Molecular Analysis of a Novel Opioid Peptide Transporter

新型阿片肽转运蛋白的分子分析

• 批准号: 7127794
• 负责人: VADIVEL GANAPATHY
• 金额: $ 14.65万
• 依托单位: AUGUSTA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-01 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7127794
• 关键词: AIDS; cell line; human; infection; lead; neural transmission; neurotransmitters; peptides; proteins

DESCRIPTION (provided by applicant): Endogenous opioids involved in opioidergic neurotransmission are peptides consisting of 5 or more amino acids. It is generally assumed that clearance of these peptides from the synapse and consequent termination of neurotransmission are mediated by peptidases. Recently, we have identified a novel sodium (Na)-and chlorine (Cl)-coupled transporter that is specific for opioid peptides. Such a transporter has not been described in the literature for this class of neurotransmitters. The expression of this transporter is demonstrable in the rat brain and in human neuronal cell lines. The characteristics of this transporter are similar to those of previously known other neurotransmitter transporters. We hypothesize that this transporter represents a new, hitherto unrecognized, component of opioidergic neurotransmission and hence a key determinant of opioid function and opiate addiction. Tat, the transactivator protein coded by the human immunodeficiency virus HIV-1, markedly up-regulates this transporter in cell lines. Since opioids exhibit immunomodulatory function and influence the pathogenesis and progression of AIDS, we predict that Tat-induced changes in the expression of the opioid peptide transporter may have clinical relevance to patients with HIV-1 infection. Even though the functional evidence for the existence of the transporter is unequivocal, nothing is known at present on the molecular nature of the protein responsible for the transport function. Here we propose to clone the transporter using functional expression strategy and establish the molecular identity of the transporter. We will then elucidate the functional features of the transporter using heterologous expression systems with mammalian cells and Xenopus oocytes. Successful cloning of the transporter will lead to the molecular identity of the gene and the protein responsible for the novel transport function. This will facilitate development of tools such as nucleotide probes and antibodies necessary to perform molecular studies on the expression and localization of the transporter in intact animals and in cultured cells. We will also compare the expression and activity of the transporter in control mice and in Tat-transgenic mice to assess the influence of HIV-1 Tat on opioid function in intact animals. These studies will lead to the discovery of a previously uncharacterized novel protein which might function as a key player not only in the area of opioid biology and opiate addiction but also in the progression of HIV-1 infection and AIDS in opiate addicts.

描述（由申请人提供）：参与阿片能神经传递的内源性阿片样物质是由5种或更多氨基酸组成的肽。一般认为，这些肽从突触的清除和随后的神经传递终止是由肽酶介导的。最近，我们已经确定了一种新的钠（Na）和氯（Cl）偶联转运体，它是特异性的阿片肽。这种转运体在这类神经递质的文献中尚未被描述。该转运体在大鼠脑和人类神经细胞系中均有表达。这种转运蛋白的特性与以前已知的其他神经递质转运蛋白相似。我们假设这种转运体代表了一种新的，迄今为止未被认识的阿片能神经传递成分，因此是阿片功能和阿片成瘾的关键决定因素。结果，由人类免疫缺陷病毒HIV-1编码的反激活蛋白在细胞系中显著上调该转运蛋白。由于阿片类药物具有免疫调节功能并影响艾滋病的发病和进展，我们预测tat诱导的阿片类肽转运体表达的变化可能与HIV-1感染患者具有临床相关性。尽管转运蛋白存在的功能证据是明确的，但目前对负责转运功能的蛋白质的分子性质一无所知。在此，我们提出利用功能表达策略克隆转运蛋白，并建立转运蛋白的分子身份。然后，我们将利用哺乳动物细胞和非洲爪蟾卵母细胞的异种表达系统阐明转运蛋白的功能特征。转运体的成功克隆将导致基因和负责新转运功能的蛋白质的分子身份。这将促进诸如核苷酸探针和抗体等工具的开发，这些工具用于在完整动物和培养细胞中对转运体的表达和定位进行分子研究。我们还将比较该转运体在对照小鼠和Tat转基因小鼠中的表达和活性，以评估HIV-1 Tat对完整动物阿片功能的影响。这些研究将导致发现一种以前未被表征的新蛋白，该蛋白可能不仅在阿片样物质生物学和阿片成瘾领域发挥关键作用，而且在阿片成瘾者中HIV-1感染和艾滋病的进展中发挥关键作用。