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Cell Surface Calcium‐Sensing Receptor Heterodimers: Mutant Gene Dosage Affects Ca 2+ Sensing but Not G Protein Interaction
Author(s) -
Goolam Mahvash A.,
Brown Alice P.,
Edwards Kimberly T.,
Gregory Karen J.,
Leach Katie,
Conigrave Arthur D.
Publication year - 2022
Publication title -
journal of bone and mineral research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.882
H-Index - 241
eISSN - 1523-4681
pISSN - 0884-0431
DOI - 10.1002/jbmr.4651
Subject(s) - calcium sensing receptor , mutant , hek 293 cells , g protein coupled receptor , receptor , microbiology and biotechnology , chemistry , transfection , calcium , extracellular , endoplasmic reticulum , mutation , wild type , transmembrane domain , signal transduction , calcium signaling , biology , biochemistry , calcium metabolism , gene , organic chemistry
The calcium‐sensing receptor is a homodimeric class C G protein‐coupled receptor (GPCR) that senses extracellular Ca 2+ (Ca 2+ o ) via a dimeric extracellular Venus flytrap (VFT) unit that activates G protein‐dependent signaling via twin Cysteine‐rich domains linked to transmembrane heptahelical (HH) bundles. It plays a key role in the regulation of human calcium and thus mineral metabolism. However, the nature of interactions between VFT units and HH bundles, and the impacts of heterozygous or homozygous inactivating mutations, which have implications for disorders of calcium metabolism are not yet clearly defined. Herein we generated CaSR‐GABA B1 and CaSR‐GABA B2 chimeras subject to GABA B ‐dependent endoplasmic reticulum sorting to traffic mutant heterodimers to the cell surface. Transfected HEK‐293 cells were assessed for Ca 2+ o ‐stimulated Ca 2+ i mobilization using mutations in either the VFT domains and/or HH bundle intraloop‐2 or intraloop‐3. When the same mutation was present in both VFT domains of receptor dimers, analogous to homozygous neonatal severe hyperparathyroidism (NSHPT), receptor function was markedly impaired. Mutant heterodimers containing one wild‐type (WT) and one mutant VFT domain, however, corresponding to heterozygous familial hypocalciuric hypercalcemia type‐1 (FHH‐1), supported maximal signaling with reduced Ca 2+ o potency. Thus two WT VFT domains were required for normal Ca 2+ o potency and there was a pronounced gene‐dosage effect. In contrast, a single WT HH bundle was insufficient for maximal signaling and there was no functional difference between heterodimers in which the mutation was present in one or both intraloops; ie, no gene‐dosage effect. Finally, we observed that the Ca 2+ o ‐stimulated CaSR operated exclusively via signaling in‐trans and not via combined in‐trans and in‐cis signaling. We consider how receptor asymmetry may support the underlying mechanisms. © 2022 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).