The non-catalytic domain of ras-GAP inhibits transformation induced by G protein coupled receptors.
We have studied the relationship between ras-GAP and G protein coupled receptors in a proliferative setting comprised of NIH3T3 expressing transfected muscarinic receptors (mAChRs). GAP expression plasmids were engineered to encode wild-type GAP, its carboxyl-terminal catalytic domain, a mutant lacking a portion of the catalytic domain, and an amino-terminal domain which contained the hydrophobic region as well as SH2-SH3 domains. Cotransfection of each GAP expression plasmid into NIH3T3 cells did not affect the transforming ability of the v-mos oncogene, but plasmids encoding wild-type GAP or the mutant consisting of an intact catalytic domain inhibited transformation induced by normal c-ras. Wild-type GAP also prevented transformation by m1 mAChRs, whereas the mutant consisting of only its catalytic domain lacked any demonstrable effect. In contrast, the N-terminal non-catalytic domain of GAP effectively prevented m1-induced focus-formation. Cell lines coexpressing m1 receptors and each of the GAP constructs revealed that GAP proteins do not affect m1 receptor density, receptor ligand binding characteristics or coupling to the PI-PLC signal transduction pathway. Thus, our findings suggest a role for the N-terminal non-catalytic domain of GAP in regulating biological functions mediated by G protein-coupled receptors.
Xu
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N
Molecular Signaling Unit, National Institute of Dental Research, National Institutes of Health, Bethesda, Maryland 20892.
McCormick
F
F
Gutkind
J S
JS
D016475:3T3 Cells; D000818:Animals; D002471:Cell Transformation, Neoplastic / Q000235:genetics*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D020690:GTPase-Activating Proteins; D006801:Humans; D051379:Mice; D011506:Proteins / Q000235:genetics / Q000502:physiology*; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics / Q000502:physiology*; D011976:Receptors, Muscarinic / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology; D014162:Transfection; D020703:ras GTPase-Activating Proteins
D020690:GTPase-Activating Proteins; D011506:Proteins; D011976:Receptors, Muscarinic; D020703:ras GTPase-Activating Proteins; D019204:GTP-Binding Proteins; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras)