HCMV, CELL GROWTH STATE, AND TRANSCRIPTION FACTOR E2F

描述：病毒和宿主细胞因素的相互作用决定了 病毒感染的命运。阿齐兹汗博士一直在研究人类 巨细胞病毒感染及其即刻早期表达的研究 细胞周期/生长调节二氢叶酸还原酶基因 吉恩。E2F转录因子家族参与了基本的 Dhfr基因的转录和细胞周期调控。E2F绑定 72 kDa IE1基因产物反式激活DHFR所需的位点 (IE72)，E2F-1和E2F-2在细胞内和无细胞实验中与IE72相互作用。 对HCMV IE72和E2F进行了进一步的功能和分子分析 确定调节病毒复制的巨细胞病毒-宿主细胞相互作用。至 为此，他们将：(1)描述IE72及其E2F介导的作用 IE72在人巨细胞病毒感染中的作用 增殖、细胞周期动力学和细胞基因表达；以及(3) 人巨细胞病毒激活E2F依赖转录机制的研究 IE72。 为了确定IE72在人巨细胞病毒感染中的确切作用 相关的细胞反应，他们将决定病毒的效果 对E2F合成的影响及其与其他细胞蛋白的关系。 为了确定IE72与E2FS和激酶的相互作用 IE72的活性是病毒复制所必需的，显性阴性 排除这些活动的IE72突变体将被使用。实验以实现 解决第二个目标是利用稳定的IE72诱导细胞系 表情。IE72对宿主细胞增殖和基因表达的影响 宿主细胞生长状态在调节其反应中的表达和作用 将被确定为IE72。解决第三个目标的实验包括 建立体外转录系统，以确定是否存在 IE72和E2F依赖的转录效应。蛋白水解酶的作用 IE72的活性及其从特定口袋蛋白中解离E2F 在转录激活方面将得到解决。 总之，他们将E2F确定为HCMV-IE72靶标 以探讨E2F作为病毒作用的重要介体的作用。 这些实验的目的是要确定人巨细胞病毒是如何影响 细胞转录因子及其他参与细胞周期的因子 控制与宿主细胞感染性和两种细胞的激活有关 以及病毒DNA合成和基因表达。

DESCRIPTION: The interaction of viral and host cell factors determines the fate of viral infection. Dr. Azizkhan has been studying the effect of human cytomegalovirus (HCMV) infection and expression of HCMV-immediate early (IE) genes on the cell cycle/growth regulated dihydrofolate reductase (DHFR) gene. The E2F family of transcription factors are involved in basal transcription and cell cycle regulation of the DHFR gene. The E2F binding site is required for trans activation of DHFR by the 72 kDa IE1 gene product (IE72), and E2F-1 and -2 interact with IE72 in cells and in cell-free assay. Further functional and molecular analyses of HCMV IE72 and E2F are proposed to identify HCMV-host cell interactions that regulate viral replication. To this end, they will: (1) characterize the role of IE72 and its E2F-mediated effects in HCMV infection; (2) characterize the effects of IE72 on cell proliferation, cell cycle kinetics and cellular gene expression; and (3) determine the mechanism of activation of E2F-dependent transcription by HCMV IE72. In order to determine the precise role of IE72 in HCMV infection and the associated cellular response, they will determine the effect of viral infection on E2F synthesis and its association with other cellular proteins. To determine whether the interaction of IE72 with E2Fs and the kinase activity of IE72 are essential for viral replication, dominant negative mutants of IE72 that preclude these activities will be used. Experiments to address the second aim utilize a stable cell line with inducible IE72 expression. The effect of IE72 on host cell proliferation and gene expression and the role of host cell growth state in modulating its response to IE72 will be ascertained. Experiments to address the third aim involve establishing an in vitro transcription system to determine whether the effects of IE72 and E2F-dependent transcription. The role of the kinase activity of IE72 and its dissociation of E2F from specific pocket proteins in transcriptional activation will be addressed. In summary, their identification of E2F as a target of HCMV-IE72 directed them to explore the role of E2F as an essential mediator of virus effects. The goal of these experiments is to establish how the effects of HCMV on cellular transcription factors and other factors involved in cell cycle control relate to host cell infectivity and to activation of both cellular and viral DNA synthesis and gene expression.