Cannabinoid receptors and associated proteins

大麻素受体和相关蛋白

基本信息

批准号：
10205004
负责人：
ALLYN C HOWLETT
金额：
$ 63.05万
依托单位：
WAKE FOREST UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-08-15 至 2023-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10205004
关键词：
2-arachidonylglycerol Agonist Antibodies Arl proteins Arrestins Attenuated Binding Binding Proteins Binding Sites Brain C-terminal CNR1 gene Carrier Proteins Cell Density Cell membrane Cell model Cell surface Cells Cholates Co-Immunoprecipitations Complex Corpus striatum structure Coupling Crystallization Cyclic AMP Data Detergents Development Disease Management Drug Design Endocannabinoids Epilepsy Family member G-Protein-Coupled Receptors GTP Binding GTP-Binding Protein alpha Subunits, Gs GTP-Binding Proteins Go Alpha Subunit Hippocampus (Brain)Immunoprecipitation In Vitro Individual Intervention Investigation Knowledge Length Ligands Marijuana Membrane Molecular Motor Neurodegenerative Disorders Neuromodulator Neurons Pain Pain management Peptides Pertussis Toxin Pharmaceutical Preparations Phosphorylation Phosphorylation Site Process Protein Family Proteins Proteomics Publishing Receptor Signaling Recombinant Proteins Recombinants Regulation Reporting Resistance Resolution Role Self Medication Signal Pathway Signal Transduction Site Specificity Structure Surface Synapses Synaptosomes Testing Therapeutic Agents Variant X-Ray Crystallography addiction anandamide base cannabinoid receptor cannabinoid receptor interacting protein 1a cell type design experimental study genetic regulatory protein knock-down member mutant nervous system disorder neuroblastoma cell neuropathology neuroprotection neurotransmission novel overexpression peptidomimetics preference recombinant peptide response small molecule targeted treatment therapeutic target trafficking

项目摘要

Summary The CB1 receptor (CB1R) a therapeutic target for treatment of addictions, neurodegenerative disorders and pain management, but medicinal compounds based upon the CB1R have been limited. The functions of the CB1R to regulate neuronal processes in development, retrograde signaling in neurotransmission, and cellular mechanisms of neuroprotection are critical to brain function. Endocannabinoid ligands 2-arachidonoylglycerol and anandamide are the primary neuromodulators of synaptic activity, but the full understanding of how CB1R signaling can be regulated by associated proteins in specific cell types is just beginning to be appreciated. The Scientific Premise is that CRIP1a modulation of the CB1R can be understood at the structural and functional level such that drug design based on peptide or small molecule interventions can target the CRIP1a-CB1R interaction. Our recently published studies have demonstrated that CRIP1a reduces the density of cell surface CB1R, attenuates the agonist-dependent but not the constitutive internalization processes by competing with β-arrestins for binding to C-terminal sites, and curtails the trafficking of newly- synthesized CB1R to the cell surface after prolonged WIN55212-2 but not CP55940. Other studies demonstrated that CRIP1a has a critical role in regulating CB1R cellular signaling by altering the preference for coupling from Gi3 & Go, which require the C-terminus for activation, to Gi1 & Gi2, which do not. In unpublished studies, we have determined the high resolution structure from X-ray crystallography, and found that CRIP1a is a member of the family of carriers for myristoylated or isoprenylated proteins. Based upon this major advance in knowledge of the structure and function of CRIP1a, we hypothesize that the function of CRIP1a is to interact with the CB1R–G-protein complex in ways that can be regulated by G-alpha and/or G-gamma subunit specificity, phosphorylation, and interaction with other regulatory proteins that are known to release cargo. We propose to investigate the CB1R associated proteins in the N18TG2 neuroblastoma cell model which endogenously expresses the CB1R and its associated proteins, as well as in in vitro experiments of purified recombinant proteins and peptides derived therefrom. The aims of this project are to investigate: the interaction of CRIP1a with the CB1R; the structural and functional interaction of CRIP1 with G- proteins; and the regulation of CRIP1a function by cargo-releasing proteins and phosphorylation. The results of the proposed investigation should prove to be transformative for the field by providing evidence that CB1R and associated CRIP1a interact to direct cellular signaling pathways. From this understanding, novel peptides and small molecules could be developed as therapeutic agents for neurological diseases in which both CB1R and CRIP1a co-exist in neurons.

概括 CB1受体（CB1R）是治疗成瘾、神经退行性疾病和疼痛管理，但基于 CB1R 的药物化合物有限。 CB1R 调节发育中的神经元过程、神经传递中的逆行信号传导和细胞神经保护机制对大脑功能至关重要。和 anandamide 是突触活动的主要神经调节剂，但充分了解 CB1R 如何信号传导可以通过特定细胞类型中的相关蛋白进行调节才刚刚开始被认识到。科学前提是 CB1R 的 CRIP1a 调节可以从结构和功能水平，使得基于肽或小分子干预的药物设计可以针对我们最近发表的研究表明 CRIP1a 可以降低 CRIP1a-CB1R 相互作用。细胞表面 CB1R 的密度，减弱激动剂依赖性但不减弱组成性内化通过与 β-arrestins 竞争结合 C 端位点来进行处理，并减少新的贩运在延长 WIN55212-2 后，将 CB1R 合成到细胞表面，但 CP55940 没有。 CRIP1a 通过改变对 CB1R 的偏好在调节 CB1R 细胞信号传导中发挥关键作用从需要 C 端激活的 Gi3 和 Go 到不需要 C 端激活的 Gi1 和 Gi2，未发表。研究中，我们通过X射线晶体学确定了高分辨率结构，并发现CRIP1a是基于这一重大进展，它是肉豆蔻酰化或异戊二烯化蛋白质载体家族的成员。在了解 CRIP1a 的结构和功能后，我们发现 CRIP1a 的功能是以可由 G-α 和/或 G-gamma 调节的方式与 CB1R-G-蛋白复合物相互作用亚基特异性、磷酸化以及与已知的其他调节蛋白的相互作用我们建议研究 N18TG2 神经母细胞瘤细胞中的 CB1R 相关蛋白。内源表达CB1R及其相关蛋白的模型以及体外实验该项目的目的是研究： CRIP1a 与 CB1R 的相互作用；CRIP1 与 G- 的结构和功能相互作用；蛋白质；以及通过货物释放蛋白质和磷酸化调节 CRIP1a 功能。拟议调查的结果应证明对该领域具有变革性，提供证据表明 CB1R 和相关的 CRIP1a 相互作用以指导细胞信号传导途径。肽和小分子可以开发为神经系统疾病的治疗剂，其中 CB1R 和 CRIP1a 共存于神经元中。