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 α 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)P₃]. ET_B‐receptor‐transfected cells also produced Ins(1,4,5)P₃ on stimulation by ET‐1. The ET‐1‐induced production of Ins(1,4,5)P₃ was markedly higher in Gα_q‐cotransfected or Gα₁₁‐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 Gα_s co‐transfection with each receptor. In contrast, when Gα_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 Gα_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, G₁₁, G_s, and G_i2, and suggest that endothelin receptors stimulate multiple effectors 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.