Characterizing exon 11 promoter of the mu opioid receptor gene, OPRM

描述（由申请人提供）：本提案的长期目标是了解mu阿片受体（Oprm）基因的调控机制，并深入了解这种调控的药理学和生理学意义。早期的药理学研究已经提出了几种mu阿片受体亚型：mu1、mu2和吗啡-62-葡糖苷（M6G）。然而，目前只发现了一个mu阿片受体基因，这就提出了Oprm基因的选择性前mrna剪接和多个启动子可能负责多个mu阿片受体的可能性。在过去的十年中，我们已经从小鼠Oprm基因中鉴定出25个剪接变体，从大鼠Oprm基因中鉴定出13个剪接变体，从人类Oprm基因中鉴定出12个剪接变体。剪接变异的功能意义是由区域和细胞特异性表达、激动剂诱导的G蛋白偶联和受体内化的差异所支持的。通过分离一个新的启动子，外显子11启动子（E11启动子）进一步证明了Oprm基因的多样性。通过在大鼠和人类Oprm基因中鉴定这些小鼠同源物，也证实了E11相关变异和E11启动子的保守性。我们最近的敲入/敲除研究表明，在缺乏外显子11的小鼠中，M6G、6-乙酰吗啡和海洛因镇痛作用大大减弱，而吗啡的反应保持不变，这意味着外显子11相关剪接变异体和外显子11启动子在介导一类mu阿片类药物的作用中发挥了主要的功能作用。本提案将继续探索E11启动子的调控和功能，提出以下具体目标：1)。对小鼠E11启动子进行表征；2）. 研究人类E11启动子的结构和功能；3)。采用双基因靶向小鼠模型探讨E11和E1启动子的药理作用。本课题所获得的知识将有助于我们更好地了解Oprm基因调控的复杂性和功能重要性，建立基因靶向动物模型，研究该基因调控和功能的潜在机制，并为开发用于控制疼痛和滥用药物的新型药物提供潜在靶点。本研究的主要目的是进一步研究mu阿片受体（Oprm）基因中一个新的启动子——外显子11启动子的调控和功能。本课题所获得的知识将有助于我们更好地了解Oprm基因调控的复杂性和功能重要性，建立基因靶向动物模型，研究该基因调控和功能的潜在机制，并为开发用于控制疼痛和滥用药物的新型药物提供潜在靶点。

DESCRIPTION (provided by applicant): The long term goal of this proposal is to understand the mechanisms by which the mu opioid receptor (Oprm) gene is regulated and to gain insights into the pharmacological and physiological significance of this regulation. Early pharmacological studies have proposed several mu opioid receptor subtypes: mu1, mu2 and morphine-62-glucuronide (M6G). However, only one mu opioid receptor gene has been identified, raising the possibility that alternative pre-mRNA splicing and multiple promoters of the Oprm gene may be responsible for the multiple mu opioid receptors. Over the last ten years, we have identified 25 splice variants from the mouse Oprm gene, 13 splice variants from the rat Oprm gene and 12 from the human Oprm gene. The functional significance of the splice variants is supported by differences in regional and cell-specific expression, agonist-induced G protein coupling and receptor internalization. Diversity of the Oprm gene was further demonstrated by isolation of a new promoter, exon 11 promoter (E11 promoter). Conservation of the E11-associated variants and the E11 promoter has also been confirmed by identifying these mouse homologs in the rat and human Oprm genes. Our recent knockin/knockout study showed that in mice lacking exon 11, M6G, 6-acetylmorphine and heroin analgesia were greatly diminished, while morphine's response remained unchanged, implying a major functional role for exon 11-associated splice variants and exon 11 promoter in mediating the actions of a subset of mu opioids. This proposal will continue to explore the regulations and functions of E11 promoters by proposing the following specific aims: 1). To characterize the mouse E11 promoter; 2). To investigate structure and function of the human E11 promoter; 3). To explore the pharmacological function of E11 and E1 promoters using double gene targeting mouse model. The knowledge gained from this proposal will help us obtain a better understanding of the complexity and functional importance of Oprm gene regulation, establish the gene targeted animal models for studying the underlying mechanisms of this gene regulation and function, and provide potential targets for developing novel drugs used in control of pain and drug of abuse. The primary goal of this proposal is to further investigate the regulations and functions of a new promoter, exon 11 promoter, in the mu opioid receptor (Oprm) gene. The knowledge gained from this proposal will help us obtain a better understanding of the complexity and functional importance of Oprm gene regulation, establish the gene targeted animal models for studying the underlying mechanisms of this gene regulation and function, and provide potential targets for developing novel drugs used in control of pain and drug of abuse.