Functional Coupling of G Proteins to Endothelin Receptors Is Ligand and Receptor Subtype Specific

1. The aims of the present study were (a) to determine the identity of the G proteins with which the endothelin receptor interacts and whether this interaction is subtype specific and (b) to determine whether agonist exposure can result in specific coupling between the endothelin receptor and G proteins.2. Coupling between endothelin A (ET_A) or endothelin B (ET_B) receptors and G proteins was assessed in two fibroblast cell lines, each expressing one receptor subtype. Four ligands, ET-1, ET-3, SRTXb, and SRTXc, were used for receptor stimulation. The G protein -subunit coupled to the receptor was identified by immunoprecipitation with an antibody against the endothelin receptor and immunoblotting with specific antibodies against different G protein -subunits.3. Unstimulated ET_A and ET_B receptors (ET_AR and ET_BR, respectively) were barely coupled to G_o. The unstimulated ET_AR coimmunoprecipitated with G_i3, whereas the unstimulated ET_BR was much less strongly coupled to G_i3. The coupling of ET_BR to G_i1G_i2 -subunits was much stronger than the coupling of ET_AR to these -subunits. Stimulation with the different ET agonists also resulted in differential coupling of G proteins to the receptor subtypes. All four ligands caused a strong increase in coupling of the ET_BR to G_i3, whereas coupling of the ET_AR to this subunit was not affected by ET-1 and was even decreased by SRTXc. On the other hand, all four ligands caused a much greater increase in the coupling of ET_AR to G_q/G₁₁ than in the coupling of ET_BR to these -subunits. Ligand-induced coupling between the receptors and the G_i1 and G_i2 -subunits is similar for the two receptor subtypes. The same was true for ligand-induced coupling of the receptors to G_o, except that ET-3 increased the coupling of this -subunit to ET_BR and decreased the coupling to ET_AR. Taken together, the results of this study show that coupling between ET receptors and G proteins is ligand and receptor subtype specific.4. It remains to be established whether this diversity of receptor–G protein coupling is of relevance for the various endothelin signaling pathways and/or pathological states.