Mechanistics Analysis of Papillomavirus E2 Functions

描述(申请人提供)：乳头瘤病毒(PV)是一组小的DNA病毒，可在包括人类在内的各种高等脊椎动物中引起良性病变。某些乳头状瘤病毒，包括人类乳头瘤病毒(HPV)，如HPV16和HPV18，也与宫颈癌和其他人类肿瘤的发病机制有关。乳头瘤病毒建立持续感染，在这种感染中，病毒基因组作为自主复制的质粒在增殖的宿主细胞的细胞核中保持在低拷贝数。在牛乳头瘤病毒(BPV1)转化的小鼠细胞中，病毒基因组也以多拷贝核质粒的形式复制，这些核质粒可以在很长一段时间内保持细胞的转化状态。为了确保在感染和转化的细胞中持续存在，复制的病毒基因组必须在有丝分裂后被分割并维持在子细胞的核中。如果没有一种机制来确保核重组后病毒基因组在核膜内的定位，病毒基因组可能会留在细胞质中，并在细胞分裂后因降解或稀释而丢失。乳头瘤病毒E2蛋白是一种重要的调节蛋白，在病毒转录和病毒DNA复制中发挥关键作用，也是病毒基因组在分裂细胞中维持所必需的。在细胞分裂过程中，E2和PV基因组与有丝分裂染色体密切相关。最近，我们利用蛋白质组学的方法对体内与E2相关的细胞蛋白进行了系统的研究，发现Brd4(溴结构域包含蛋白4)是一种与E2相互作用的蛋白，并表明它是BPV1E2的有丝分裂染色体受体。这项研究拨款的重点是扩大对PVE2蛋白的研究，并研究E2与Brd4相互作用对病毒生命周期其他方面的功能意义。破坏E2与Brd4的结合可以阻断BPV1对细胞的转化以及E2和病毒DNA与有丝分裂染色体的结合。将进行化学遗传筛选，以确定E2/Brd4结合的小分子抑制剂是新的治疗性抗病毒化合物的潜在先导分子。

DESCRIPTION (provided by applicant): The papillomaviruses (PVs) are a group of small DNA viruses that induce benign lesions in variety of higher vertebrates, including humans. Certain papillomaviruses, including the human papillomaviruses (HPVs) such as HPV16 and 18, are also associated with the pathogenesis of cervical cancer and other human tumors. Papillomaviruses establish persistent infections in which the viral genomes are maintained as autonomous replicating plasmids at a low copy number in the nuclei of proliferating host cells. In bovine papillomavirus (BPV1) transformed mouse cells, the viral genomes also replicate as multicopy nuclear plasmids that can persist over long periods of time to maintain the transformation status of the cells. To ensure persistence in both infected and transformed cells, the replicated viral genomes must be partitioned and maintained in the nuclei of daughter cells following mitosis. Without a mechanism to ensure the localization of the viral genome within the nuclear envelope following nuclear reassembly, the viral genome could be left behind in the cytoplasm and lost either through degradation or dilution after cell division. The papillomavirus E2 protein, which is an important regulatory protein that plays critical roles in viral transcriptional and viral DNA replication, is also required for viral genome maintenance in dividing cells. E2, as well as the PV genomes, are closely associated with mitotic chromosomes in dividing cells. Utilizing a proteomic approach to systematically characterize cellular proteins that associate with E2 in vivo, we recently identified Brd4 (bromodomain-containing protein 4) as an E2 interacting protein and showed that it functions as the mitotic chromosome receptor for BPV1 E2. The focus of this research grant is to extend these studies on the PV E2 proteins and to examine the functional significance of the interaction of E2 with Brd4 to other aspects of the viral life cycle. Disruption of the binding of E2 to Brd4 can block the transformation of cells by BPV1 and the association of E2 and the viral DNA with mitotic chromosomes. Chemical genetic screens will be conducted to identify small molecule inhibitors of E2/Brd4 binding as potential lead molecules for novel therapeutic antiviral compounds.