Spatiotemporally resolved GPCR interactome uncovers unique mediators of receptor agonism
Maria M. Shchepinova, Rachel Richardson, Jack W. Houghton, Abigail R. Walker, Mohammed Safar, Daniel Conole, Aylin C. Hanyaloglu, Edward W. Tate
Abstract
Cellular signaling by membrane G protein-coupled receptors (GPCRs) is governed by a complex and diverse array of mechanisms. The dynamics of a GPCR interactome, as it evolves over time and space in response to an agonist, provide a unique perspective on pleiotropic signaling decoding and functional selectivity at the cellular level. In this study, we utilized proximity-based APEX2 proteomics to investigate the interaction network of the luteinizing hormone receptor (LHR) on a minute-to-minute timescale. We developed an analytical approach that integrates quantitative multiplexed proteomics with temporal reference profiles, creating a platform to identify the proteomic environment of APEX2-tagged LHR at the nanometer scale. LHR activity is finely regulated spatially, leading to the identification of putative interactors, including the Ras-related GTPase RAP2B, which modulate both receptor signaling and post-endocytic trafficking. This work provides a valuable resource for spatiotemporal nanodomain mapping of LHR interactors across subcellular compartments.