Cellular Pharmacology of Kappa Opioid Receptor

Kappa 阿片受体的细胞药理学

• 批准号: 7064864
• 负责人: LEE-YUAN LIU-CHEN
• 金额: $ 33.07万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-05-01 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7064864
• 关键词: binding sites; biological signal transduction; cell line; genetic manipulation; guanine nucleotide binding protein; maleimides; microtubule associated protein; opioid receptor; pharmacology; phosphorylation; posttranslational modifications; protein localization; protein protein interaction; protein structure function; protein transport; receptor binding; receptor expression; sodium hydrogen exchanger; stimulant /agonist; transfection; transport proteins; tubulin; ubiquitin

DESCRIPTION (provided by applicant): Activation of (kappa) opioid receptors produces many effects, including analgesia, dysphoria and water diuresis and kappa agonists may be useful as analgesics, water diuretics and antipruritic drugs. Kappa opioid receptors are coupled through pertussis toxin-sensitive G proteins to affect a variety of effectors. Repeated or continuous administration of Kappa agonists leads to tolerance and dependence. Part of tolerance can be accounted for at the receptor level. The applicant's long term objectives are to understand at the molecular level the biochemical events that occur following chronic exposure to opioid drugs. After exposure to the selective agonist U50, 488H, the human kappa opioid receptor (hkor) undergoes internalization and the internalized receptors are recycled to plasma membranes or sorted to lysosomes and proteasomes for degradation. Whether there are signals directing the sorting of the internalized receptors is not well understood. The central hypothesis of this application is that the hkor is associated with EBP50/NHERF-1 and GABARAPL1 and undergoes ubiquitination, all of which play important roles in the regulation, trafficking and signaling of the receptor. The specific aims are as follows. (1) To investigate the interaction of the hkor with EBP50/NHERF-1, a PDZ domain-containing protein. The residues in the hkor involved in the interaction with EBP50/NHERF-1 will be determined by mutagenesis studies. Effects of receptor phosphorylation and ubiquitination on its association with EBP50/NHERF-1 will be assessed and the role of EBP50/NHERF in the U50,488H-induced stimulation of Na+,H+-exchange will be examined. (2) To examine the interaction of the hkor with GABARAPL 1. Whether there are interactions between GABARAPL1 and tubulin, N-ethylmaleimide-sensitive factor, EBP50/NHERF-1, rab5 or rab7 will be investigated. The regions of GABARAPL 1 and the hkor involved in the interaction will be determined. The functional significance of the hkor-GABARAPL1 interaction will be studied in terms of trafficking, surface expression, ligand binding and signal transduction of the hkor. (3) To delineate the role of ubiquitination in the signaling and trafficking of the hkor. Effects of different agonists and receptor phosphorylation on ubiquitination of the hkor will be examined. The sites of ubiquitination in the hkor will be determined by mutagenesis. The impact of ubiquitination on the functional properties, regulation and trafficking of the hkor will be investigated. Elucidation of the functional significance of EBP50/NHERF-1 and GABARAPL 1 and receptor ubiquitination in the trafficking and signaling of the hkor will provide better understanding of cell biology of the hkor and will have implications for development of kappa agonists and antagonists as therapeutic agents. In addition, such understanding will have important ramifications for other G protein coupled receptors since these molecules may play similar roles for other receptors.

描述（由申请人提供）：（KAPPA）阿片受体的激活产生许多作用，包括镇痛，吞咽困难和水利尿剂和Kappa激动剂，可能可作为镇痛药，水利尿剂和抗尿毒药物。 Kappa阿片受体通过百日咳毒素敏感的G蛋白偶联，以影响各种效应子。重复或连续给药kappa激动剂会导致耐受性和依赖性。可以在受体水平上解释一部分耐受性。申请人的长期目标是在分子水平上了解长期暴露于阿片类药物后发生的生化事件。在接触选择性激动剂U50，488H之后，人类Kappa阿片受体（HKOR）经历了内在化，并且内部化受体被回收至质膜或分类为溶酶体和蛋白酶，以降解。是否有指导内部受体分类的信号尚不清楚。该应用的中心假设是HKOR与EBP50/NHERF-1和GABARAPL1相关，并经历泛素化，所有这些都在受体的调节，运输和信号传导中起着重要作用。具体目标如下。 （1）研究HKOR与EBP50/NHERF-1（一种含PDZ结构域的蛋白质）的相互作用。与EBP50/NHERF-1相互作用的HKOR中的残基将由诱变研究确定。将评估受体磷酸化和泛素化对其与EBP50/NHERF-1关联的影响，并将检查EBP50/NHERF在U50,488H诱导的Na+，H+-Exchange中的作用。 （2）检查HKOR与Gabarapl 1的相互作用。是否将研究Gabarapl1和微管蛋白之间存在相互作用，N-乙基甲米酰胺敏感因子，EBP50/NHERF-1，RAB5或RAB7之间是否存在相互作用。将确定Gabarapl 1的区域和参与相互作用的HKOR。 HKOR-GABARAPL1相互作用的功能显着性将在运输，表面表达，配体结合和HKOR的信号转导方面进行研究。 （3）描述泛素化在HKOR的信号传导和贩运​​中的作用。将检查不同激动剂和受体磷酸化对HKOR泛素化的影响。 HKOR中泛素化的部位将由诱变确定。泛素化对HKOR的功能性能，调节和贩运的影响将得到研究。阐明EBP50/NHERF-1和GABARAPL 1的功能意义以及HKOR的贩运和信号传导中的受体泛素化将提供对HKOR的细胞生物学的更好理解，并将对Kappa激动剂和拮抗剂作为治疗剂的发展产生影响。此外，这种理解将对其他G蛋白偶联受体产生重要的影响，因为这些分子可能对其他受体起着相似的作用。