AP2σ Mutations Impair Calcium-Sensing Receptor Trafficking and Signaling, and Show an Endosomal Pathway to Spatially Direct G-Protein Selectivity

Spatial control of G-protein-coupled receptor (GPCR) signaling, which is used by cells to translate complex information into distinct downstream responses, is achieved by using plasma membrane (PM) and endocytic-derived signaling pathways. The roles of the endomembrane in regulating such pleiotropic signaling via multiple G-protein pathways remain unknown. Here, we investigated the effects of disease-causing mutations of the adaptor protein-2 σ subunit (AP2σ) on signaling by the class C GPCR calcium-sensing receptor (CaSR). These AP2σ mutations increase CaSR PM expression yet paradoxically reduce CaSR signaling. Hypercalcemia-associated AP2σ mutations reduced CaSR signaling via Gα_q/11 and Gα_i/o pathways. The mutations also delayed CaSR internalization due to prolonged residency time of CaSR in clathrin structures that impaired or abolished endosomal signaling, which was predominantly mediated by Gα_q/11. Thus, compartmental bias for CaSR-mediated Gα_q/11 endomembrane signaling provides a mechanistic basis for multidimensional GPCR signaling. Gorvin et al. show that the class C GPCR calcium-sensing receptor (CaSR) mediates signaling from plasma membranes using Gα_q/11 and Gα_i/o and from endosomes by using only Gα_q/11. Adaptor protein-2 σ subunit (AP2σ) mutations impair CaSR internalization, leading to reduced sustained endosomal signaling and hypercalcemia in humans.