Searchable abstracts of presentations at key conferences in endocrinology
Endocrine Abstracts (2016) 44 P47 | DOI: 10.1530/endoabs.44.P47

SFEBES2016 Poster Presentations Bone and Calcium (20 abstracts)

Studies of an Autosomal Dominant Hypocalcemia type-1 (ADH1) associated calcium-sensing receptor (CaSR) mutation, Arg680Gly, provides insights into biased signalling

Caroline Gorvin 1 , Valerie Babinsky 1 , Anders Schou 2 , Peter Nissen 3 , Fadil Hannan 4 & Rajesh Thakker 1


1University of Oxford, Academic Endocrine Unit, Oxford Centre for Diabetes, Endocrinology and Metabolism, Radcliffe Department of Medicine, Oxford, UK; 2Hans Christian Andersen Children’s Hospital, Odense University Hospital, Odense, Denmark; 3Department of Clinical Biochemistry, Aarhus University Hospital, Aarhus, Denmark; 4University of Liverpool, Institute of Ageing and Chronic Disease, Liverpool, UK.


The CaSR, a G-protein-coupled receptor that regulates extracellular calcium (Ca2+o), predominantly signals via G-protein-αq/11 (Gαq/11), initiating IP3-mediated intracellular calcium (Ca2+i) accumulation, and mitogen-activated protein kinase (MAPK) signalling. CaSR also activates MAPK signalling via Gαi/o, or by associating with the scaffolding protein β-arrestin. CaSR gain-of-function mutations cause autosomal dominant hypocalcaemia type-1 (ADH1). Mutational analysis of CASR in a seven-year-old male and his father with ADH1 identified a novel heterozygous mutation (p.Arg680Gly) in both patients. The variant was functionally characterised in HEK293 cells transiently expressing WT (Arg680) or mutant Gly680 CaSRs using: a flow cytometry assay to measure Ca2+i; and a SRE luciferase reporter gene to assess MAPK signalling, in response to Ca2+o elevations. In contrast to reported ADH1 mutations, Gly680 had no effect on Ca2+i, but did significantly elevate MAPK responses. Measurements of the IP3 breakdown product IP1 showed no significant difference between WT and Gly680 expressing cells, confirming Gαq/11- signalling was not responsible for elevated Gly680 MAPK responses. We hypothesized such differences could instead be due to biased signalling by Gαi/o or a G-protein-independent β-arrestin-mediated pathway. To test the former, we assessed the effect of the Gαi/o-blocking agent pertussis toxin (PTx) on SRE reporter responses. PTx reduced SRE responses in both WT and Gly680 expressing cells to similar levels, indicating Gαi/o is not responsible for elevated MAPK signalling. To investigate G-protein-independent mechanisms we used a β-arrestin-1 siRNA, which had no effect on WT SRE reporter activity, but significantly reduced responses in Gly680 expressing cells. Structural homology modelling predicts Arg680 to form a salt bridge with CaSR transmembrane domain-7, which limits binding of β-arrestin; thus loss of this salt bridge would enhance β-arrestin binding and MAPK signalling. In conclusion, we report a CaSR mutation associated with hypocalcaemia that enhances MAPK signalling via a unique G-protein-independent mechanism involving β-arrestin-1.

Volume 44

Society for Endocrinology BES 2016

Brighton, UK
07 Nov 2016 - 09 Nov 2016

Society for Endocrinology 

Browse other volumes

Article tools

My recent searches

No recent searches.