Phosphorylation of the regulator of G protein signaling RGS9-1 by protein kinase A is a potential mechanism of light- and Ca2+-mediated regulation of G protein function in photoreceptors

Biochemistry. 2001 Oct 23;40(42):12619-27. doi: 10.1021/bi015624b.


In vertebrate photoreceptors, photoexcited rhodopsin interacts with the G protein transducin, causing it to bind GTP and stimulate the enzyme cGMP phosphodiesterase. The rapid termination of the active state of this pathway is dependent upon a photoreceptor-specific regulator of G protein signaling RGS9-1 that serves as a GTPase activating protein (GAP) for transducin. Here, we show that, in preparations of photoreceptor outer segments (OS), RGS9-1 is readily phosphorylated by an endogenous Ser/Thr protein kinase. Protein kinase C and MAP kinase inhibitors reduced labeling by about 30%, while CDK5 and CaMK II inhibitors had no effect. cAMP-dependent protein kinase (PKA) inhibitor H89 reduced RGS9-1 labeling by more than 90%, while dibutyryl-cAMP stimulated it 3-fold, implicating PKA as the major kinase responsible for RGS9-1 phosphorylation in OS. RGS9-1 belongs to an RGS subfamily also including RGS6, RGS7, and RGS11, which exist as heterodimers with the G protein beta subunit Gbeta5. Phosphorylated RGS9-1 remains associated with Gbeta5L, a photoreceptor-specific splice form, which itself was not phosphorylated. RGS9-1 immunoprecipitated from OS was in vitro phosphorylated by exogenous PKA. The PKA catalytic subunit could also phosphorylate recombinant RGS9-1, and mutational analysis localized phosphorylation sites to Ser(427) and Ser(428). Substitution of these residues for Glu, to mimic phosphorylation, resulted in a reduction of the GAP activity of RGS9-1. In OS, RGS9-1 phosphorylation required the presence of free Ca(2+) ions and was inhibited by light, suggesting that RGS9-1 phosphorylation could be one of the mechanisms mediating a stronger photoresponse in dark-adapted cells.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Calcium / metabolism
  • Calcium / physiology*
  • Cattle
  • Chelating Agents / pharmacology
  • Cyclic AMP-Dependent Protein Kinases / physiology*
  • Egtazic Acid / pharmacology
  • GTP-Binding Proteins / metabolism*
  • GTP-Binding Proteins / physiology
  • GTPase-Activating Proteins / physiology
  • Light*
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Phosphorylation / drug effects
  • Photoreceptor Cells, Vertebrate / drug effects
  • Photoreceptor Cells, Vertebrate / metabolism*
  • Photoreceptor Cells, Vertebrate / physiology
  • RGS Proteins / genetics
  • RGS Proteins / metabolism*
  • RGS Proteins / physiology
  • Rod Cell Outer Segment / drug effects
  • Rod Cell Outer Segment / enzymology
  • Rod Cell Outer Segment / metabolism
  • Serine / genetics
  • Serine / metabolism
  • Vision, Ocular / genetics
  • Vision, Ocular / physiology*


  • Chelating Agents
  • GTPase-Activating Proteins
  • RGS Proteins
  • regulator of g-protein signaling 9
  • Serine
  • Egtazic Acid
  • Cyclic AMP-Dependent Protein Kinases
  • GTP-Binding Proteins
  • Calcium