Constitutive Gαi Coupling Activity of Very Large G Protein-coupled Receptor 1 (VLGR1) and Its Regulation by PDZD7 Protein

Background: The signaling and regulatory mechanism of the orphan receptor VLGR1 remains elusive. Results: The cleaved VLGR1 -subunit constitutively coupled to G(i) and was regulated by the VLGR1 -subunit, a disease-associated mutation, and PDZD7. Conclusion: The VLGR1 -subunit signals independently and is regulated at multiple levels. Significance: The identified new signaling mechanism may aid in the design of a VLGR1-targeted therapy.The very large G protein-coupled receptor 1 (VLGR1) is a core component in inner ear hair cell development. Mutations in the vlgr1 gene cause Usher syndrome, the symptoms of which include congenital hearing loss and progressive retinitis pigmentosa. However, the mechanism of VLGR1-regulated intracellular signaling and its role in Usher syndrome remain elusive. Here, we show that VLGR1 is processed into two fragments after autocleavage at the G protein-coupled receptor proteolytic site. The cleaved VLGR1 -subunit constitutively inhibited adenylate cyclase (AC) activity through G(i) coupling. Co-expression of the G(iq) chimera with the VLGR1 -subunit changed its activity to the phospholipase C/nuclear factor of activated T cells signaling pathway, which demonstrates the G(i) protein coupling specificity of this subunit. An R6002A mutation in intracellular loop 2 of VLGR1 abolished G(i) coupling, but the pathogenic VLGR1 Y6236fsx1 mutant showed increased AC inhibition. Furthermore, overexpression of another Usher syndrome protein, PDZD7, decreased the AC inhibition of the VLGR1 -subunit but showed no effect on the VLGR1 Y6236fsx1 mutant. Taken together, we identified an independent G(i) signaling pathway of the VLGR1 -subunit and its regulatory mechanisms that may have a role in the development of Usher syndrome.