Regulation of GPCR Recycling at the Plasma Membrane

质膜上 GPCR 回收的调控

• 批准号: 8432875
• 负责人: Guillermo Ariel Yudowski
• 金额: $ 22.98万
• 依托单位: UNIVERSITY OF PUERTO RICO MED SCIENCES
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-03-01 至 2016-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8432875
• 关键词: ADRB2 gene; Adrenergic Receptor; Agonist; Amino Acid Sequence; Area; Award; Behavior; Cell Surface Receptors; Cell membrane; Cell model; Cell surface; Cells; Characteristics; Complex; Docking; Endocytosis; Event; Family; Frequencies; G-Protein-Coupled Receptors; Image; Institution; Laboratories; Lateral; Life; Ligands; Measurement; Mediating; Molecular; Nature; Neuraxis; Neurons; Pathologic Processes; Pathway interactions; Pharmaceutical Preparations; Physiological; Physiological Processes; Positioning Attribute; Process; Proteins; Receptor Activation; Receptor Signaling; Recycling; Regulation; Relative (related person); Research; Signal Transduction; Sorting - Cell Movement; Specificity; Stimulus; Tail; Techniques; Testing; Therapeutic; Total Internal Reflection Fluorescent; Universities; Vesicle; Work; beta-2 Adrenergic Receptors; drug of abuse; genetic regulatory protein; insight; mu opioid receptors; novel strategies; receptor; receptor recycling; response

signals into the cell. GPCRs mediate many important signaling functions in the central nervous system (CMS) and mediate the effects of many therapeutic and abused drugs. A basic mechanism by which these receptors are regulated is by ligand-induced endocytosis. This process reduces the number of activated receptors at the cell surface therefore regulating receptor signaling. While endocytosis is strongly regulated by physiological ligands and drugs, it is generally believed that recycling is a nonregulated process. New evidence from our laboratory and others are challenging this 'default1 hypothesis of GPCR recycling. We have developed a technique to investigate single insertion events at the cell surface by combining live total reflection microscopy (TIRF) with a pH sensitive extracelullar tag fused to the extracelullar terminus of GPCRs. Our preliminary results indicates that receptor recycling is a tightly regulated process involved in the homeostatic response to extracelullar stimuli. Furtheremore, regulation of recycling is dependent on receptor activation, which ultimately decreases the frequency of recycling events reducing receptor number at the cell surface. I propose to extend and further develop this novel approach to examine: a) The characteristics of the vesicles mediating receptor recycling, b) Identify the machinery involved in the fusion process, and its possible regulation by receptor activity, c) Investigate if the regulation of receptor recycling is a general process for GPCRs, d) Define and investigate the functionality of newly inserted receptors and the possible existence of a molecular complex associated to reinserted receptors. This work will be perfomed using an experimentally advantageous heterologous cell model, and the results will be latter examined in hippocamapal neurons. By investigating the regulation of receptor insertion to the cell surface, our work has the potential to provide fundamental new insight to key cellular events likely to be relevant to a wide variety of physiological and pathological processes. Obtaining the K99/ROO award will be an extraordinary opportunity to transition from my postdoctoral studies to an independent position in a university/research institution.

信号进入细胞。GPCR介导中枢神经系统中许多重要的信号传导功能 (CMS)并介导许多治疗和滥用药物的作用。一种基本机制， 这些受体通过配体诱导的内吞作用来调节。这个过程减少了 激活细胞表面的受体，从而调节受体信号传导。虽然内吞作用是 由于受到生理配体和药物的强烈调节，通常认为再循环是一种非调节的 过程来自我们实验室和其他机构的新证据正在挑战这种“默认假设 GPCR回收。我们已经开发了一种技术来调查单插入事件的细胞 通过将活全反射显微镜（TIRF）与pH敏感的细胞外标签结合， GPCR的细胞外末端。我们的初步结果表明，受体再循环是一个紧密的， 参与对细胞外刺激的稳态反应的调节过程。此外，监管 循环的频率依赖于受体的激活，这最终会降低循环的频率。 减少细胞表面受体数量的事件。我建议扩展和进一步发展这部小说 检查方法：a）介导受体再循环的囊泡的特征，B）鉴定 参与融合过程的机制，及其通过受体活性的可能调节，c）研究是否 受体回收的调节是GPCR的一般过程，d）定义和研究 新插入的受体的功能和可能存在的分子复合物， 重新插入的受体。这项工作将使用实验上有利的异源细胞进行 模型，结果将在后面的海马神经元中进行检查。通过研究 受体插入细胞表面，我们的工作有可能提供基本的新见解的关键 可能与多种生理和病理过程相关的细胞事件。 获得K99/ROO奖将是一个非凡的机会，从我的博士后过渡 在大学/研究机构中获得独立职位。