Paramyxovirus F protein mediated membrane fusion

描述（由申请方提供）：膜融合是病毒进入的关键过程，由副粘病毒融合（F）蛋白促进。所有的F蛋白都含有一些在融合中起基本作用的共同特征。然而，F蛋白之间存在显着的差异，与蛋白质活化和促进膜融合，许多关键的问题仍然是关于融合的机制，促进这些重要的病毒蛋白。我们的长期研究目标是了解副粘病毒F蛋白促进膜融合的确切机制。我们的总体假设是，融合促进的关键域的功能将在不同的F蛋白保守，但在这些领域的相互作用将调节触发机制。为了解决这个重要的假设，我们将追求以下具体目标：1。我们将建立在我们的令人兴奋的初步数据，这表明三聚体形成的孤立亨德拉F跨膜（TM）结构域，以确定的作用，TM-TM相互作用的糖蛋白折叠和融合。因此，我们将描述TM-TM相互作用的关键残基，并评估这种相互作用在Hendra F折叠、运输和融合中的作用;分析HRB或细胞质尾添加对TM-TM相互作用稳定性的影响;比较PIV 5和HMPV F蛋白的TM-TM相互作用与Hendra F TM的相互作用;并确定来自副粘病毒附着蛋白的TM结构域是否与自身或与F蛋白TM结构域相互作用; 2.）我们将定义低pH和内吞作用在HMPV病毒进入中的作用，并描述HMPV F低pH诱导的构象变化和重组病毒进入中HRB接头区中静电排斥的作用; 3）我们将分析HRA与F2保守区之间稳定相互作用的作用，我们对PIV 5 F的研究表明，F2保守区可以在触发中发挥作用，通过定义PIV 5 F中该结构域的关键相互作用，评估受影响的融合阶段，分析该区域突变对病毒进入的影响，并分析突变对其他F蛋白中该区域的影响。实现这些目标将提供关键信息的区域，控制参与F蛋白促进膜融合过程的分子事件，并可能确定新的抗病毒治疗的目标。副粘病毒家族既包括已确定的人类病原体，如麻疹病毒和呼吸道合胞病毒，也包括新出现的人类病原体，包括高致病性亨德拉病毒和尼帕病毒以及最近发现的人类偏肺病毒（HMPV）。对融合机制的详细了解可能会导致新的抗病毒试剂的开发。

DESCRIPTION (provided by applicant): Membrane fusion, a process critical for viral entry, is promoted by the paramyxovirus fusion (F) proteins. All F proteins contain a number of common features that play fundamental roles in fusion. However, significant variations exist between F proteins related to protein activation and promotion of membrane fusion, and many critical questions remain concerning the mechanism by which fusion is promoted by these important viral proteins. The long-term objective of our research is to understand the precise mechanism(s) of paramyxovirus F protein-promoted membrane fusion. Our overall hypothesis is that the function of domains critical for fusion promotion will be conserved in diverse F proteins, but that interactions in these domains will modulate triggering mechanisms. To address this important hypothesis we will pursue the following specific aims: 1.) We will build on our exciting preliminary data which demonstrates trimer formation of isolated Hendra F transmembrane (TM) domains to define the role of TM-TM interactions in glycoprotein folding and fusion. We will therefore delineate the residues critical for TM-TM interactions, and evaluate the role of this interaction in Hendra F folding, trafficking and fusion; analyze the effect of HRB or cytoplasmic-tail additions on stability of TM-TM interactions; compare TM-TM interactions for the PIV5 and HMPV F proteins to those observed for the Hendra F TM; and determine if TM domains from paramyxovirus attachment proteins interact with themselves or with the F protein TM domains; 2.) We will define the role of low pH and endocytosis in HMPV viral entry and delineate the role of electrostatic repulsion in the HRB linker region in HMPV F low pH-induced conformational changes and entry of recombinant viruses and; 3.) We will analyze the role of stabilizing interactions between HRA and a conserved region of F2, which our studies of PIV5 F indicate can play a role in triggering, by defining key interactions in this domain in PIV5 F, assessing the stage of fusion which is affected, analyzing the effect of mutations in this region on viral entry, and analyzing the effect of mutations to this region in other F proteins. Accomplishing these goals will provide crucial information on the regions that control the molecular events involved in the F protein-promoted membrane fusion process, and potentially identify new targets for antiviral therapy. PUBLIC HEALTH RELEVANCE The paramyxovirus family contains both established human pathogens, such as measles virus and respiratory syncytial virus, and newly emerged human pathogens, including the highly pathogenic Hendra and Nipah viruses and the recently identified human metapneumovirus (HMPV). A detailed understanding of the fusion mechanism could lead to development of new antiviral reagents.