Biochemical analysis of papillomavirus replication

描述（由申请人提供）：本提案的总体目标是了解乳头瘤病毒如何复制其基因组。近年来，乳头瘤病毒已成为人类疾病中非常重要的致病因子。这些病毒作为其正常生命周期的一部分，感染并转化上皮细胞，引起良性肿瘤，良性肿瘤以低但显著的频率可以变成恶性肿瘤。更深入地了解生命周期，特别是DNA复制，对于了解疾病及其传播，并最终制定有效的治疗措施至关重要。我们正在研究乳头瘤病毒在体内和体外的DNA复制。我们正在对病毒DNA复制所需的病毒蛋白质和序列元件进行遗传、生物化学和结构分析。在本提案中，我们将继续深入分析乳头瘤病毒复制子，重点是负责病毒复制起点的有序识别、扭曲和解旋的组装途径。我们建议（i）表征从E12E22复合物到E1DT的转变。(ii)表征E1 DT并分析其向DH的过渡。(iii)研究E1 DH的性质并表征其活性。(iv)分析核提取物激活E1 DH的形成。公共卫生相关性：乳头瘤病毒是人类疾病的重要病原体。病毒DNA复制机制是药物治疗的少数潜在靶点之一。我们的研究，这是针对详细了解病毒DNA复制机制将提供新的潜在目标。

DESCRIPTION (provided by applicant): The overall goal of this proposal is to understand how papillomviruses replicate their genomes. Papillomaviruses have in recent years emerged as a very important causal agent in human disease. These viruses as part of their normal life cycle infect and transform cells in the epithelium causing benign tumors that with a low, but significant, frequency can become malignant. A deeper understanding of the life cycle in general, and DNA replication in particular, is of critical importance for the understanding of the disease, its transmission and ultimately for the development of effective therapeutic measures. We are studying DNA replication of papillomaviruses in vivo and in vitro. We are combining genetic biochemical and structural analyses of the viral proteins and sequence elements that are required for viral DNA replication. In this proposal we will continue our in depth analysis of the papillomavirus replicon with emphasis on the assembly pathways responsible for the ordered recognition, distortion and unwinding of the viral origin of replication. We propose to (i) characterize the transition from the E12E22 complex to the E1 DT. (ii) To characterize the E1 DT and to analyze its transition to the DH. (iii) To investigate the properties of the E1 DH and to characterize its activity. (iv) To analyze nuclear extract activated E1 DH formation. PUBLIC HEALTH RELEVANCE: Papillomaviruses are very important causative agents of human disease. The viral DNA replication machinery presents one of the few potential targets for drug therapy. Our studies, which are directed towards understanding the viral DNA replication machinery in detail will provide new such potential targets.