Activating Mutations of the G-protein Subunit Α 11 Interdomain Interface Cause Autosomal Dominant Hypocalcemia Type 2.

Context: Autosomal dominant hypocalcemia types 1 and 2 (ADH1 and ADH2) are caused by germline gain-of-function mutations of the calcium-sensing receptor (CaSR) and its signaling partner, the G-protein subunit alpha(11) (G alpha(11)), respectively. More than 70 different gain-of-function CaSR mutations, but only 6 different gain-of-function G alpha(11) mutations are reported to date. Methods: We ascertained 2 additional ADH families and investigated them for CaSR and G alpha(11) mutations. The effects of identified variants on CaSR signaling were evaluated by transiently transfecting wild-type (WT) and variant expression constructs into HEK293 cells stably expressing CaSR (HEK-CaSR), and measuring intracellular calcium (Ca-i(2+)) and MAPK responses following stimulation with extracellular calcium (Ca-e(2+)). Results: CaSR variants were not found, but 2 novel heterozygous germline G alpha(11) variants, p.Gly66Ser and p.Arg149His, were identified. Homology modeling of these revealed that the Gly66 and Arg149 residues are located at the interface between the G alpha(11) helical and GTPase domains, which is involved in guanine nucleotide binding, and this is the site of 3 other reported ADH2 mutations. The Ca-i(2+) and MAPK responses of cells expressing the variant Ser66 or His149 G alpha(11) proteins were similar to WT cells at low Ca-e(2+), but significantly increased in a dose-dependent manner following Ca-e(2+) stimulation, thereby indicating that the p.Gly66Ser and p.Arg149His variants represent pathogenic gain-of-function G alpha(11) mutations. Treatment of Ser66- and His149-G alpha(11) expressing cells with the CaSR negative allosteric modulator NPS 2143 normalized Ca-i(2+) and MAPK responses. Conclusion: Two novel ADH2-causing mutations that highlight the G alpha(11) interdomain interface as a hotspot for gain-of-function G alpha(11) mutations have been identified.