Reciprocal regulation of the platelet-derived growth factor receptor-beta and G protein-coupled receptor kinase 5 by cross-phosphorylation: effects on catalysis.
Signaling by the platelet-derived growth factor receptor-beta (PDGFRbeta) is diminished when the PDGFRbeta is phosphorylated on seryl residues by G protein-coupled receptor kinase-5 (GRK5), but mechanisms for GRK5 activation by the PDGFRbeta remain obscure. We therefore tested whether the PDGFRbeta is able to tyrosine-phosphorylate and thereby activate GRK5. Purified GRK5 was tyrosine-phosphorylated by the wild-type PDGFRbeta to a stoichiometry of 0.8 mol phosphate/mol GRK5, an extent approximately 5 times greater than observed with a Y857F PDGFRbeta mutant that fails to phosphorylate exogenous substrates but autophosphorylates and activates Src normally. The degree of PDGFRbeta-mediated phosphorylation of GRK5 correlated with GRK5 activity, as assessed by seryl phosphorylation of the PDGFRbeta in purified protein preparations, in intact cells expressing a tyrosine-to-phenylalanine GRK5 mutant, and in GRK5 peptide phosphorylation assays. However, tyrosyl phosphorylation of GRK5 was not necessary for GRK5-mediated phosphorylation of the beta(2)-adrenergic receptor, even though beta(2)-adrenergic receptor activation promoted tyrosyl phosphorylation of GRK5 in smooth muscle cells. Phosphorylation of the PDGFRbeta by GRK5 in smooth muscle cells or in purified protein preparations reduced PDGFRbeta-mediated peptide phosphorylation. In contrast, phosphorylation of GRK5 by the PDGFRbeta enhanced the V(max) of GRK5-mediated peptide phosphorylation, by 3.4-fold, without altering the GRK5 K(M) for peptide. We conclude that GRK5 tyrosyl phosphorylation is required for the activation of GRK5 by the PDGFRbeta, but not by the beta(2)-adrenergic receptor, and that by activating GRK5, the PDGFRbeta triggers its own desensitization.
Cai
Xinjiang
X
Departments of Medicine/Cardiology, Duke University Medical Center, Durham, North Carolina, USA.
Wu
Jiao-Hui
JH
Exum
Sabrina T
ST
Oppermann
Martin
M
Premont
Richard T
RT
Shenoy
Sudha K
SK
Freedman
Neil J
NJ
D000595:Amino Acid Sequence; D000818:Animals; D002384:Catalysis; D002417:Cattle; D002460:Cell Line; D054774:G-Protein-Coupled Receptor Kinase 5 / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D010766:Phosphorylation / Q000502:physiology; D020797:Receptor, Platelet-Derived Growth Factor beta / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D018411:Spodoptera; D013379:Substrate Specificity / Q000502:physiology; D014443:Tyrosine / Q000378:metabolism
D014443:Tyrosine; D020797:Receptor, Platelet-Derived Growth Factor beta; D054774:G-Protein-Coupled Receptor Kinase 5