Cytomegalovirus UL80 Proteins:Interactions and Modifications that Impact Function

描述(申请人提供)：我们建议回答有关分子相互作用的问题，这些问题指导着传染性巨细胞病毒形成的最早步骤。我们的重点是由开放阅读框UL80a编码的两种基因相关蛋白，称为组装蛋白前体(Pap)和衣壳成熟蛋白前体(PPR)。这些蛋白质相互作用，并与其他蛋白质相互作用，协调新生的前衣壳的组装和成熟--这是产生传染性病毒所必需的过程。我们最近克服了一个溶解性问题，这使得在体外使用纯化的蛋白质变得复杂，并将应用这种新的方法来寻找线索和测试基于试点研究的令人鼓舞的结果的模型。我们将通过三个特定的目标来做到这一点：(I)测试预测的磷酸化对Pap和PPR关键相互作用的影响；(Ii)验证“羧基盘绕结构域”在Pap和PPR自我相互作用中的差异优势，并确定其生物学意义和相关性；以及(Iii)确定Pap和PPR中是否需要CMV特异性的第二核定位信号(NLS2)来与衣壳门户蛋白(PUL104)相互作用。从这项工作中获得的信息将有助于确定在衣壳形成和成熟过程中驱动UL80蛋白结合和解离的变化序列，并有助于开发破坏(例如用于治疗)或利用(例如基因/药物输送载体)疱疹病毒组装的新策略的长期目标。 与公共卫生相关：巨细胞病毒是一种疱疹病毒群病毒，它对免疫系统薄弱的人构成威胁，包括非常年轻和非常年长的人。这项研究试图了解两种基因相关的蛋白质如何指导形成传染性病毒的最早步骤。所获得的信息将与驱动衣壳形成的分子相互作用有关，将有助于确定病毒形成的生物学机制，并将有助于确定从治疗上干预这一过程的新方法。

DESCRIPTION (provided by applicant): We propose to answer questions about molecular interactions guiding the earliest steps in formation of infectious cytomegalovirus. Our focus is on two genetically related proteins encoded by open reading frame UL80a, called the assembly protein precursor (pAP) and the capsid maturational protease precursor (pPR). Together these proteins interact with themselves and with other proteins to coordinate assembly and maturation of the nascent procapsid shell - a process essential to produce infectious virus. We have recently overcome a solubility problem that complicates working with the purified proteins in vitro, and will apply this new approach to pursue leads and test models based on encouraging results from pilot studies. We will do this through three Specific Aims: (i) test predicted effects of phosphorylation on key interactions of pAP and pPR; (ii) verify the differential dominance of the "carboxyl coiled-coil domain" in the self-interaction of pAP versus pPR self-interaction, and determine its biological importance and relevance; and (iii) establish whether a CMV-specific second nuclear localization signal (NLS2) in pAP and pPR is required for their interaction with the capsid portal protein (pUL104). Information gained from this work will help define the sequence of changes that drive UL80 protein associations and dissociations during capsid formation and maturation, and contribute to the longer-term goal of developing new strategies for disrupting (e.g., for therapeutics) or exploiting (e.g., gene/drug delivery vectors) herpesvirus assembly. PUBLIC HEALTH RELEVANCE: Cytomegalovirus is a herpes-group virus that is a threat to people with weakened immune systems, including the very young and the very old. This research seeks to understand how two genetically-related proteins guide the earliest steps in forming infectious virus. Information obtained will relate the molecular interactions driving capsid formation will the biological mechanism of virus formation, and will help identify new ways to interfere therapeutically with that process.