Endocrine Abstracts

Loss-of-function mutations of the calcium-sensing receptor (CaSR), a G-protein-coupled receptor (GPCR), result in familial hypocalciuric hypercalcaemia (FHH), a disorder of extracellular calcium homeostasis affecting the parathyroids and kidneys. However, around 35% of FHH patients do not have CaSR mutations. A form of FHH, designated FHH2, has been mapped to chromosome 19p. The GNA11 gene, encoding G-protein α11 (Gα₁₁), a component of the CaSR signalling pathway, resides on 19p and is therefore a candidate gene for FHH2. We investigated the FHH2 kindred for GNA11 mutations and identified an in-frame isoleucine deletion (Ile199del). GNA11 mutational analysis was also undertaken in nine previously reported FHH patients who did not have CaSR mutations, and this revealed a Leu135Gln missense mutation in one of these patients. To assess the functional consequences of these mutations, wild-type and mutant Gα₁₁ proteins were expressed in HEK293 cells stably transfected with CaSR, and the intracellular calcium response to changes in extracellular calcium was measured. The Ile199del and Leu135Gln mutations both led to a rightward shift of the concentration-response curves with significantly (P<0.0001) increased mean EC50 values of 2.52 mM (95% confidence interval (CI)=2.49–2.56) and 3.46 mM (95%CI =3.40–3.51), respectively, when compared to the wild-type Gα₁₁ EC50 of 2.29 mM (95%CI=2.24–2.34). These findings indicated that the Ile199del and Leu135Gln mutations were associated with a loss of Gα₁₁ function. An examination of the crystal structure of Gα_q, which has 90% amino acid identity to Gα₁₁, indicated that both of these mutations were located in Gα structural motifs that are critical for GPCR signalling. In particular, the Ile199del mutation disrupted hydrogen bonding within a peptide loop that comprises part of the Gα-GPCR interface. This study has identified the genetic abnormality causing FHH2, and further increased our understanding of extracellular calcium homeostasis and the structure–function relationships of G-protein α subunits.

Declaration of funding: Yes.

Details: This work was supported by Medical Research Council programme grants – G9825289 and G1000467, National Institute for Health Research Oxford Biomedical Research Centre Programme, and European Commission Seventh Framework Programme – FP7-264663. One of the co-authors is a Wellcome Trust Clinical Research Training Fellow.

DOI: 10.1530/endoabs.31.P1