A calcium-sensing receptor mutation causing hypocalcemia disrupts a transmembrane salt bridge to activate...

Area of Science:

• Endocrinology and Metabolism
• Molecular and Cellular Biology
• Biochemistry

Background:

• The calcium-sensing receptor (CaSR), a G protein-coupled receptor (GPCR), regulates extracellular calcium homeostasis through Gq/11 and Gi/o signaling pathways.
• CaSR mutations cause familial hypocalciuric hypercalcemia type 1 (FHH1) and autosomal dominant hypocalcemia type 1 (ADH1), demonstrating biased signaling.
• FHH1-associated mutations favor MAPK signaling, while ADH1 mutations enhance Ca2+i responses, indicating differential pathway activation.

Purpose of the Study:

• To investigate a newly identified ADH1-associated CaSR mutation, R680G.
• To elucidate the structural basis and signaling mechanisms underlying CaSR biased signaling.
• To identify novel CaSR signaling pathways independent of canonical G protein interactions.

Main Methods:

• Expression of CaSR variants in HEK 293 cells.
• Measurement of cytosolic calcium (Ca2+i) and mitogen-activated protein kinase (MAPK) signaling.
• Homology modeling and site-directed mutagenesis to analyze structural-functional relationships.

Main Results:

• The R680G CaSR mutation enhanced MAPK signaling without affecting Ca2+i responses, indicating biased signaling.
• This enhanced MAPK activity was mediated by a noncanonical pathway involving β-arrestin proteins, independent of Gq/11 and Gi/o.
• Disruption of an Arg680-Glu767 salt bridge in transmembrane domain 3 and extracellular loop 2, respectively, was identified as the structural basis for selective β-arrestin signaling.

Conclusions:

• This study demonstrates a novel β-arrestin-mediated signaling pathway for the CaSR.
• The Arg680-Glu767 salt bridge is crucial for mediating CaSR signaling bias.
• Understanding these biased signaling mechanisms provides new insights into calcium homeostasis regulation and related disorders.