Molecular Mechanism Of Very Large G Protein Coupled Receptor (VLGR1) Signal Transduction And Its Function

The very large G protein coupled receptor (VLGR1) is a member of adhesion receptors or large N-terminal family B-7 transmembrane helixes (LNB7TM) receptors within the seven trans-membrane receptor superfamily. VLGR1 is the largest GPCR identified to date; its mRNA spans 19kb and encodes 6300 amino acids. VLGR1 is a core component of ankle-link complex in inner ear hair cells. Knock-out and mutation mouse models show that loss of VLGR1 function leads to abnormal stereociliary development and hearing loss, indicating crucial roles of VLGR1 in hearing transduction or audiroty system development. Over the past 10 or so years, human genetics data suggested that VLGR1 mutations cause Usher syndromes and seizures. Although significant progresses have been made, the details of VLGR1’s function in hair cells, its signaling cascade, and the mechanisms underlying causative effects of VLGR1 mutations in human diseases remain elusive and ask for further investigation.In our study, we show that VLGR1 is processed into two fragments after autocleavage at the G-protein-coupled receptor proteolytic site (GPS). The cleaved VLGR1 β-subunit constitutively inhibited adenylate cyclase (AC) activity through Gαi coupling. Co-expression of the Gαiq chimera with the VLGR1 β-subunit changes its activity to the PLC/NFAT signaling pathway, which demonstrates the Gαi protein-coupling specificity of this subunit. A R6002A mutation in intracellular loop 2 (ICL2) of VLGR1 abolished Gαi 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 β-subunit but showed no effect on the VLGR1 Y6236fsxl mutant. Taken together, we identified an independent Gα1; signaling of the VLGR1 J3-subunit and its regulatory mechanisms, which may have a role in the development of Usher syndrome.