THE ROLE OF INTRACELLULAR TRAFFIC IN HIV INFECTION

细胞内运输在 HIV 感染中的作用

• 批准号: 3917443
• 负责人: J A HANOVER
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3917443
• 关键词: cell nucleus; drug resistance; genetic manipulation; glycoproteins; human immunodeficiency virus; intracellular membranes; lymphocytic leukemia; membrane permeability; molecular cloning; plasmids; protein sequence; receptor; surface antigens; synthetic peptide; tissue /cell culture; transfection; virus infection mechanism

An investigation of CD4, the presumed T-cell receptor for the human immunodeficiency virus, has shown that surface expression of this protei is blocked by tunicamycin, a potent inhibitor of glycosylation, under conditions where alternate surface receptors are unaffected. Initial studies employing acute lymphoblastic leukemic cells have beer extended by the successful transfection of a plasmid containing the cDNA For CD4 into Chinese hamster ovary cells. Subsequent cotransfection with another plasmid containing the multiple drug resistant gene, has permitted the isolation of stable clones expressing large amounts of CD4. Preliminary data on this material indicates the presence in CD4 of complex or multi- antennary hybrid oligosaccharides. A targeting sequence has been identified which is sufficient to allow protein molecules as large as 0.5 x 10 to pass through the nuclear envelope. This has been assessed in our laboratory by chemically coupling peptides having the nuclear localization sequence to a highly fluorescent cytoplasmic protein. The sequence Pro-Lys-Lys-Lys-Arg-Lys-Va-Cys is sufficient to target these conjugates to the nucleus. The tat and art gene products of HIV have sequences very similar to this targeting domain. The approach we have taken is to chemically synthesize peptides corresponding to the tat sequence examined the ability of this peptide to confer nuclear localization. In addition to these studies, we have begun to examine the structure of the nuclear pore complex, across which the many nucleocytoplasmic exchange processes must occur. The structure of the pore complex is explored using recombinant DNA techniques. The major nuclear pore glycoprotein np62 has been cloned and a large segment sequenced.

CD 4是人类T细胞受体的研究 免疫缺陷病毒，已经显示了这种表面表达 蛋白酶被衣霉素阻断，衣霉素是一种有效的蛋白酶抑制剂， 糖基化，在交替的表面受体 不受影响。 采用急性淋巴细胞性 白血病细胞由于成功的转染而延长了寿命， 将含有CD 4 cDNA的质粒导入中国仓鼠卵巢 细胞 随后用另一种质粒共转染， 多重耐药基因，已经允许分离出 表达大量CD 4的稳定克隆。 初步数据 表明CD 4中存在复合物或多- 触角杂合寡糖。 已经鉴定了靶向序列，其足以 允许大至0.5 x 10的蛋白质分子通过 核膜 这已经在我们的实验室进行了评估， 具有核定位的化学偶联肽 高荧光细胞质蛋白的序列。 序列 Pro-Lys-Lys-Lys-Arg-Lys-Va-Cys足以靶向这些 与原子核结合。 HIV的达特和art基因产物 与这个靶向结构域非常相似。 的方法 我们采取的方法是化学合成 检测了这种肽赋予 核定位 除了这些研究，我们还开始 检查核孔复合体的结构， 许多核质交换过程必须发生。 的 利用重组DNA探索孔复合物的结构 技术. 主要的核孔糖蛋白np 62已经被 克隆并测序了一个大片段。