Phosphorylation of the platelet-derived growth factor receptor-β by G protein-coupled receptor kinase-2 reduces receptor signaling and interaction with the Na+/H+ exchanger regulatory factor

Abstract

G protein-coupled receptor kinase-2 (GRK2) can phosphorylate and desensitize the platelet-derived growth factor receptor-β (PDGFRβ) in heterologous cellular systems. To determine whether GRK2 regulates the PDGFRβ in physiologic systems, we examined PDGFRβ signaling in mouse embryonic fibroblasts from GRK2-null and cognate wild type mice. To discern a mechanism by which GRK2-mediated phosphorylation can desensitize the PDGFRβ, but not the epidermal growth factor receptor (EGFR), we investigated effects of GRK2-mediated phosphorylation on the association of the PDGFRβ with the Na+/H+ exchanger regulatory factor (NHERF), a protein shown to potentiate dimerization of the PDGFRβ, but not the EGFR. Physiologic expression of GRK2 diminished (a) phosphoinositide hydrolysis elicited through the PDGFRβ but not heterotrimeric G proteins; (b) Akt activation evoked by the PDGFRβ but not the EGFR; and (c) PDGF-induced tyrosyl phosphorylation of the PDGFRβ itself. PDGFRβ desensitization by physiologically expressed GRK2 correlated with a 2.5-fold increase in PDGF-promoted PDGFRβ seryl phosphorylation. In 293 cells, GRK2 overexpression reduced PDGFRβ/ NHERF association by 60%. This effect was reproduced by S1104D mutation of the PDGFRβ, which also diminished PDGFRβ activation and signaling (like the S1104A mutation) to an extent equivalent to that achieved by GRK2-mediated PDGFRβ phosphorylation. GRK2 overexpression desensitized only the wild type but not the S1104A PDGFRβ. We conclude that GRK2-mediated PDGFRβ seryl phosphorylation plays an important role in desensitizing the PDGFRβ in physiologic systems. Furthermore, this desensitization appears to involve GRK2-mediated phosphorylation of PDGFRβ Ser1104, with consequent dissociation of the PDGFRβ from NHERF.