PHOSPHORYLATION AND DESENSITIZATION OF HUMAN ENDOTHELIN-A AND ENDOTHELIN-B RECEPTORS - EVIDENCE FOR G-PROTEIN-COUPLED RECEPTOR KINASE SPECIFICITY

Although endothelin-l can elicit prolonged physiologic responses, accu mulating evidence suggests that rapid desensitization affects the prim ary G protein-coupled receptors mediating these responses, the endothe lin A and B receptors (ETA-R and ETB-R). The mechanisms by which this desensitization proceeds remain obscure, however. Because some intrace llular domain sequences of the ETA-R and ETB-R. differ substantially, we tested the possibility that these receptor subtypes might. be diffe rentially regulated by G protein-coupled receptor kinases (GRKs). Homo logous, or receptor-specific, desensitization occurred within 4 min bo th in the ETA-R-expressing A10 cells and in 293 cells transfected with either the human ETA-R or ETB-R, In 293 cells, this desensitization c orresponded temporally with agonist-induced phosphorylation of each re ceptor, assessed by receptor immunoprecipitation from P-32(i)-labeled cells. Agonist-induced receptor phosphorylation was not substantially affected by PKC inhibition but was reduced 40% (p < 0.03) by GRK inhib ition, effected by a dominant negative GRK2 mutant, Inhibition of agon ist-induced phosphorylation abrogated agonist-induced ETA-R desensitiz ation. Overexpression of GRK2, -5, or -6 in 293 cells augmented agonis t-induced ET-R phosphorylation similar to 2-fold (p < 0.02), but each kinase reduced receptor-promoted phosphoinositide hydrolysis different ly. While GRK5 inhibited ET-R signaling by only similar to 25%, GRK2 i nhibited ET-R signaling by 80% (p < 0.01), Congruent with its superior efficacy in suppressing ET-R signaling, GRK2, but not GRK5, co-immuno precipitated with the ET-Rs in an agonist-dependent manner. ma conclud e that both the ETA-R and ETB-R can be regulated indistinguishably by GRK-initiated desensitization, We propose that because of its affinity for ET-Rs demonstrated by co-immunoprecipitation, GRK2 is the most li kely of the GRKs to initiate ET-R desensitization.