The Role of Caspase Activation in the Differentiation-Dependent Life Cycle of HPV

高危人乳头瘤病毒(HPV)是宫颈癌的病原体。HPV的生命周期是 与其宿主细胞角质形成细胞的分化状态有关。差异化触发生产阶段 包括病毒基因组扩增、晚期基因表达和病毒粒子产生。在……里面 与正常细胞在分化后退出细胞周期不同，HPVE6和E7的表达 蛋白质主要通过降解P53和pRb来维持细胞周期中的一部分细胞的活动， 这个项目的长期目标分别是了解调节激活的机制 依赖于分化的病毒事件。为此，我已经证明了HPV31刺激低谷 分化时半胱氨酸天冬氨酸酶的激活水平，这是线粒体凋亡途径的特征 并且是通过切割病毒复制蛋白E1来扩增病毒基因组所必需的。在……里面 此外，我最近发现，HPV诱导了一种ATM依赖的DNA损伤反应 分化细胞，这是有效的病毒基因组扩增和caspase激活所必需的 通过自动柜员机目标Chk2。基于这一发现，caspase激活以及Chk2激活可以 被E6和E7刺激，并鉴定了几种HPV蛋白中的caspase裂解位点，即 假设HPV蛋白E6和E7通过DNA损伤反应诱导caspase激活 激活后期病毒事件。检验这一假设的具体目的是：1)确定机制(S)，通过 HPV通过检测E6和E7对DNA损伤的影响来激活分化细胞中的caspase 线粒体膜完整性的反应和改变，以及通过执行突变分析 确定Chk2激活和caspase激活所需的结构域；2) 确定HPV蛋白的caspase裂解如何在生命的分化依赖事件中起作用 通过体外循环和体内循环研究。E6和E7介导细胞活化的途径 导致caspase激活的DNA损伤反应将为HPV如何调控细胞凋亡提供洞察力 机器不仅促进了病毒的生命周期，而且可能促进了病毒的转化。

High-risk human papillomaviruses (HPV) are the causative agents of cervical cancer. The life cycle of HPV Is tied to the differentiation status of its host cell, the keratinocyte. Differentiation triggers the productive phase of the life cycle, which includes viral genome amplification, late gene expression and virion production. In contrast to normal cells, which exit the cell cycle upon differentiation, expression of the HPV E6 and E7 proteins maintain a subset of cells active in the cell cycle largely through degradation of p53 and pRb, respectively The long-term goal of this project is to understand the mechanisms that regulate the activation of differentiation-dependent viral events. Toward this end, I have demonstrated that HPV31 stimulates a low level of caspase activation upon differentiation that is characteristic of the mitochondrial apoptotic pathway and necessary for amplification of viral genomes through cleavage of the viral replication protein E1. In addition, I have recently found that HPV induces an ATM-dependent DNA damage response in differentiating cells, which is necessary for efficient viral genome amplification, as well as caspase activation through the ATM target Chk2. Based on the finding that caspase activation, as well as Chk2 activation, can be stimulated by both E6 and E7, and the identification of caspase cleavage sites in several HPV proteins, I hypothesize that the HPV proteins E6 and E7 induce caspase activation through a DNA damage response to activate late viral events. Specific aims to test this hypothesis are: 1) Identify the mechanism(s) by which HPV activates caspases in differentiating cells by examining the effect of E6 and E7 on the DNA damage response and alteration of mitochondrial membrane integrity, as well as by performing a mutational analysis of E6 and E7 to identify domains that are necessary for Chk2 activation and caspase activation; 2) Determine how caspase cleavage of HPV proteins contributes to differentiation-dependent events in the life cycle by using in vitro and in vivo studies. Delineating the pathway by which E6 and E7 mediate activation of a DNA damage response to lead to caspase activation will offer insight as to how HPV modulates apoptotic machinery to facilitate not only the viral life cycle, but possibly transformation as well.