Comprehensive genetic dissection of poxvirus membrane assembly and function

痘病毒膜组装和功能的全面基因解剖

基本信息

批准号：
10575027
负责人：
Kartik Chandran
金额：
$ 25.2万
依托单位：
ALBERT EINSTEIN COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-12-01 至 2024-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10575027
关键词：
Actins Amino Acids Behavior assessment Binding Biochemical Biological Assay Biology C-Type Lectins C-terminal Cell surface Cells Collaborations Complement Complex Cowpox virus Cytoplasm Cytoplasmic Tail Data Defect Development Dissection GPA33 gene Gene Combinations Generations Genetic Genetic Screening Genetic study Growth Human Immunity Individual Infection Internet Intracellular Membranes Libraries Mammals Mass Spectrum Analysis Mediating Membrane Proteins Molecular Monkeypox virus Morphogenesis Mutagenesis Mutate Mutation Nature Oncolytic Orthopoxvirus Persons Phenotype Point Mutation Population Poxviridae Proteins Proteomics Public Health Reporting Risk Role Sensitivity and Specificity Serial Passage Series Site Smallpox Surface Therapeutic Therapeutic antibodies Vaccinated Vaccination Vaccines Vaccinia virus Viral Viral Proteins Virion Virulence Virus Work Zoonoses cancer therapy cell motility extracellular fitness genomic locus improved in vivo loss of function member membrane assembly membrane biogenesis mutant mutation screening new epidemic next generation sequencing novel oncolytic vector particle poxvirus vectors prevent protein protein interaction prototype reverse genetics tissue tropism transmission process virus genetics virus host interaction virus virus interaction

Actins Amino Acids Behavior assessment Binding Biochemical Biological Assay Biology C-Type Lectins C-terminal Cell surface Cells Collaborations Complement Complex Cowpox virus Cytoplasm Cytoplasmic Tail Data Defect Development Dissection GPA33 gene Gene Combinations Generations Genetic Genetic Screening Genetic study Growth Human Immunity Individual Infection Internet Intracellular Membranes Libraries Mammals Mass Spectrum Analysis Mediating Membrane Proteins Molecular Monkeypox virus Morphogenesis Mutagenesis Mutate Mutation Nature Oncolytic Orthopoxvirus Persons Phenotype Point Mutation Population Poxviridae Proteins Proteomics Public Health Reporting Risk Role Sensitivity and Specificity Serial Passage Series Site Smallpox Surface Therapeutic Therapeutic antibodies Vaccinated Vaccination Vaccines Vaccinia virus Viral Viral Proteins Virion Virulence Virus Work Zoonoses cancer therapy cell motility extracellular fitness genomic locus improved in vivo loss of function member membrane assembly membrane biogenesis mutant mutation screening new epidemic next generation sequencing novel oncolytic vector particle poxvirus vectors prevent protein protein interaction prototype reverse genetics tissue tropism transmission process virus genetics virus host interaction virus virus interaction

展开

项目摘要

The zoonotic potential of members of the genus Orthopoxvirus (orthopoxviruses) of mammal- infecting poxviruses represent a continued threat to global public health. A hallmark of the complex biology of orthopoxvirus multiplication in the host cell cytoplasm is the generation of two distinct infectious forms—intracellular mature virions (IMV), which remain largely cell- associated, and enveloped virions (EV), which are critical for extracellular viral spread. EVs bear a unique complement of eight virus-encoded membrane proteins at their surface that influence tissue tropism, long-range in vivo dissemination, and ultimately, virulence; provide key targets for vaccine-induced and therapeutic antibodies; and are important determinants of the therapeutic potential of oncolytic poxvirus vectors. Despite their importance, a comprehensive understanding of EVs is challenged by the complex networks of virus-host and virus-virus interactions that underlie their morphogenesis and function. Our overall objective is to unravel these functional protein interaction networks through a combination of genetic and proteomic approaches. Using a new platform for unbiased and deep mutagenesis of any genomic locus in the prototypic orthopoxvirus, vaccinia virus (VACV), we conducted deep-mutational scans (DMS) of two EV proteins, A33 and A34, in the context of infectious viral particles and identified novel regiospecific mutations that enhance EV fitness. We will elucidate the mechanisms of action of these mutants. In parallel, we will perform genetic modifier and proteomic screens to uncover new functional interactions of A33/A34 with viral and host proteins. Collectively, these studies will provide new information on how EVs—critical for both the virulence of orthopoxviruses and their utility as oncolytic vectors—assemble, egress, and spread to new cells during infection.

哺乳动物感染痘病毒的正托病毒属（正托病毒）成员的人畜共患潜力代表了对全球公共卫生的持续威胁。宿主细胞细胞质中正托病毒繁殖复杂生物学的标志是产生两种不同的感染性形式 - 感染性成熟病毒（IMV），这些病毒（IMV）在很大程度上与细胞相关，并且包膜病毒（EV），这对于细胞外病毒外传播至关重要。电动汽车在其表面上具有八种病毒编码的膜蛋白的独特补体，这些膜蛋白会影响组织的向潮流，体内远距离传播，并最终导致病毒。提供疫苗诱导和治疗性抗体的关键靶标；并且是溶瘤毒病毒载体的治疗潜力的重要决定剂。尽管它们的重要性，但对电动汽车的全面了解受到了病毒宿主和病毒 - 病毒相互作用的复杂网络的挑战，这些网络是其形态发生和功能的基础。我们的总体目标是通过遗传和蛋白质组学方法的结合来揭示这些功能性蛋白质相互作用网络。我们使用一个新的平台，用于对原型正托病毒中任何基因组基因座的无偏和深诱变，vacicinia病毒（VACV），我们对两种EV蛋白A33和A34进行了深入的扫描（DMS），该扫描是在感染性病毒颗粒和标识的新型调节性范围的上下文中，以增强了EVEVEVENDESS，以增强了新型病毒颗粒和标识的verectialsspectialspec。我们将阐明这些突变体的作用机理。同时，我们将执行遗传修饰剂和蛋白质组学筛选，以发现A33/A34与病毒和宿主蛋白的新功能相互作用。总的来说，这些研究将提供有关在感染期间的EVS（对正托病毒病毒及其作为溶瘤载体的效用至关重要）的新信息。