Proper targeting and activity of a nonfunctioning thyroid-stimulating hormone receptor (TSHr) combining an inactivating and activating TSHr mutation in one receptor

Patrizia Agretti, Giuseppina De Marco, Paola Collecchi, Luca Chiovato, Paolo Vitti, Aldo Pinchera, Massimo Tonacchera

Research output: Contribution to journalArticle

Abstract

Activating mutations of the thyroid-stimulating hormone receptor (TSHr) have been identified as a cause of toxic adenomas. Germline-inactivating TSHr mutations have been described as a cause of congenital hypothyroidism. The effects of combining activating and inactivating mutations within a single receptor was studied. The double mutant T477I/P639S contained an activating TSHr mutation (P639S) together with an inactivating one (T477I). The other one (I486M/P639S) contained two activating mutations. Constructs were expressed in COS-7 cells and basal and TSH-stimulated cyclic AMP (cAMP) accumulation and inositol phosphate (IP) production were determined. The expression at the cell surface was studied both with binding and fluorescence-activated cell scanning analysis. Our results show that the effect of combining the two activating mutations is an increase in the constitutive activity only for the cAMP pathway and not for the IP pathway suggesting that different mutations result in receptor conformations with different relative abilities to couple to G s-alpha or Gq-alpha. Surprisingly the double mutant containing the T477I behaves as an activating receptor with constitutive activity both for the cAMP and IP pathways. These data show that an inactive form of the TSHr which is trapped inside a cell after transfection is able to gain the membrane surface when combined with an activated form of the receptor.

Original languageEnglish
Pages (from-to)3839-3847
Number of pages9
JournalEuropean Journal of Biochemistry
Volume270
Issue number18
DOIs
Publication statusPublished - Sep 2003

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Keywords

  • Constitutive activity
  • G-protein-coupled receptors
  • Germline mutations
  • Site-directed mutagenesis
  • Somatic mutations
  • TSH receptor

ASJC Scopus subject areas

  • Biochemistry

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