MOLECULAR-IDENTIFICATION OF GUANINE-NUCLEOTIDE-BINDING REGULATORY PROTEINS WHICH COUPLE TO ENDOTHELIN RECEPTORS

The coupling of two endothelin receptor subtypes (ET(A) and ET(B)) to several types of guanine-nucleotide-binding regulatory protein (G protein) was examined. Two subtypes of receptor cDNAs were transfected alone or together with four different G protein alpha subunit cDNAs in COS-7 cells. In ET(A) receptor-transfected cells, endothelin-1 (ET-1) activated phosphatidylinositol-specific phospholipase C as measured by the production of phosphatidylinositol 1,4,5-trisphosphate [Ins(1,4,5)P-3]. ET(B)-receptor-transfected cells also produced Ins(1,4,5)P-3 on stimulation by ET-1. The ET-1-induced production of Ins(1,4,5)P-3 was markedly higher in G alpha(q)-cotransfected or G alpha(11)-cotransfected cells than in cells transfected with each receptor alone. ET-1 also stimulated production of cAMP in ET(A) or ET(B) receptor-transfected cells. The production of cAMP was synergistically amplified by G alpha(s) co-transfection with each receptor. In contrast, when G alpha(i2) was co-transfected with the ET(A) or ET(B) receptor, ET-1 displayed an inhibitory action on forskolin-stimulated cAMP accumulation. Pertussis-toxin treatment of the G alpha(i2)-transfected cells resulted in abolition of the endothelin-induced inhibition of cAMP accumulation. These observations indicate that both ET(A) and ET(B) receptors are able to couple to G(q), G(11), G(s) and G(i5), and suggest that endothelin receptors stimulate multiple effecters via several types of G protein simultaneously. The overall effects induced by endothelin may differ in cell types depending on the level of expression of each G-protein subtype in the cell.