Interactions of muscarinic receptors with the heterotrimeric G proteins Gq and G12: transduction of proliferative signals.
The proliferative and transforming properties of m2 and m5 muscarinic acetylcholine receptors and a series of wild-type, chimeric, and mutant G proteins were measured alone or in combination in NIH 3T3 cells to determine which G proteins mediate these signals and to what extent these signals can be influenced by changing the stoichiometry of receptors and G proteins. Responses were measured using the focus-forming assay and a novel assay called R-SAT (Receptor Selection and Amplification Technology) in which proliferative responses are monitored using a reporter gene. Individually, GTPase-deficient mutants (*) of G alpha q and G alpha 12, wild-type G alpha q, and m5 were active in R-SAT. G alpha 12* and m5 also induced focus formation. m2 was inactive in both assays. The ability of m5 to induce foci was significantly reduced by coexpression of G alpha q*. Synergistic effects of receptor/ G protein combinations were not observed in focus-forming assays but were readily detected by R-SAT. Coexpression of G alpha q with m5 induced constitutive activity in R-SAT and increased the potency of agonists at m5 by 90-fold. G alpha q also evoked agonist-dependent responses from m2 but not constitutive activity. Agonist potency was increased 10-fold at m2 and decreased 15-fold at m5 when these receptors were coexpressed with G alpha qi5, a chimeric G protein containing the five C-terminal residues of G alpha i2, compared with coexpression with G alpha q. Both G alpha q and G alpha qi5 had biphasic effects on the proliferative responses to m5 and m2, respectively, inhibiting responses at high agonist concentrations. Coexpression of G alpha 12 or G alpha 12i5 had no effect on the concentration-response relationships of m5, but both elicited weak responses from m2. We conclude that although G alpha 12 is a more potent oncogene, G alpha q transduces m5-driven cellular responses. The demonstrations that proliferative responses can be elicited from a nonmitogenic receptor by altering the type and concentration of available G proteins and that constitutive responses can be induced by G proteins imply that both the magnitude and type of receptor-initiated signal can be regulated at the level of G proteins in vivo.
Burstein
E S
ES
Department of Psychiatry, University of Vermont, Burlington, USA.
Bräuner-Osborne
H
H
Spalding
T A
TA
Conklin
B R
BR
Brann
M R
MR
D016475:3T3 Cells / Q000737:chemistry / Q000502:physiology; D000818:Animals; D002217:Carbachol / Q000494:pharmacology; D020558:GTP Phosphohydrolases / Q000235:genetics; D019204:GTP-Binding Proteins / Q000502:physiology*; D015870:Gene Expression / Q000187:drug effects / Q000502:physiology; D017930:Genes, Reporter; D051379:Mice; D018721:Muscarinic Agonists / Q000494:pharmacology; D011976:Receptors, Muscarinic / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology*
D018721:Muscarinic Agonists; D011976:Receptors, Muscarinic; D002217:Carbachol; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins