Selective regulation of Gαq/11 by an RGS domain in the G protein-coupled receptor kinase, GRK2

G protein-coupled receptor kinases (GRKs) are well characterized regulators of G protein-coupled receptors, whereas regulators of G protein signaling (RGS) proteins directly control the activity of G protein alpha subunits. Interestingly, a recent report (Siderovski, D. P., Hessel, A., Chung, S., Mak, T. W., and Tyers, M. (1996) Curr. Biol. 6, 211-212) identified a region within the N terminus of GRKs that contained homology to RGS domains. Given that RGS domains demonstrate AlF4--dependent binding to G protein alpha subunits, we tested the ability of G proteins from a crude bovine brain extract to bind to GRK affinity columns in the absence or presence of AlF4-. This revealed the specific ability of bovine brain G alpha(q/11) to bind to both GRK2 and GRK3 in an AlF4--dependent manner. In contrast, G alpha(s), G alpha(i), and G alpha(12/13) did not bind to GRK2 or GRK3 despite their presence in the extract. Additional studies revealed that bovine brain G alpha(q/11) could also bind to an N-terminal construct of GRK2, while no binding of G alpha(q/11) G alpha(s), G alpha(i), or G alpha(12/13) to comparable constructs of GRK5 or GRK6 was observed. Experiments using purified G alpha(q) revealed significant binding of both G alpha(q) GDP/AlF4- and G alpha(q)(GTP gamma S), but not G alpha(q)(GDP), to GRK2. Activation-dependent binding was also observed in both COS-1 and HEK293 cells as GRK2 significantly co-immunoprecipitated constitutively active G alpha(q)(R183C) but not mild type G alpha(q). ln vitro analysis revealed that GRK2 possesses weak GAP activity toward G alpha(q) that is dependent on the presence of a G protein-coupled receptor. However, GRK2 effectively inhibited G alpha(q)-mediated activation of phospholipase C-beta both in. vitro and in cells, possibly through sequestration of activated G alpha(q). These data suggest that a subfamily of the GRKs may be bifunctional regulators of G protein-coupled receptor signaling operating directly on both receptors and G proteins.