Regulatory mechanism of HIV-ltranscription and its therapeutic control

为了鉴定HIV编码的达特的细胞靶基因，我们进行了基因表达谱分析，并发现了OGG 1，该基因编码一种负责修复8-氧代-dG的酶，8-氧代-dG是一种由于放射性氧物质（ROS）作用而受损的鸟嘌呤残基。在位于OGG 1基因启动子内的as-regulatory元件中，我们已经确定AP-4位点负责Tat介导的OGG 1上调。当AP-4位点突变时，OGG 1基因表达上调，表明AP-4在OGG 1基因表达中起负作用，并且达特的作用被消除。通过染色质免疫沉淀（ChIP）实验，我们发现AP-4与OGG 1启动子上的这些AP-4位点结合，并且在达特转导后AP-4从OGG 1启动子上释放。我们还发现，达特结合AP-4的免疫沉淀，然后通过蛋白质印迹。当测量8-氧代-dG水平的量时，我们发现在达特转导后8-氧代-dG的显著降低。这些发现表明达特可能在维持HIV感染细胞和HIV前病毒DNA的基因组完整性方面发挥作用。我们还研究了一种新的IKK抑制剂的作用，该抑制剂主要参与信号诱导的潜伏感染细胞的HIV复制。通过刺激感染HIV的细胞，HIV的产生大大增加，用ACHP预处理细胞在非细胞毒性浓度下大大抑制了病毒的产生，IC 50和CC 50值分别为0.5 μM和15 μM（治疗窗约为30）。这些结果表明，ACHP及其衍生物可能具有治疗作用，以防止艾滋病的发展，通过阻止艾滋病毒复制从潜伏感染的细胞。

In an attempt to identify the cellular target genes for HIV-encocled Tat, we have performed gene expression profile analysis and fund OGG1, the gene encoding an enzyme responsible for the repair of 8-oxo-dG, an damaged Guanine residue due to the action of radieal oxygen species (ROS). Among the as-regulatory elements located within the promoter of OGG1 gene, we have identified AP-4 sites to be responsible for the Tat-mediated OGG1 upregulation. When AP-4 sites were mutated the OGG1 gene expression was upregulated, indicating that AP-4 plays a negative role in OGG1 gene expression, and the effect of Tat was abolished. Using chromatin immunoprecipitation (ChIP) assay we fund that AP-4 binds to these AP-4 sites on the OGG1 promoter and AP-4 was reliesed from the OGG1 promoter upon Tat transduction. We also fund that Tat binds to AP-4 by immunoprecipitation followed by Western blotting. When the amounts of 8-oxo-dG level were measured, we found the dramatic decrease of the 8-oxo-dG upon Tat transduction. These findings indicate a possibility that Tat plays a role in Maintaining the genomic integrity of the HIV-infected cells and HIV proviral DNA.We also examined the effect of a novel inhibitor of IKK that is primarily involved in the signal induced HIV replication from the latently infected cells. The HIV production was greatly augmented by stimulating cells infected with HIV and the pretreatment of cells with ACHP greatly inhibited the viral production under non-cytotoxic concentrations with IC 50 and CC50 values of 0.5 μM and 15 μM, respectively (therapeutic window being approximately 30). These findings suggest that ACHP and its derivatives may have therapeutic effect to prevent AIDS development by preventing HIV replication from the latently infected cells.