The Role of the Adaptor Protein-2 Sigma Subunit (AP2σ) in Calcium Homeostasis (360G-Wellcome-205011_Z_16_Z)

G-protein coupled receptors (GPCRs) facilitate cellular responses to extracellular stimuli, and GPCR mutations result in many diseases, including endocrine disorders. Mutations of the calcium-sensing receptor (CaSR), a pivotal GPCR in calcium homeostasis, resulting in loss-of-function or gain-of-function cause familial hypocalciuric hypercalcaemia type-1 (FHH1) and autosomal dominant hypocalcaemia type-1 (ADH1), respectively. FHH and ADH are genetically heterogeneous, and loss-of-function or gain-of-function mutations of the G-protein-alpha11 utilised by CaSR, cause FHH2 and ADH2, respectively; while loss-of-function mutations of adaptor protein-2 sigma subunit (AP2sigma), which is critical for clathrin-mediated endocytosis, cause FHH3. To date, the reported AP2sigma mutations, which cause only FHH3 but not ADH, all affect residue Arg15 and impair CaSR signalling and trafficking. However, the Thakker group have recently identified seven other AP2sigma variants (Arg3His, Ala44Thr, Phe52Tyr, Arg61His, Thr112Met, Met117Ile, Glu120Gly), and my goal is to determine the roles of these AP2sigma variants in calcium homeostasis by: 1) Characterising their structural-functional relationships by in vitro studies that assess effects on three-dimensional models and on CaSR signalling and trafficking pathways. 2) Determining in vivo phenotypes of AP2sigma mutations by generating knock-in mouse models, using CRISPR-Cas. 3) Assessing pharmacological effects of drugs, e.g. velcalcetide, using above AP2sigma in vitro and in vivo models.