Heterotrimeric G proteins containing G alpha i3 regulate multiple effector enzymes in the same cell. Activation of phospholipases C and A2 and inhibition of adenylyl cyclase.
Previous studies have shown that a single type of transmembrane receptor is able to regulate multiple effectors through the activation of heterotrimeric G proteins. For example, the m2 muscarinic acetylcholine receptor (mAChR) expressed in Chinese hamster ovary (CHO) cells inhibits adenylyl cyclase, stimulates phospholipase C-dependent intracellular Ca2+ release, and activates phospholipase A2 through pertussis toxin-sensitive G proteins. However, it is unclear whether multiple effector enzymes can be regulated by one type of heterotrimeric G protein within a single cell. To investigate this question, we constructed a derivative of G alpha i3 (termed G alpha i3 C > S) in which the carboxyl-terminal cysteine residue, the site for pertussis toxin modification, was changed to a serine. Following pertussis toxin treatment of transfected CHO cells expressing the m2 mAChR, we found that the G alpha i3 C > S protein underwent guanine nucleotide exchange in response to the muscarinic agonist carbachol, while the m2 mAChR failed to activate the endogenous G alpha i2 and G alpha i3 proteins. Moreover, coupling of heterotrimeric G proteins containing G alpha i3 C > S to the m2 mAChR resulted in pertussis toxin-resistant inhibition of adenylyl cyclase, stimulation of phospholipase C-induced intracellular Ca2+ release, and phospholipase A2-mediated arachidonic acid release. Therefore, these studies provide conclusive evidence that heterotrimeric G proteins containing just G alpha i3 can regulate multiple effector enzymes within the same cell type.
Department of Biochemistry and Molecular Biology, Harvard University, Cambridge, Massachusetts 02138.
T W
TW
Hunt
R C
RC
Carroll
E G
EG
Peralta
D037361:Adenylate Cyclase Toxin; D000067956:Adenylyl Cyclase Inhibitors; D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism; D006224:Cricetinae; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D037342:Pertussis Toxin; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D011976:Receptors, Muscarinic / Q000502:physiology; D015398:Signal Transduction; D010738:Type C Phospholipases / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D037361:Adenylate Cyclase Toxin; D000067956:Adenylyl Cyclase Inhibitors; D011976:Receptors, Muscarinic; D010566:Virulence Factors, Bordetella; D037342:Pertussis Toxin; D010741:Phospholipases A; D054467:Phospholipases A2; D010738:Type C Phospholipases; D019204:GTP-Binding Proteins; D002118:Calcium