G protein-coupled receptor kinase 5 phosphorylation of hip regulates internalization of the chemokine receptor CXCR4

Biochemistry. 2011 Aug 16;50(32):6933-41. doi: 10.1021/bi2005202. Epub 2011 Jul 12.

Abstract

Regulation of the magnitude, duration, and localization of G protein-coupled receptor (GPCR) signaling responses is controlled by desensitization, internalization, and downregulation of the activated receptor. Desensitization is initiated by the phosphorylation of the activated receptor by GPCR kinases (GRKs) and the binding of the adaptor protein arrestin. In addition to phosphorylating activated GPCRs, GRKs have been shown to phosphorylate a variety of additional substrates. An in vitro screen for novel GRK substrates revealed Hsp70 interacting protein (Hip) as a substrate. GRK5, but not GRK2, bound to and stoichiometrically phosphorylated Hip in vitro. The primary binding domain of GRK5 was mapped to residues 303-319 on Hip, while the major site of phosphorylation was identified to be Ser-346. GRK5 also bound to and phosphorylated Hip on Ser-346 in cells. While Hip was previously implicated in chemokine receptor trafficking, we found that the phosphorylation of Ser-346 was required for proper agonist-induced internalization of the chemokine receptor CXCR4. Taken together, Hip has been identified as a novel substrate of GRK5 in vitro and in cells, and phosphorylation of Hip by GRK5 plays a role in modulating CXCR4 internalization.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Base Sequence
  • Carrier Proteins / metabolism*
  • Cell Line
  • Endocytosis*
  • G-Protein-Coupled Receptor Kinase 5
  • Humans
  • Immunoprecipitation
  • Phosphorylation
  • RNA, Small Interfering
  • Receptors, CXCR4 / metabolism*
  • Serine / metabolism
  • Substrate Specificity
  • Tumor Suppressor Proteins / metabolism*

Substances

  • CXCR4 protein, human
  • Carrier Proteins
  • RNA, Small Interfering
  • Receptors, CXCR4
  • ST13 protein, human
  • Tumor Suppressor Proteins
  • Serine
  • G-Protein-Coupled Receptor Kinase 5