The beta(3)-adrenergic receptor activates mitogen-activated protein kinasein adipocytes through a G(i)-dependent mechanism

Promiscuous coupling between G protein-coupled receptors and multiple speci es of heterotrimeric G; proteins provides a potential mechanism for expandi ng the diversity of G protein-coupled receptor signaling. We have examined the mechanism and functional consequences of dual G(s)/G(i) protein couplin g of the beta(3)-adrenergic receptor (beta(3)AR) in 3T3-F442A adipocytes. T he beta(3)AR selective agonist disodium (R,R)-5-[2[[2-(3-chlorophenyl)-2-hy droxyethyl]-amino]propyl]-1,3-benzodioxole-2,2-dicarboxylate (CL316,243) st imulated a dose-dependent increase in cAMP production in adipocyte plasma m embrane preparations, and pretreatment of cells with pertussis toxin result ed in a further a-fold increase in cAMP production by CL316,243, CL316,243 (5 mu M) stimulated the incorporation of 8-azido-[P-32]GTP into G alpha(s) (1.57 +/- 0.12; n = 3) and G alpha(i) (1.68 +/- 0.13; n = 4) in adipocyte p lasma membranes, directly demonstrating that beta(3)AR stimulation results in G(i)-GTP exchange. The beta(3)AR-stimulated increase in 8-azido-[P-32]GT P labeling of G alpha(i) was equivalent to that obtained with the A(1)-aden osine receptor agonist N-6-cyclopentyladenosine (1.56 +/- 0.07; n = 4), whe reas inclusion of unlabeled GTP (100 mu M) eliminated all binding. Stimulat ion of the beta(3)AR in 3T3-F442A adipocytes led to a 2-3-fold activation o f mitogen-activated protein (MAP) kinase, as measured by extracellular sign al-regulated kinase-1 and -2 (ERK1/2) phosphorylation. Pretreatment of cell s with pertussis toxin (PTX) eliminated MAP kinase activation by beta(3)AR, demonstrating that this response required receptor coupling to Gi, Express ion of the human beta(3)AR in HEK-293 cells reconstituted the PTX-sensitive stimulation of MAP kinase, demonstrating that this phenomenon is not exclu sive to adipocytes or to the rodent beta(3)AR, ERK1/2 activation by the bet a(3)AR was insensitive to the cAMP-dependent protein kinase inhibitor H-89 but was abolished by genistein and AG1478. These data indicate that constit utive beta(3)AR coupling to G(i) proteins serves both to restrain G(s)-medi ated activation of adenylyl cyclase and to initiate additional signal trans duction pathways, including the ERK1/2 MAP kinase cascade.