Molecular basis for parainfluenza virus infection and host response

描述（由申请方提供）：人副流感病毒（HPIV 1、2和3）是美国因呼吸道病毒感染导致儿科住院的第二大原因。在免疫功能低下的人中，这些病原体也会导致长期疾病，甚至死亡。我们的长期目标是了解副流感病毒如何引起疾病并诱导免疫力，以便开发目前无法获得的特定抗病毒药物和疫苗。副流感病毒融合（F）包膜糖蛋白存在于病毒粒子和感染细胞的表面。我们推测副流感病毒F蛋白通过其促进膜融合和免疫原性的多种功能活性来调节病毒感染和宿主反应。最近确定的高分辨率结构的副流感病毒F蛋白融合前（天然）和融合后（发夹）的形式，现在提供了一个结构基础，以调查的分子机制，F蛋白调节副流感病毒的生物学。本申请的目的是了解副流感病毒F蛋白的结构变化是如何调节的（特异性目的1），以及它们如何帮助确定致病性（特异性目的2）和免疫原性（特异性目的3）。在具体目标1，我们将确定如何HPIV 3 F蛋白结构的变化，在膜融合过程中进行调节。我们将测试的假设，F蛋白的重折叠和膜融合的区域，经历了巨大的结构变化之间的融合前和融合后的F蛋白结构的残基调节。将对HPIV 3 F蛋白中的七肽重复（HR）区域进行突变分析。了解F蛋白结构如何稳定，然后在膜融合过程中触发重新折叠，将提供对蛋白质介导的膜融合的更好理解，这是所有包膜病毒共同的基本机制。在特定目标2中，我们将研究F蛋白的融合性如何导致仙台病毒在小鼠中的致病性。我们将确定超致流重组仙台病毒变异体rSeV-F-L179 V和其他病毒变异体在小鼠中诱导更大致病性的机制。研究仙台病毒在其自然宿主小鼠中的发病机制，将有助于了解F蛋白在发病机制中的作用。这些研究也将支持用新疗法治疗人类副流感病毒感染的努力。在特定目标3中，我们将提高表达HPIV 3 F蛋白的仙台病毒疫苗的免疫原性。我们将确定仙台病毒载体的修饰和HPIV 3 F蛋白抗原的表达结构形式如何帮助确定免疫原性和潜在的免疫病理学。由于F蛋白在所有副粘病毒中是保守的，因此了解HPIV 3 F蛋白如何决定免疫力和免疫病理学可能有助于开发其他重要呼吸道副粘病毒（如其他人副流感病毒（HPIV 1、HPIV 2和HPIV 4）和呼吸道合胞病毒（RSV））的疫苗。

DESCRIPTION (provided by applicant): Human parainfluenza viruses (HPIV1, 2, and 3) are the second leading cause of pediatric hospitalization in the United States due to respiratory viral infection. In the immunocompromised, these pathogens also cause prolonged illness and often death. Our long-term goal is to understand how parainfluenza viruses cause disease and induce immunity so that specific antiviral drugs and vaccines, which are currently unavailable, can be developed. The parainfluenza virus fusion (F) envelope glycoprotein is found on the surfaces of virions and infected cells. We hypothesize that the parainfluenza virus F protein regulates viral infection and the host response by its multiple functional activities of promoting membrane fusion and immunogenicity. Recent determinations of high-resolution structures of parainfluenza virus F proteins in both prefusion (native) and postfusion (hairpin) forms now provides a structural basis to investigate molecular mechanisms by which the F protein regulates the biology of parainfluenza viruses. The objective of this application is to understand how structural changes by parainfluenza virus F proteins are regulated (Specific Aim 1) and how they help determine pathogenicity (Specific Aim 2) and immunogenicity (Specific Aim 3). In Specific Aim 1, we will determine how HPIV3 F protein structural changes are regulated during membrane fusion. We will test the hypothesis that F protein refolding and membrane fusion are regulated by residues in regions that undergo dramatic structural changes between prefusion and postfusion F protein structures. Mutational analyses on heptad repeat (HR) regions in the HPIV3 F protein will be performed. Understanding how the F protein structure is stabilized and then triggered to refold during membrane fusion will provide a greater understanding of protein-mediated membrane fusion, a fundamental mechanism common to all enveloped viruses. In Specific Aim 2, we will investigate how F protein fusogenicity causes Sendai virus pathogenicity in mice. We will determine mechanisms by which a hyperfusogenic recombinant Sendai virus variant rSeV-F-L179V and other virus variants induce greater pathogenicity in mice. Studying Sendai virus pathogenesis in its natural host, the mouse, will provide an understanding of the role of the F protein in pathogenesis. These studies will also support efforts to treat human parainfluenza virus infection with novel therapeutics. In Specific Aim 3, we will increase the immunogenicity of a Sendai virus vaccine that expresses the HPIV3 F protein. We will determine how modifications of the Sendai virus vector and the expressed structural form of the HPIV3 F protein antigen help determine immunogenicity and potential immunopathology. Because the F protein is conserved among all of the paramyxoviruses, an understanding of how the HPIV3 F protein determines immunity and immunopathology may assist in the development of vaccines for other important respiratory paramyxoviruses like the other human parainfluenza viruses (HPIV1, HPIV2, and HPIV4) and respiratory syncytial virus (RSV).