Bioengineering new BRET-based biosensors to analyze GPCR signal transduction from different intracellular compartments

• Grant number: 15/09131-6
• Support Opportunities: Scholarships abroad - Research Internship - Post-doctor
• Start date: June 10, 2015
• End date: June 09, 2016
• Field of knowledge: Biological Sciences - Pharmacology - Biochemical and Molecular Pharmacology
• Principal Investigator: Claudio Miguel da Costa Neto
• Grantee: Lucas Tabajara Parreiras e Silva
• Supervisor: Michel Bouvier
• Host Institution: Faculdade de Medicina de Ribeirão Preto (FMRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Institution abroad: Université de Montréal, Canada
• Associated to the scholarship: 13/01261-2 - Involvement of biased agonism on trafficking and signaling of Angiotensin II AT1 receptor, BP.PD

Abstract

G protein-coupled receptors (GPCRs) constitute the largest family of signal transducer proteins in mammals and are considered a major target for drug discovery. After stimulation, GPCRs activate G proteins and then recruit ²-arrestins, which lead to disruption of G protein signaling and initiation of the internalization process. Although b-arrestin was first discovered for its role in interrupting G protein-mediated signaling it is now clear that the GPCR/²-arrestin complex can also lead to G protein-independent signaling and can persist even in the internalized endosomes. Recent advances in GPCR pharmacology led to the identification of ligands that preferentially activate one of these pathways that are known as "biased agonists". Moreover, although it is assumed that G protein signaling is halted upon GPCR internalization, very recent data suggest that G protein signaling may be also triggered by GPCRs localized at the endosomes and other intracellular compartments. These mechanisms may provide spatiotemporal regulation of signaling cascades with important consequences for normal and pathological responses and for the development of better drugs. However, very few tools are available to address that. In an effort to address these issues, the general aim of this study is to investigate the spatiotemporal regulation of this second round of G protein activation and its importance for cell physiology. We also want to address if biased agonists can also influence G protein activation from intracellular compartments with possible functional relevance. For this purpose, we will construct new BRET-based biosensors to evaluate G protein activation by GPCRs localized in distinct intracellular compartments. (AU)