Gα_q-coupled Receptor Internalization Specifically Induced by Gα_q Signaling

In the present report, we investigated the effect of ezrin-radixin-moesin-binding phosphoprotein 50 (EBP50) expression on the agonist-induced internalization of the thromboxane A₂ β receptor (TPβ receptor). Interestingly, we found that EBP50 almost completely blocked TPβ receptor internalization, which could not be reversed by overexpression of G protein-coupled receptor (GPCR) kinases and arrestins. Because we recently demonstrated that EBP50 can bind to and inhibit Gα_q, we next studied whether Gα_q signaling could induce TPβ receptor internalization, addressing the long standing question about the relationship between GPCR signaling and their internalization. Expression of a constitutively active Gα_q mutant (Gα_q-R183C) resulted in a robust internalization of the TPβ receptor, which was unaffected by expression of dominant negative mutants of arrestin-2 and -3, but inhibited by expression of EBP50 or dynamin-K44A, a dominant negative mutant of dynamin. Phospholipase Cβ and protein kinase C did not appear to significantly contribute to internalization of the TPβ receptor, suggesting that Gα_q induces receptor internalization through a phospholipase Cβ- and protein kinase C-independent pathway. Surprisingly, there appears to be specificity in Gα protein-mediated GPCR internalization. Gα_q-R183C also induced the internalization of CXCR4 (Gα_q-coupled), whereas it failed to do so for the β₂-adrenergic receptor (Gα_s-coupled). Moreover, Gα_s-R201C, a constitutively active form of Gα_s, had no effect on internalization of the TPβ, CXCR4, and β₂-adrenergic receptors. Thus, we showed that Gα protein signaling can lead to internalization of GPCRs, with specificity in both the Gα proteins and GPCRs that are involved. Furthermore, a new function has been described for EBP50 in its capacity to inhibit receptor endocytosis.