteins c-raf; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D011518:Proto-Oncogene Proteins; D010566:Virulence Factors, Bordetella; D006153:Guanosine Diphosphate; D006165:Guanylyl Imidodiphosphate; D006160:Guanosine Triphosphate; D037342:Pertussis Toxin; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; D017027:Protein Tyrosine Phosphatases; D019204:GTP-Binding Proteins