Cannabinoid receptors and associated proteins

总结 CB 1受体（CB 1 R）是用于治疗成瘾、神经退行性疾病和神经退行性疾病的治疗靶点， 疼痛管理，但基于CB 1 R的药用化合物受到限制。的职能 CB 1 R调节发育中的神经元过程，神经传递中的逆行信号传导，以及细胞 神经保护机制对脑功能至关重要。内源性大麻素配体2-花生四烯酸甘油 和花生四烯酸是突触活动的主要神经调节剂，但全面了解CB 1 R 在特定的细胞类型中，信号传导可以被相关的蛋白质调节，这一点刚刚开始被认识到。的 科学假设是，CB 1 R的CRIP 1a调节可以在结构和功能上理解。 功能水平，使得基于肽或小分子干预的药物设计可以靶向 CRIP 1a-CB 1 R相互作用。我们最近发表的研究表明，CRIP 1a减少了 细胞表面CB 1 R的密度，减弱激动剂依赖性但不是组成性内化 通过与β-arrestins竞争结合到C-末端位点，并减少新的- 在延长的WIN 55212 -2后，细胞表面合成CB 1 R，但不合成CP 55940。其他研究 证明CRIP 1a在调节CB 1 R细胞信号传导中具有关键作用， 从需要C-末端激活的Gi 3和G 0偶联到不需要C-末端激活的Gi 1和Gi 2。在未出版的 研究中，我们已经确定了高分辨率结构的X射线晶体学，并发现CRIP 1a是 肉豆蔻酰化或异戊二烯化蛋白质的载体家族成员。基于这一重大进展 在CRIP 1a的结构和功能的知识，我们假设CRIP 1a的功能是， 与CB 1 R-G-蛋白复合物相互作用的方式可以由G-α和/或G-γ调节 亚基特异性、磷酸化以及与其他已知 释放货物。我们拟研究N18 TG 2神经母细胞瘤细胞中CB 1 R相关蛋白 内源性表达CB 1 R及其相关蛋白的模型，以及体外实验 纯化的重组蛋白和由其衍生的肽。该项目的目的是调查： CRIP 1a与CB 1 R的相互作用; CRIP 1与G- CRIP 1a的功能通过货物释放蛋白和磷酸化调节。的 拟议调查的结果应该通过提供证据证明对该领域具有变革性， CB 1 R和相关的CRIP 1a相互作用以指导细胞信号传导途径。从这个角度来看，小说 肽和小分子可被开发为神经疾病的治疗剂， CB 1 R和CRIP 1a在神经元中共存。

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 In unpublished studies, we have determined the high resolution structure from X-ray crystallography, and found that CRIP1a is a member of the family of carriers for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.