GENETIC INTERACTION OF HUMAN HERPESVIRUS-6 WITH HIV-1

人类疱疹病毒 6 (HHV-6) 或人类相互作用的研究 巨细胞病毒 (HCMV) 与人类免疫缺陷病毒 1 型 (HIV-1) 鉴于 HHV-6 或之间建议的关系感兴趣 CMV 感染和艾滋病的发病机制。 HHV-6 和 HIV-1 可以 高效感染 CD4 T 细胞，共感染加速细胞病变 影响。 进行研究是为了了解这些相互作用如何 发生在分子水平。 我们测试了可能影响的 HHV-6 基因 HIV-1 复制。 通过 DNA 转染到人类 T 细胞和猴子中 肾细胞，我们已经证明 HHV-6 转化基因片段， 本实验室先前鉴定出的 ZVH14 可以反式激活 HIV-1 LTR。 这种反式激活是通过 Sp1 结合位点介导的 在 HIV-1 长末端重复序列中。 115 个氨基酸的开放阅读框， ZVH14 DNA 中的 B115 显示出与 ZVH14。 体外表达研究发现 B115 基因产物 预测 Mr = 18 kDa。 该蛋白质的突变体是由 位点特异性诱变以确认其在反式激活中的作用。 一个 特定突变体从 5' 端删除了 5 个核苷酸 ORF 显示 HIV-1 LTR 反式激活显着减少。 这一观察结果支持了我们的发现：B115 编码反式激活蛋白 蛋白质。 体外表达的蛋白质或 B115 提取物 转染Cos-7（SV40转化猴肾细胞）细胞没有 在凝胶迁移分析中显示与 HIV-1 LTR 的任何结合。 在后续行动中 研究表明，B115 也可以反式激活 CMV 立即 人成纤维细胞和猴肾细胞的早期启动子。 Sp1结合 正如我们报道的那样，位点似乎介导了这种反式激活 HIV-1 LTR 激活。 这些数据表明 HHV-6 转化基因 ZVH14 片段编码一种反式激活蛋白，可以激活 来自异源启动子和 Sp1 结合位点的转录是 对于这种反式激活至关重要。 这项研究对于更好地了解 HHV-6 在 艾滋病和巨细胞病毒视网膜炎的发病机制。 该项目还将提供 开发新型抗病毒疗法和安全的知识和工具 用于艾滋病基因治疗的病毒和质粒载体。

Studies on interactions of human herpesvirus-6 (HHV-6) or human cytomegalovirus (HCMV) with human immunodeficiency virus type 1 (HIV-1) are of interest in view of the suggested relationship between HHV-6 or CMV infections and the pathogenesis of AIDS. HHV-6 and HIV-1 can productively infect CD4+T cells and coinfection accelerates cytopathic effects. Studies were conducted to understand how these interactions occur at the molecular level. We tested HHV-6 genes that may affect HIV-1 replication. By DNA transfection into human T-cells and monkey kidney cells, we have shown that an HHV-6 transforming gene segment, ZVH14, previously identified in this laboratory, can transactivate the HIV-1 LTR. This transactivation is mediated through Sp1 binding sites in the HIV-1 long terminal repeat. A 115-amino acid open reading frame, B115, in the ZVH14 DNA showed a similar transactivation capacity as ZVH14. In vitro expression studies found the B115 gene product of the predicted Mr = 18 kDa. Mutants of this protein were generated by site-specific mutagenesis to confirm its role in transactivation. A specific mutant having a deletion of 5 nucleotides from the 5' - end of the ORF showed significant reduction in transactivation of the HIV-1 LTR. This observation supports our finding that B115 encodes a transactivator protein. In vitro expressed protein or the extracts from B115 transfected Cos-7 (SV40 transformed monkey kidney cells) cells did not show any binding to the HIV-1 LTR in gel shift assays. In a follow up study, we have shown that B115 can also transactivate the CMV immediate early promoter in human fibroblasts and monkey kidney cells. Sp1 binding sites seem to mediate this transactivation as we have reported with the HIV-1 LTR activation. These data suggest that HHV-6 transforming gene segment, ZVH14, encodes a transactivator protein which can activate transcription from heterologous promoters and Sp1 binding sites are essential for this transactivation. This study is important to better understand the role of HHV-6 in the pathogenesis of AIDS and CMV retinitis. The project will also provide knowledge and tools for development of novel antiviral therapies and safe viral and plasmid vectors for gene therapy of AIDS.