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Insights into Activation Mechanisms of G Protein-Coupled and Atypical β-Arrestin-Coupled Chemokine Receptors

深入了解 G 蛋白偶联和非典型 β-抑制蛋白偶联趋化因子受体的激活机制

基本信息

批准号：
9899267
负责人：
Tracy M Handel
金额：
$ 45.2万
依托单位：
SCRIPPS RESEARCH INSTITUTE, THE
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-04-01 至 2023-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9899267
关键词：
Agonist Arrestins Binding Binding Proteins Biophysics CXCL12 gene CXCR4 gene Cardiovascular system Clinical Trials Coupled Couples Coupling Data Development Disease Docking Drug Targeting Fluorescence Fluorescence Spectroscopy G-Protein-Coupled Receptors GTP-Binding Protein alpha Subunits, Gs GTP-Binding Proteins Immunologic Surveillance Immunosuppression Impairment Inflammation Inflammatory Ligand Binding Ligands Malignant Neoplasms Medical Methods Modeling Molecular Conformation Monitor Mutagenesis Mutation Neoplasm Metastasis Output Pharmaceutical Preparations Pharmacology Phenotype Physiology Point Mutation Property Proteins Public Health Receptor Activation Research Role Sampling Signal Transduction Structural Models Structure Testing Therapeutic Time angiogenesis arrestin 2 beta-arrestin cancer therapy cell motility chemokine chemokine receptor conformational conversion drug action drug development improved inhibitor/antagonist innovation insight novel receptor receptor binding receptor function response side effect single molecule small molecule tumor growth virtual

项目摘要

The chemokine CXCL12 and its G protein-coupled receptor (GPCR), CXCR4, regulate cell migration during development, immune surveillance and inflammation in normal physiology. They are also notorious for their roles in disease, particularly cancer. Recently, the "atypical" chemokine receptor, ACKR3, was identified as a second receptor for CXCL12 that does not signal through G proteins but instead couples to β-arrestin. Like CXCR4, ACKR3 is expressed during development and up-regulated in cancer. Despite their medical importance, the mechanisms by which CXCR4 and ACKR3 are activated to elicit distinct functional responses are poorly understood. Biophysical, computational and mutagenesis studies have shown that CXCR4 and ACKR3 recognize CXCL12 in a structurally similar manner. However, activation of CXCR4 is sensitive to even single point mutations of the chemokine and the receptor-binding pocket, whereas virtually all ligands tested activate ACKR3. Thus, CXCR4 and ACKR3 appear to function by different mechanisms. We hypothesize that CXCR4 activation involves a precise network of residues that stabilize the active conformation of the receptor, whereas ACKR3 activation occurs by a “wedge-like” mechanism, such that whenever any ligand docks in the receptor-binding pocket, it activates by destabilizing the inactive conformation. We propose to use single- molecule fluorescence (SMF) spectroscopy to explore the conformational dynamics and different activation mechanisms of these two receptors. We will also investigate how ligands and effectors (G proteins and β- arrestin) control the conformations of CXCR4 and ACKR3 and thus their signaling responses. The underlying hypothesis is that GPCRs and ACRs are intrinsically dynamic, sampling multiple conformations, and that ligands and effectors mutually regulate each other to influence the receptor conformation and signaling output. Strong preliminary data support this hypothesis. Our central hypothesis will be pursued with three specific aims. 1: Establish SMF methods to monitor the conformational dynamics of CXCR4 and ACKR3 in real-time, and probe their mechanisms of activation. 2: Investigate structural mechanisms of ACKR3 activation and the allostery between agonist binding and β-arrestin coupling. 3: Investigate structural mechanisms of CXCR4 activation and the allostery between agonist binding and G protein coupling. The innovation of this proposal is that novel SMF methods will provide experimental information on receptor dynamics and allostery that cannot be obtained with other methods. Moreover, these approaches have never been applied to chemokine receptors and very little is known about the relationship between conformational dynamics and atypical receptor activation. The studies are significant because they will provide unique insights into the distinct activation mechanisms of two therapeutically important receptors, one that is a G protein-coupled receptor and one that is β-arrestin-coupled. Understanding how ligands and effectors control the conformational state and signaling output of these receptors should ultimately inform drug development.

趋化因子CXCL12及其G蛋白偶联受体（GPCR） CXCR4调节细胞迁移