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Vasoactive intestinal polypeptide type-1 receptor regulation. Desensitization, phosphorylation, and sequestration.

Publication ,  Journal Article
Shetzline, MA; Walker, JK; Valenzano, KJ; Premont, RT
Published in: The Journal of biological chemistry
July 2002

The vasoactive intestinal polypeptide type-1 (VPAC(1)) receptor is a class II G protein-coupled receptor, distinct from the adrenergic receptor superfamily. The mechanisms involved in the regulation of the VPAC(1) receptor are largely unknown. We examined agonist-dependent VPAC(1) receptor signaling, phosphorylation, desensitization, and sequestration in human embryonic kidney 293 cells. Agonist stimulation of cells overexpressing this receptor led to a dose-dependent increase in cAMP that peaked within 5-10 min and was completely desensitized after 20 min. Cells cotransfected with the VPAC(1) receptor (VPAC(1)R) and G protein-coupled receptor kinases (GRKs) 2, 3, 5, and 6 exhibited enhanced desensitization that was not evident with GRK 4. Immunoprecipitation of the epitope-tagged VPAC(1) receptor revealed dose-dependent phosphorylation that was increased with cotransfection of any GRK. Agonist-stimulated internalization of the VPAC(1)R peaked in 10 min, and neither overexpressed beta-arrestin nor its dominant-negative mutant altered internalization. However, a dynamin-dominant negative mutant did inhibit VPAC(1) receptor internalization. Interestingly, VPAC(1)R specificity in desensitization was not evident by study of the overexpressed receptor; however, we determined that human embryonic kidney 293 cells express an endogenous VPAC(1)R that did demonstrate dose-dependent GRK specificity. Therefore, VPAC(1) receptor regulation involves agonist-stimulated, GRK-mediated phosphorylation, beta-arrestin translocation, and dynamin-dependent receptor internalization. Moreover, study of endogenously expressed receptors may provide information not evident in overexpressed systems.

Duke Scholars

Published In

The Journal of biological chemistry

ISSN

0021-9258

Publication Date

July 2002

Volume

277

Issue

28

Start / End Page

25519 / 25526

Location

united states

Related Subject Headings

  • beta-Arrestins
  • Receptors, Vasoactive Intestinal Polypeptide, Type I
  • Receptors, Vasoactive Intestinal Peptide
  • Protein Transport
  • Precipitin Tests
  • Phosphorylation
  • Microscopy, Fluorescence
  • Humans
  • Cell Line
  • Biochemistry & Molecular Biology
 

Citation

APA
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MLA
NLM
Shetzline, M. A., Walker, J. K., Valenzano, K. J., & Premont, R. T. (2002). Vasoactive intestinal polypeptide type-1 receptor regulation. Desensitization, phosphorylation, and sequestration. The Journal of Biological Chemistry, 277(28), 25519–25526.
Shetzline, M. A., J. K. Walker, K. J. Valenzano, and R. T. Premont. “Vasoactive intestinal polypeptide type-1 receptor regulation. Desensitization, phosphorylation, and sequestration.The Journal of Biological Chemistry 277, no. 28 (July 2002): 25519–26.
Shetzline MA, Walker JK, Valenzano KJ, Premont RT. Vasoactive intestinal polypeptide type-1 receptor regulation. Desensitization, phosphorylation, and sequestration. The Journal of biological chemistry. 2002 Jul;277(28):25519–26.
Shetzline, M. A., et al. “Vasoactive intestinal polypeptide type-1 receptor regulation. Desensitization, phosphorylation, and sequestration.The Journal of Biological Chemistry, vol. 277, no. 28, July 2002, pp. 25519–26.
Shetzline MA, Walker JK, Valenzano KJ, Premont RT. Vasoactive intestinal polypeptide type-1 receptor regulation. Desensitization, phosphorylation, and sequestration. The Journal of biological chemistry. 2002 Jul;277(28):25519–25526.

Published In

The Journal of biological chemistry

ISSN

0021-9258

Publication Date

July 2002

Volume

277

Issue

28

Start / End Page

25519 / 25526

Location

united states

Related Subject Headings

  • beta-Arrestins
  • Receptors, Vasoactive Intestinal Polypeptide, Type I
  • Receptors, Vasoactive Intestinal Peptide
  • Protein Transport
  • Precipitin Tests
  • Phosphorylation
  • Microscopy, Fluorescence
  • Humans
  • Cell Line
  • Biochemistry & Molecular Biology