Stable association of G proteins with beta(2)AR is independent of the state of receptor activation

beta(2)-Adrenergic receptors expressed in Sf9 cells activate endogenous Gs and adenylyl cyclase [Mouillac B., Caron M., Bonin H., Dennis M. and Bouvie r M. (1992) J. Biol. Chem.. 267, 21733-21737]. However, high affinity agoni st binding is not detectable under these conditions suggesting an improper stoichiometry between the receptor and the G protein and possibly the effec tor molecule as well. In this study we demonstrate that when beta(2)-adrene rgic receptors were co expressed with various mammalian G protein subunits in Sf9 cells using recombinant baculoviruses signalling properties found in native receptor systems were reconstituted. For example, when beta(2)AR wa s co-expressed with the Gs alpha subunit, maximal receptor-mediated adenyly l cyclase stimulation was greatly enhanced (60 +/- 9.0 versus 150 +/- 52 pm ol cAMP/min/mg protein) and high affinity, GppNHp-sensitive, agonist bindin g was detected. When G beta gamma subunits were co-expressed with Gs alpha and the beta(2)AR, receptor stimulated GTPase activity was also demonstrate d, in contrast to when the receptor was expressed alone, and this activity was higher than when beta(2)AR was co-expressed with Gs alpha alone. Other properties of the receptor, including receptor desensitization and response to inverse agonists were unaltered. Using antisera against an epitope-tagg ed beta(2)AR, both Gs alpha and beta gamma subunits could be co-immunopreci pitated with the beta(2)AR under conditions where subunit dissociation woul d be expected given current models of G protein function. A desensitization -defective beta(2)AR (S261, 262, 345, 346A) and a mutant which is constitut ively desensitized (C341G) could also co-immunoprecipitate G protein subuni ts. These results will be discussed in terms of a revised view of G protein -mediated signalling which may help address issues of specificity in recept or/G protein coupling. (C) 1999 Elsevier Science Inc.