Visualization of the functional rescuing of GPCR homo/hetero-dimer by “silent” α 1B adrenoceptor FRET sensor

Abstract

Previous studies have demonstrated that the co-expression of two mutant G protein-coupled receptors (GPCRs) can potentially restore their impaired functionality. In this study, we investigated the rescue of functional deficits in homo- and heterodimers of α1B-adrenergic receptors (ARs) using a combination of biophysical and biochemical approaches. Initially, we constructed an Förster resonance energy transfer (FRET) sensor for α1B-AR that lacked ligand-binding capability (referred to as “silent”), and a mutant α1B-AR incapable of downstream activation (referred to as “defective”). Our results revealed that the co-expression of the “defective” and “silent” α1B-ARs led to the activation of the “silent” sensor and restoration of functional impairments in both receptors. Similarly, we conducted experiments involving the co-expression of a “defective” CXCR4 receptor lacking downstream signaling and G-protein recruitment with the “silent” α1B-AR FRET sensor, which also resulted in the rescue of their functional deficiencies. These findings highlight that upon dimerization with the “defective receptor, the “silent” receptor undergoes agonist-like structural changes, leading to functional restoration. This study provides direct evidence for the functional complementation mechanism mediated by GPCR dimers and offers new insights into the intervention strategies targeting GPCR defect-related diseases.