THE EFFECTS OF STRESS RESPONSE GENES ON THE REGULATION OF HIV-1 GENE EXPRESSION

该项目将通过以下方式检查HIV-1基因表达的激活 DNA损伤剂和其他压力。对人体细胞的处理 DNA损伤剂，如紫外线辐射，可导致激活 潜伏的病毒和增加的病毒产量。此激活是 可能由多种机制介导，包括诱导一种 细胞类TAT因子与至少一种和可能的诱导 更多与HIV启动子中的特定元件结合的蛋白质。我们 计划应用我们在DNA损伤诱导(DDI)基因和 它们的启动子研究参与诱导的机制 HIV-1启动子。该项目的一部分将涉及使用 不同的HIV启动子构建融合到报告基因，如 氯霉素乙酰转移酶(CAT)。这些构造将是 稳定整合在人类细胞中，以及各种缺失和 将对启动子的突变进行分析。各种因素的影响 将使用诸如激酶抑制剂之类的抑制剂来描绘 不同的激酶和其他蛋白质的作用。实验，如 条带移位分析将被用来鉴定结合因子； 与转录结合的各种序列的竞争实验 因素，包括我们最近拥有的新的DDI元素 将采取行动，初步确定共同因素。 不同类型的DNA损伤剂对诱导的影响将是 研究：这可能为参与的机制提供证据 诱导，并且在计划治疗时也可能具有临床重要性 适用于患有恶性肿瘤的HIV感染患者。

This project will examine the activation of HIV-1 gene expression by DNA-damaging agents and other stresses. Treatment of human cells with DNA-damaging agents, such as UV radiation, can lead to the activation of latent virus and to increased virus production. This activation is probably mediated by multiple mechanisms including the induction of a cellular Tat-like factor and the induction of at least one and probably more proteins that bind to specific elements in the HIV promoter. We plan to apply our expertise with DNA-damage-inducible (DDI) genes and their promoters to study the mechanisms involved in the induction of the HIV-1 promoter. One portion of this project will involve the use of various HIV promoter constructs fused to reporter genes such as chloramphenicol acetyltransferase (CAT). These constructs will be stably integrated in human cells and the effect of various deletions and mutations in the promoter will be analyzed. The effect of various inhibitors such as kinase inhibitors will be employed to delineate the role of different kinases and other proteins. Experiments such as band-shift assays will be employed to identify binding-factor elements; competition experiments with various sequences, which bind transcription factors, including novel DDI elements, which we have recently identified, will be undertaken to tentatively identify common factors. The effect of various types of DNA-damaging agents on induction will be studied: this may provide evidence for the mechanisms involved in induction and may also be of clinical importance when planning therapy for HIV-infected patients with malignancies.