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）：阿片类神经传递涉及的内源性阿片类药物是由5种或更多氨基酸组成的肽。通常认为，从突触中清除这些肽，因此神经传递的终止是由肽酶介导的。最近，我们确定了针对阿片类肽的新型钠（Na）和氯（Cl）耦合转运蛋白。在这类神经递质的文献中尚未描述这种转运蛋白。该转运蛋白的表达在大鼠脑和人类神经元细胞系中都是可证明的。该转运蛋白的特征与先前已知的其他神经递质转运蛋白的特征相似。我们假设该转运蛋白代表了阿片神经传递的一种新的，迄今未识别的成分，因此是阿片类药物功能和阿片类药物成瘾的关键决定因素。 TAT是由人免疫缺陷病毒HIV-1编码的反式激活蛋白，明显上调了该转运蛋白在细胞系中。由于阿片类药物具有免疫调节功能并影响艾滋病的发病机理和进展，因此我们预测TAT诱导的阿片类肽转运蛋白表达的变化可能与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，但目前在负责转运功能的蛋白质的分子性质上尚无任何人知道。在这里，我们建议使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，我们将使用具有哺乳动物细胞和爪蟾卵母细胞的异源表达系统来阐明转运蛋白的功能特征。转运蛋白的成功克隆将导致基因的分子身份和负责新型运输功能的蛋白质。这将有助于开发有关对完整动物和培养细胞中转运蛋白的表达和定位进行分子研究所必需的核苷酸探针和抗体的开发。我们还将比较转运蛋白在对照小鼠和TAT-转基因小鼠中的表达和活性，以评估HIV-1 TAT对完整动物阿片类药物功能的影响。这些研究将导致发现先前未表征的新型蛋白质，该蛋白可能不仅在阿片类生物学和阿片类药物的领域，而且在HIV-1感染的进展和阿片类药物的进展中都起着关键作用。