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Phosphorylation and desensitization of human endothelin A and B receptors. Evidence for G protein-coupled receptor kinase specificity.

Publication ,  Journal Article
Freedman, NJ; Ament, AS; Oppermann, M; Stoffel, RH; Exum, ST; Lefkowitz, RJ
Published in: J Biol Chem
July 11, 1997

Although endothelin-1 can elicit prolonged physiologic responses, accumulating evidence suggests that rapid desensitization affects the primary G protein-coupled receptors mediating these responses, the endothelin A and B receptors (ETA-R and ETB-R). The mechanisms by which this desensitization proceeds remain obscure, however. Because some intracellular domain sequences of the ETA-R and ETB-R differ substantially, we tested the possibility that these receptor subtypes might be differentially regulated by G protein-coupled receptor kinases (GRKs). Homologous, or receptor-specific, desensitization occurred within 4 min both in the ETA-R-expressing A10 cells and in 293 cells transfected with either the human ETA-R or ETB-R. In 293 cells, this desensitization corresponded temporally with agonist-induced phosphorylation of each receptor, assessed by receptor immunoprecipitation from 32Pi-labeled cells. Agonist-induced receptor phosphorylation was not substantially affected by PKC inhibition but was reduced 40% (p << 0.03) by GRK inhibition, effected by a dominant negative GRK2 mutant. Inhibition of agonist-induced phosphorylation abrogated agonist-induced ETA-R desensitization. Overexpression of GRK2, -5, or -6 in 293 cells augmented agonist-induced ET-R phosphorylation approximately 2-fold (p << 0.02), but each kinase reduced receptor-promoted phosphoinositide hydrolysis differently. While GRK5 inhibited ET-R signaling by only approximately 25%, GRK2 inhibited ET-R signaling by 80% (p << 0.01). Congruent with its superior efficacy in suppressing ET-R signaling, GRK2, but not GRK5, co-immunoprecipitated with the ET-Rs in an agonist-dependent manner. We conclude that both the ETA-R and ETB-R can be regulated indistinguishably by GRK-initiated desensitization. We propose that because of its affinity for ET-Rs demonstrated by co-immunoprecipitation, GRK2 is the most likely of the GRKs to initiate ET-R desensitization.

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Published In

J Biol Chem

DOI

ISSN

0021-9258

Publication Date

July 11, 1997

Volume

272

Issue

28

Start / End Page

17734 / 17743

Location

United States

Related Subject Headings

  • beta-Adrenergic Receptor Kinases
  • Substrate Specificity
  • Receptors, Endothelin
  • Receptor, Endothelin B
  • Receptor, Endothelin A
  • Receptor Protein-Tyrosine Kinases
  • Rats
  • Protein Serine-Threonine Kinases
  • Phosphorylation
  • Phosphatidylinositols
 

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Freedman, N. J., Ament, A. S., Oppermann, M., Stoffel, R. H., Exum, S. T., & Lefkowitz, R. J. (1997). Phosphorylation and desensitization of human endothelin A and B receptors. Evidence for G protein-coupled receptor kinase specificity. J Biol Chem, 272(28), 17734–17743. https://doi.org/10.1074/jbc.272.28.17734
Freedman, N. J., A. S. Ament, M. Oppermann, R. H. Stoffel, S. T. Exum, and R. J. Lefkowitz. “Phosphorylation and desensitization of human endothelin A and B receptors. Evidence for G protein-coupled receptor kinase specificity.J Biol Chem 272, no. 28 (July 11, 1997): 17734–43. https://doi.org/10.1074/jbc.272.28.17734.
Freedman NJ, Ament AS, Oppermann M, Stoffel RH, Exum ST, Lefkowitz RJ. Phosphorylation and desensitization of human endothelin A and B receptors. Evidence for G protein-coupled receptor kinase specificity. J Biol Chem. 1997 Jul 11;272(28):17734–43.
Freedman, N. J., et al. “Phosphorylation and desensitization of human endothelin A and B receptors. Evidence for G protein-coupled receptor kinase specificity.J Biol Chem, vol. 272, no. 28, July 1997, pp. 17734–43. Pubmed, doi:10.1074/jbc.272.28.17734.
Freedman NJ, Ament AS, Oppermann M, Stoffel RH, Exum ST, Lefkowitz RJ. Phosphorylation and desensitization of human endothelin A and B receptors. Evidence for G protein-coupled receptor kinase specificity. J Biol Chem. 1997 Jul 11;272(28):17734–17743.

Published In

J Biol Chem

DOI

ISSN

0021-9258

Publication Date

July 11, 1997

Volume

272

Issue

28

Start / End Page

17734 / 17743

Location

United States

Related Subject Headings

  • beta-Adrenergic Receptor Kinases
  • Substrate Specificity
  • Receptors, Endothelin
  • Receptor, Endothelin B
  • Receptor, Endothelin A
  • Receptor Protein-Tyrosine Kinases
  • Rats
  • Protein Serine-Threonine Kinases
  • Phosphorylation
  • Phosphatidylinositols