Regulation of the Platelet-derived Growth Factor Receptor-β by G Protein-coupled Receptor Kinase-5 in Vascular Smooth Muscle Cells Involves the Phosphatase Shp2

Abstract

Smooth muscle cell (SMC) proliferation and migration are substantially controlled by the platelet-derived growth factor receptor-␤ (PDGFR␤), which can be regulated by the Ser/Thr kinase G protein-coupled receptor kinase-2 (GRK2). In mouse aortic SMCs, however, we found that prolonged PDGFR␤ activation engendered down-regulation of GRK5, but not GRK2; moreover, GRK5 and PDGFR␤ were coordinately up-regulated in SMCs from atherosclerotic arteries. With SMCs from GRK5 knock-out and cognate wild type mice (five of each), we found that physiologic expression of GRK5 increased PDGF-promoted PDGFR␤ seryl phosphorylation by 3-fold and reduced PDGFR␤-promoted phosphoinositide hydrolysis, thymidine incorporation, and overall PDGFR␤ tyrosyl phosphorylation by ϳ35%. Physiologic SMC GRK5 activity also increased PDGFR␤ association with the phosphatase Shp2 (8-fold), enhanced phosphorylation of PDGFR␤ Tyr 1009 (the docking site for Shp2), and reduced phosphorylation of PDGFR␤ Tyr 1021. Consistent with having increased PDGFR␤-associated Shp2 activity, GRK5-expressing SMCs demonstrated greater PDGF-induced Src activation than GRK5-null cells. GRK5-mediated desensitization of PDGFR␤ inositol phosphate signaling was diminished by Shp2 knock-down or impairment of PDGFR␤/Shp2 association. In contrast to GRK5, physiologic GRK2 activity did not alter PDGFR␤/Shp2 association. Finally, purified GRK5 effected agonist-dependent seryl phosphorylation of partially purified PDGFR␤s. We conclude that GRK5 mediates the preponderance of PDGF-promoted seryl phosphorylation of the PDGFR␤ in SMCs, and, through mechanisms involving Shp2, desensitizes PDGFR␤ inositol phosphate signaling and enhances PDGFR␤-triggered Src activation.

β-Arrestins Regulate Atherosclerosis and Neointimal Hyperplasia by Controlling Smooth Muscle Cell Proliferation and Migration

“…22 Mitogenic signaling in primary SMCs more than doubles when agonists for G q/11 -and G i -coupled 7-transmembrane receptors are added to PDGF, even at receptor-saturating PDGF concentrations. 11,24 By altering SMC mitogenic signaling elicited by 7-transmembrane receptors, ␤-arrestin activity should augment (␤-arrestin2) or reduce (␤-arrestin1) mitogenic signaling elicited by the combination of PDGF and plateletderived agonists for 7-transmembrane receptors. Furthermore, because PDGFRs can be transactivated by 7-transmembrane receptors, 22 ␤-arrestin-mediated regulation of 7-transmembrane receptors may alter levels of SMC PDGFR activation.…”

“…For carotid endothelial denudation, we used congenic age-and sex-matched C57BL/6 WT, ␤-arrestin1 Ϫ/Ϫ , ␤-arrestin2 Ϫ/Ϫ , and green fluorescent protein (GFP)-transgenic mice. Bone marrow transplantation, SMC migration, 10 RNA interference, 11 and cell proliferation assays 11 are described in the expanded Materials and Methods section in the online data supplement, available at http://circres.ahajournals.org.…”

Protein Tyrosine Phosphatase SHP-2 Is Positively Involved in Platelet-Derived Growth Factor–Signaling in Vascular Neointima Formation via the Reactive Oxygen Species–Related Pathway

“…SHP-2 is believed to be a target of protein tyrosine kinases. It can bind directly to PDGF-R (15,16) and is thought to contribute to the proximal signaling events of PDGF-R (17,18). Although most PTPs increase the dephosphorylation of proteins and act as negative regulators in response to extracellular stimuli, SHP-2 is believed to be a positive mediator of the PDGF signal (19,20).…”

G Protein–Coupled Receptor Kinase GRK5 Phosphorylates Moesin and Regulates Metastasis in Prostate Cancer

“…GRK5 phosphorylates the non-GPCR substrate nucleophosmin, thereby altering sensitivity of breast cancer cells to polo-like kinase inhibitor-induced apoptosis (9). Also, GRK5 phosphorylates receptor tyrosine kinases platelet-derived growth factor receptor (10,11) and VEGFR (12) that may play a role in cancer. Screening with shRNA library identified GRK5 as a potential regulator of cell-cycle progression (13), and GRK5 expression levels correlate with prostate cell proliferation in vitro and tumor growth in animals (14).…”