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

用DNA损伤剂如紫外线辐射处理人类细胞， 可导致潜伏病毒的激活和病毒的增加 生产 这可能对疾病进展有重要意义 在治疗HIV-1感染者期间， 使用细胞毒性药物的患者。我们在遗传毒性应激方面的专业知识 反应的研究已经应用于HIV-1的反应研究。 这 该项目由NIH校内艾滋病靶向抗病毒项目资助 但只为裴博士提供了部分工资支持 在FY 93。 最初，我们专注于细胞毒性药物诱导的 稳定整合的HIV-启动子-CAT构建体。我们发现， 培养基中更多的细胞因子可以影响DNA损伤反应; 此外，我们还发现，由UV- 照射的细胞可以激活稳定整合的HIV-CAT反式。 紫外线照射可以诱导细胞Tat样因子，我们的研究表明， 参与HIV-1调节的细胞因子主要集中在 具有Tat样特性的蛋白质，我们已经通过一个 Northwestern blotting方法检测多种TAR RNA结合蛋白（TBP） 完整的核提取物。 TBP的水平被发现在 不同的细胞系，并在许多情况下DNA损伤诱导。 与 通过这种方法，我们已经将已知的TAR结合的数量增加了一倍多。 蛋白质，并首次证明了TBP对 应力 我们计划进一步描述这些TBP及其对 遗传毒性应激，包括细胞因子在其活化中的作用。 将开发纯化这些DNA损伤诱导RNA的方法， 结合蛋白，并最终分离cDNA克隆， 蛋白质序列测定。 这些蛋白质的鉴定将 这可能为HIV-1的调节和HIV-1的免疫调节提供重要的见解。 HIV-1感染的细胞对细胞毒性剂的反应。

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 may have important implications in disease progression both from environmental sources and during treatment of HIV-l-infected patients with cytotoxic agents. Our expertise in genotoxic stress responses has been applied to the study of responses in HIV-l. This project was funded by the NIH Intramural AIDS Targeted Antiviral Program for FY91 and FY92 but only partial salary support for Dr. Bae was provided in FY93. Initially, we focused on the induction by cytotoxic agents of stably-integrated HIV-promoter-CAT constructs. We have found that one or more cytokines in the medium can affect DNA-damage responsiveness; in addition, we have also found that a soluble factor(s) produced by UV- irradiated cells can activate stably-integrated HIV-CAT in trans. Since UV-irradiation can induce a cellular Tat-like factor, our search for cellular factors involved in HIV-l regulation has focused primarily on proteins with Tat-like properties, and we have identified by a Northwestern blotting approach a variety of TAR RNA-binding proteins (TBP) in whole nuclear extracts. The levels of TBP were found to vary in different cell lines, and were DNA-damage inducible in many cases. With this approach, we have more than doubled the number of known TAR-binding proteins and have shown the first demonstration of a TBP response to stress. We plan to further characterize these TBP and their responses to genotoxic stress including the role of cytokines in their activation. Approaches will be developed to purify these DNA-damage-inducible RNA- binding proteins and to ultimately isolate cDNA clones based on partial protein sequence determinations. Identification of these proteins will probably provide important insights into the regulation of HIV-l and the responses of HIV-l-infected cells to cytotoxic agents.