Role of receptor phosphorylation in desensitization and internalization of the secretin receptor

Martin H. Holtmann, Belinda F. Roettger, Delia I. Pinon, Laurence J. Miller

Research output: Contribution to journalArticle

75 Scopus citations

Abstract

The secretin receptor is prototypic of a recently described family of G protein-coupled receptors. We recently demonstrated its phosphorylation in response to agonist stimulation and elimination of this covalent modification by C-terminal truncation (F. Ozcelebi et al. (1995) Mol. Pharmacol. 48, 818- 824). Here, we explore the functional impact of receptor phosphorylation and structural determinants for desensitization by comparing receptor behavior after agonist exposure in cell lines expressing wild-type and truncated receptor. To characterize receptor internalization, a novel fluorescent full agonist, [rat secretin-27]-Gly-rhodamine, was developed, which bound specifically and with high affinity. Both receptor constructs bound secretin normally, leading to normal G protein coupling and cAMP accumulation and prompt receptor internalization. Exposure to 10 nM secretin for 5 min or 12 h prior to washing and restimulation with a full range of concentrations demonstrated absent cAMP responses in wild-type receptor-bearing cells and responses 25 to 30% of control and shifted 1 order of magnitude to the right in the truncated receptor-bearing cells. Thus, the major mechanism of desensitization was phosphorylation-independent receptor internalization. Phosphorylation was associated with a distinct process that likely represents interference with G protein coupling, manifest as a reduced rate of cAMP stimulation. Thus, dual distinct mechanisms of desensitization exist in the secretin receptor family that should help protect receptor-bearing cells from overstimulation.

Original languageEnglish (US)
Pages (from-to)23566-23571
Number of pages6
JournalJournal of Biological Chemistry
Volume271
Issue number38
DOIs
StatePublished - Sep 30 1996

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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