Endocytic mechanisms controlling functional selectivity of the CB1R

控制 CB1R 功能选择性的内吞机制

• 批准号: 8745061
• 负责人: Guillermo Ariel Yudowski
• 金额: $ 31.5万
• 依托单位: UNIVERSITY OF PUERTO RICO MED SCIENCES
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-01 至 2019-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8745061
• 关键词: Affect; Agonist; Anxiety Disorders; Arrestin Beta 1; Arrestins; Bar Codes; Biochemical; Cannabinoids; Cannabis; Cannabis Abuse; Cannabis-Related Disorder; Cell membrane; Cells; Clathrin; Dominant-Negative Mutation; Endocytosis; Family; Flow Cytometry; Fluorescence; Fluorescence Microscopy; Future; G-Protein-Coupled Receptors; GTP-Binding Proteins; Goals; Individual; Kinetics; Life; Ligands; MAPK8 gene; Marijuana; Marijuana Dependence; Measures; Mediating; Microscopy; Molecular; Neurons; Pharmaceutical Preparations; Phosphorylation; Phosphotransferases; Physiological; Population; Process; Proteins; Proto-Oncogene Proteins c-akt; Psychotic Disorders; Rattus; Receptor Activation; Receptor Signaling; Recruitment Activity; Regulation; Role; Signal Transduction; System; Tail; Techniques; Technology; Testing; Therapeutic; Time; Tissues; Translating; Western Blotting; Work; beta-arrestin; cannabinoid receptor; cellular imaging; coated pit; mutant; prevent; public health relevance; receptor; research study

DESCRIPTION (provided by applicant): G protein-coupled receptors (GPCRs) are one of the largest families of transmembrane receptors and a major target of current therapeutic drugs. At the signaling level, it has become clear that ligands acting on the same receptor can activate multiple and sometime opposing signaling cascades; a process defined as functional selectivity or biased agonism. One of the main effectors of functional selectivity are beta-arrestins, multifunction proteins recruited to activated receptors. However, how receptor activation translates into beta-arrestin signaling is not clearly defined. Our preliminary work combining state-of-the-art live cell imaging with molecular and biochemical techniques identifies ligand-specific dwell times, the time receptors are clustered into individual endocytic pits before endocytosis, as a mechanism by which receptors can control beta-arrestin mediated signaling. Our hypothesis suggests that ligands induce specific phosphorylations at the receptor level, eliciting specific endocytic dwell times during which beta-arrestins remain recruited and engaged in signaling. We propose to define a mechanism by which the Cannabinoid 1 Receptor (CB1R), one of the most abundant receptors in the CNS and target of cannabis, controls beta-arrestin signaling during endocytic dwell times. Our aims are: 1) Characterize ligand-specific dwell times of the CB1R to test our hypothesis that ligands can elicit specific dwell times that ar independent of their endocytic efficacy. 2) Define the mechanisms underlying ligand-specific dwell times of the CB1R. We will test the hypothesis that dwell times are controlled by ligand-specific phosphorylation profiles ("bar-codes") of the receptor. Alternative mechanisms will be also investigated. 3) Determine if beta-arrestin signaling is the physiological target of ligand specific dwell times in heterologous systems and native tissue. Finally, we will test different manipulations to control arrestin signaling by altering CB1R dwell times.

描述（由申请人提供）：G蛋白偶联受体（GPCR）是最大的跨膜受体家族之一，也是目前治疗药物的主要靶点。在信号传导水平上，已经清楚的是，作用于同一受体的配体可以激活多个并且有时是相反的信号传导级联;该过程被定义为功能选择性或偏向性激动。功能选择性的主要效应物之一是β-抑制蛋白，一种被激活受体募集的多功能蛋白质。然而，受体激活如何转化为β-抑制蛋白信号转导还没有明确定义。我们的初步工作结合了最先进的活细胞成像与分子和生物化学技术，确定配体特异性停留时间，时间受体聚集成单个内吞凹内吞作用之前，作为受体可以控制β-arrestin介导的信号传导的机制。我们的假设表明，配体在受体水平诱导特异性磷酸化，引发特定的内吞停留时间，在此期间，β-抑制蛋白保持招募和参与信号传导。我们建议定义一种机制，大麻素1受体（CB 1 R）是中枢神经系统中最丰富的受体之一，也是大麻的靶点，它在内吞停留时间内控制β-arrestin信号传导。我们的目标是：1）表征CB 1 R的配体特异性停留时间以测试我们的假设，即配体可以引起不依赖于其内吞功效的特异性停留时间。2)定义CB 1 R的配体特异性停留时间的机制。我们将检验驻留时间受受体的配体特异性磷酸化特征（“条形码”）控制的假设。还将研究替代机制。3)确定β-抑制蛋白信号传导是否是异源系统和天然组织中配体特异性停留时间的生理靶点。最后，我们将测试通过改变CB 1 R停留时间来控制arrestin信号传导的不同操作。