Molecular identification and functional expression of μ3, a novel alternatively spliced variant of the human μ opiate receptor gene
Molecular identification and functional expression of μ3, a novel alternatively spliced variant of the human μ opiate receptor gene
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DOI：
10.4049/jimmunol.170.10.5118
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发表时间：
2003-05-15
影响因子：
4.4
通讯作者：
Stefano, GB
中科院分区：
文献类型：
--
作者：
Cadet, P;Mantione, KJ;Stefano, GB
我们实验室的研究揭示了一种新的μ阿片受体，μ（3），它在血管组织和白细胞中表达。mu（3）受体对阿片类生物碱具有选择性，对阿片肽不敏感。我们现在使用mu（1）受体的441-bp保守区域在分子水平上鉴定mu（3）受体。分离的cDNA的序列分析表明，它是一种新的，选择性剪接变异的μ阿片受体基因。为了确定从该cDNA表达的蛋白质是否表现出预期的μ（3）受体的生化特征，在异源系统中表达cDNA克隆。在功能水平上，用μ（3）受体cDNA转染的COS-1细胞在用吗啡处理后表现出剂量依赖性的NO释放，而不是阿片肽（即，甲硫氨酸-脑啡肽）。纳洛酮能阻断吗啡对COS-1转染细胞的作用。未转染的COS-1细胞在吗啡或类似浓度的阿片肽存在下不产生NO。[H-3]二氢吗啡的受体结合分析进一步支持阿片类生物碱的选择性和阿片肽的不敏感性，这种受体。这些数据表明，这种新的μ阿片受体cDNA编码μ（3）阿片受体，因为它表现出已知是这种受体所特有的生化特性（阿片生物碱选择性和阿片肽不敏感性）。此外，使用北方印迹，RT-PCR，和序列分析，我们已经证明了这种新的亩变种在人血管组织，单核细胞，多形核细胞，和人神经母细胞瘤的表达。细胞
Studies from our laboratory have revealed a novel mu opiate receptor, mu(3), which is expressed in both vascular tissues and leukocytes. The mu(3) receptor is selective for opiate alkaloids and is insensitive to opioid peptides. We now identify the mu(3) receptor at the molecular level using a 441-bp conserved region of the mu(1), receptor. Sequence analysis of the isolated cDNA suggests that it is a novel, alternatively spliced variant of the mu opiate receptor gene. To determine whether protein expressed from this cDNA exhibits the biochemical characteristics expected of the mu(3) receptor, the cDNA clone was expressed in a heterologous system. At the functional level, COS-1 cells transfected with the mu(3) receptor cDNA exhibited dose-dependent release of NO following treatment with morphine, but not opioid peptides (i.e., Met-enkephalin). Naloxone was able to block the effect of morphine on COS-1 transfected cells. Nontransfected COS-1 cells did not produce NO in the presence of morphine or the opioid peptides at similar concentrations. Receptor binding analysis with [H-3]dihydromorphine further supports the opiate alkaloid selectivity and opioid peptide insensitivity of this receptor. These data suggest that this new mu opiate receptor cDNA encodes the mu(3) opiate receptor, since it exhibits biochemical characteristics known to be unique to this receptor (opiate alkaloid selective and opioid peptide insensitive). Furthermore, using Northern blot, RT-PCR, and sequence analysis, we have demonstrated the expression of this new mu variant in human vascular tissue, mononuclear cells, polymorphonuclear cells, and human neuroblastoma. cells.