TRANSFORMING DOMAIN OF HHV 6 THAT TRANSACTIVATES HIV 1

越来越多的数据表明HHV-6在肝硬化进展中的辅助因子作用。 艾滋病和艾滋病相关的恶性肿瘤。重要的是，HHV-6和HIV-1 可以共感染相同的CD 4+靶T细胞，导致刺激 HIV-1复制。 此外，HHV-6感染可诱导CD 4表达， 在CD 8 + T淋巴细胞中，从而使这些细胞允许HIV-1 感染我们的研究已经确定了一个3.9kbp的Sa/I-L DNA片段， HHV-6毒株U1102来自反式激活的直接重复区 人类免疫缺陷病毒1型-长末端重复序列（HIV-1 LTR）。 Sa/I-L还转化NIH 3 T3细胞并使原代仓鼠永生化 胚胎成纤维细胞反式激活和转化活动具有 共定位于1.9kbp的Sa/I-L-HindIII亚片段（命名为 Sa/I-L-SH），其含有357个氨基酸（aa）的开放阅读框架， （命名为ORF-1）。 ORF-1可以重新激活tat缺陷的HIV-1前病毒 提示ORF-1是HHV-6基因 在艾滋病进展过程中负责HIV-1再激活的puerectin vivo.纯化的ORF-1蛋白足以反式激活HIV-1 LTR。 为了扩展我们的观察，这项建议旨在剖析 转换所需的Sa/I-L-SH内的重要区域， 反式激活，并确定其相关性的背景下， 病毒在感染过程中 我们的具体目标是：1）描述 ORF-1反式激活HIV-1的机制以及最小的 HIV-1启动子内的HHV-6反式激活因子应答元件， 突变分析将特别强调ORF-1的作用 在整个病毒的背景下反式激活潜伏的HIV-1。（二） 表征Sa/I-L-SH片段内的最小转化区域 无论是作为一个独立的实体，还是在整个病毒的背景下， 体外细胞转化和致瘤性测定，和3）确定 艾滋病相关和非艾滋病淋巴细胞中ORF-1的存在和表达 Southern印迹、聚合酶链反应和原位杂交检测恶性肿瘤 杂交分析这些研究将提供重要的新 关于特定HHV-6基因的生物学相关性的信息， 艾滋病和艾滋病相关恶性肿瘤的进展， ORF-1的转化和反式激活结构域在细胞凋亡中的意义 病毒基因组的背景。

Accumulating data suggest a co-factor role for HHV-6 in the progression of AIDS and in AIDS-related malignancies. Of significance, HHV-6 and HIV-1 can co-infect the same CD4+ target T-cells resulting in the stimulation of HIV-1 replication. Moreover, HHV-6 infection can induce expression of CD4 in CD8+ T-lymphocytes, thus rendering these cells permissive for HIV-1 infection. Our studies have identified a 3.9 kbp Sa/I-L DNA fragment of HHV-6 strain U1102 from the direct repeat region which transactivates human immunodeficiency virus type 1 -long terminal repeat (HIV-1 LTR). Sa/I-L also transforms NIH 3T3 cells and immortalizes primary hamster embryo fibroblasts. The transactivating and transforming activities have been co-localized to a 1.9 kbp Sa/I-L-Hindlll subfragment, (designated Sa/I-L-SH) containing a 357 amino acid (aa) open reading frame, (designated ORF-1). ORF-1 can reactivate tat-defective HIV-1 provirus from latently infected cells, suggesting that ORF-1 is the HHV-6 gene putatively responsible for HIV-1 reactivation during AIDS progression in vivo. Purified ORF-1 protein is sufficient to transactivate the HIV-1 LTR. To extend our observations, this proposal is designed to dissect the essential regions within Sa/I-L-SH required for transformation and transactivation and to determine their relevance within the context of the whole virus during infection. Our specific aims are to: 1) Characterize the mechanism by which ORF-1 transactivates HIV-1 as well as the minimal HHV-6 transactivator responsive element(s) within the HIV-1 promoter by mutational analysis. Specific emphasis will be placed on the role of ORF-1 within the context of the whole virus to transactivate latent HIV-1. 2) Characterize the minimal transforming region within the Sa/I-L-SH fragment both as an independent entity and in the context of the whole virus by in vitro cell transformation and tumorigenicity assays, and 3) Determine the presence and expression of ORF-1 with AIDS-related and non-AIDS lymphoid malignancies by Southern blot, polymerase chain reaction, and in situ hybridization analyses. These studies will provide significant new information about the biologic relevance of specific HHV-6 gene(s) in the progression of AIDS and AIDS-related malignancies by determining the significance of the ORF-1 transforming and transactivating domains in the context of the viral genome.