Heteromerization of angiotensin receptors changes trafficking and arrestin recruitment profiles |
| |
Authors: | Porrello Enzo R Pfleger Kevin D G Seeber Ruth M Qian Hongwei Oro Cristina Abogadie Fe Delbridge Lea M D Thomas Walter G |
| |
Institution: | a Department of Physiology, The University of Melbourne, Victoria 3010, Australiab Baker IDI Heart and Diabetes Institute, Victoria 8008, Australiac School of Biomedical Sciences, The University of Queensland, Queensland 4072, Australiad Laboratory for Molecular Endocrinology-GPCRs, Western Australian Institute for Medical Research (WAIMR) and Centre for Medical Research, University of Western Australia, WA 6009, Australiae Dimerix Bioscience Pty Ltd, Nedlands, Perth, WA 6009, Australia |
| |
Abstract: | The cardiovascular hormone angiotensin II (AngII) exerts its actions via two G protein-coupled receptor (GPCR) subtypes, AT1 and AT2, which often display antagonistic functions. Methodological constraints have so far precluded detailed analyses of the ligand-dependency, cellular localization, and functional relevance of AngII receptor interactions in live cells. In this study, we utilize a protein-fragment complementation assay (PCA) and GPCR-Heteromer Identification Technology (GPCR-HIT) to provide the first detailed investigation of the ligand-dependency and cellular localization of AngII receptor interactions in human embryonic kidney 293 cells. Fluorescent-tagged receptor constructs for PCA and GPCR-HIT displayed normal affinity and selectivity for AngII (AT1: IC50 = 1.0-1.6 nM; AT2: IC50 = 2.0-3.0 nM). Well-characterized angiotensin receptor interactions were used as positive and negative controls to demonstrate the sensitivity and specificity of these fluorescence-based assays. We report that AT1-AT2 receptor heteromers form constitutively, are localized to the plasma membrane and perinuclear compartments, and do not internalize following AngII stimulation despite arrestin being recruited specifically to the heteromer. Our findings using novel fluorescence-based technologies reveal a previously unrecognized mechanism of angiotensin receptor cross-talk involving cross-inhibition of AT1 receptor internalization through heteromerization with the AT2 receptor subtype. |
| |
Keywords: | PCA Protein-fragment Complementation Assay GPCR-HIT G protein-coupled receptor Heteromer Identification Technology |
本文献已被 ScienceDirect PubMed 等数据库收录! |
|