RAF-1 ACTIVATES TRANSCRIPTION FROM THE HIV-LONG TERMINAL REPEAT

C-Raf-1的转化活性可通过缺失或 在蛋白质的N-末端的一半插入。使用共转染 实验中，我们已经证明了c-Raf-1激酶激活转录 来自HIV-LTR的病毒。激活转化活性的突变 (例如，c-Raf-BXB)也增加了细胞的反式激活潜能 Raf-I蛋白。潜伏前病毒的激活发生在细胞 用生理诱导剂或肿瘤促进剂刺激，如 12-0-十四酰-佛波醇-13-乙酸酯(TPA)我们已经开始了实验 目的是确定c-Raf-I是否参与TPA的诱导 HIV-LTR.我们最近观察到，与公布的数据一致， HIV-LTR的转录被TPA激活。我们探索了 C-Raf-I对显性负性诱导的依赖性 C-Raf-301突变体，携带单一氨基酸替代 (Lys-Trp)定位于Raf-1蛋白的ATP结合部位，发现 它能阻断TPA的诱导。C-Raf-301阻断TPA诱导的能力 强烈提示c-Raf-I在TPA诱导HIV-LTR中起重要作用。 显然，c-Raf-I是一种重要的信号转导介质。 膜到细胞核，我们已经证明了HIV-LTR是一种 被激活的c-Raf的目标。在许多受体系统中，细胞 增殖是最终结果，随后是受体的触发 通过100%的c-Raf-1分子的过度磷酸化 手机。HIV包膜与CD4受体的关联是一种 艾滋病病毒感染的关键一步。在CD4系统中，抗体 交联会导致1%到5%的瞬时磷酸化 细胞中的c-Raf-I分子，有丝分裂信号是 流产了。因此，c-Raf-I在艾滋病毒初期阶段的作用 感染可能与其在细胞增殖中的作用不同。

The transforming activity of c-Raf-1 can be activated by deletions or insertions in the N-terminal half of the protein. Using a cotransfection assay, we have demonstrated that c-Raf-1 kinase activates transcription from the HIV-LTR. Mutations that activate the transforming activity (e.g., c-Raf-BXB) also increase the transactivation potential of the Raf-I protein. Activation of latent provirus occurs when cells are stimulated with physiological inducers or tumor promoters such as 12-0-tetradecanoyl-phorbol-13-acetate (TPA). We have begun experiments aimed at determining if c-Raf-I is involved in TPA induction of the HIV-LTR. We have recently observed, in agreement with published data, that transcription from the HIV-LTR is activated by TPA. We explored the c-Raf-I dependence of this induction utilizing a dominant negative mutant, c-Raf-301, which carries a single amino acid substitution (LYS-TRP) in the putative ATP binding site of Raf-1 kinase and found that it blocks TPA induction. The ability of c-Raf-301 to block TPA induction strongly suggests that c-Raf-I is mediating TPA induction of the HIV-LTR. Clearly, c-Raf-I is an important mediator of signal transduction from the membrane to the nucleus, and we have demonstrated that the HIV-LTR is a target for activated c-Raf. In many receptor systems where cell proliferation is the end result, triggering of the receptor is followed by hyper-phosphorylation of 100 percent of the c-Raf-1 molecules in the cell. Association of the HIV envelope with the CD4 receptor is an essential step in HIV infection. In the CD4 system, antibody cross-linking results in a transient phosphorylation of 1 to 5 percent of the c-Raf-I molecules in the cell, and the mitogenic signalling is abortive. Thus, the role of c-Raf-I in the initial stages of HIV infection may be different from its role in cellular proliferation.