Monovalent anions differentially modulate coupling of the beta2-adrenoceptor to G(s)alpha splice variants.

The beta2-adrenoceptor (beta2AR) fused to the long splice variant of G(s)alpha (G(s)alphaL), but not the beta2AR fused to the short splice variant of G(s)alpha (G(s)alphaS) shows the hallmarks of high constitutive activity, i.e., strong activation of adenylyl cyclase (AC) by GTP and strong inhibition of AC by inverse agonist. These coupling differences are the result of differences in GDP affinity of G(s)alpha splice variants. The aim of this study was to identify experimental variables that differentially affect beta2AR coupling to G(s)alphaS and G(s)alphaL. NaCl substantially reduced agonist-independent AC activation by GTP and inverse agonist inhibition and enhanced agonist stimulation of AC in Sf9 insect cell membranes expressing the beta2AR-G(s)alphaL fusion protein. Salts reduced inverse agonist inhibition and increased agonist stimulation of AC in the order of efficiency NaI approximately KI > NaBr approximately KBr > NaCl approximately LiCl approximately KCl approximately RbCl approximately CsCl approximately choline chloride, indicating that monovalent anions determine salt effects. Salts inhibited guanosine 5'-O-(3-thiotriphosphate)-mediated AC activation by G(salphaL) without beta2AR in the order of efficiency NaI > NaBr > NaCl. NaCl enhanced the affinity of G(s)alphaL for GDP. Salts had much smaller effects on beta2AR ligand regulation of AC in membranes expressing beta2AR-G(s)alphaS than in membranes expressing beta2AR-G(s)alphaL. These data are explained by a model in which anions increase the GDP affinity of G(s)alphaL more efficiently than the GDP affinity of G(s)alphaS, and, thereby, decrease the efficiency of the agonist-free beta2AR and increase the efficiency of the agonist-occupied beta2AR at promoting GDP dissociation from G(s)alphaL. Thus, monovalent anions differentially regulate beta2AR-coupling to G(s)alphaS and G(s)alphaL.