ROTAVIRUS, NORWALK VIRUS, AND ORTHOREOVIRUSES

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 我们研究项目的主要目的是了解结构与功能之间的关系 医学上重要的病毒，如轮状病毒(NIH AI36040)、杯状病毒(NIH P01 AI57788-01，与M.K.Estes博士合作)，正射病毒(NIH RO1 AI32539 in 与T.Dermody博士合作)和流感病毒(与Andrew博士合作 莱斯，正在提交赠款)。我们一直在使用NCRR资助的NCMI设施 多年来，我们将继续使用这一卓越的设施。世界一流的高分辨率 成像和图像处理设施非常适合我们的项目。 轮状病毒：轮状病毒是年轻人严重的、危及生命的胃肠炎的主要原因 孩子和动物。轮状病毒是大的(1000°)、复杂的二十面体组件。 这种病毒已经成为广泛的生化、遗传和结构研究的对象。 因为它的医学相关性，耐人寻味的结构复杂性，以及独特的 复制和形态发生。我们将继续进一步了解这一结构 轮状病毒进入细胞、内源转录、病毒质形成、基因组的基础 使用NCMI CRYO-EM设施以亚纳米分辨率进行复制和封装。 我们的重点将特别放在人类菌株上。虽然其中一些研究涉及 轮状病毒颗粒(直径1000？)，其他与病毒质形成有关的颗粒， 涉及轮状病毒非结构蛋白的基因组复制/包装， NSP5、VP1、VP2和NSP6。在受感染的细胞中，这些蛋白质在 病毒质的形成，基因组的复制/包囊。获得对以下方面的结构性见解 这些轮状病毒蛋白的功能，高分辨率的冷冻-EM分析可能是 唯一的选择。 杯状病毒：杯状病毒分为四个属，是重要的人类和兽医病毒 有可能感染人畜共患病的病原体2。在这些单链RNA病毒中，衣壳相关的功能 例如组装、抗原性和受体相互作用在单个蛋白质中编码 形成了一个二十面体衣壳。我们对获取结构信息感兴趣 关于杯状病毒科的成员。我们将继续使用NCMI设施进行调查 杯状病毒-受体以及杯状病毒-抗体的相互作用。此外，我们计划使用这些 NCMI上提供的资源可用于表征诺沃克病毒p41蛋白 解旋酶/NTPase活性。诺如病毒是流行性腹泻的病原体 人类3. 正形病毒：拟议研究的目标是确定非传染性疾病的机制 使用包膜病毒内化、拆解和膜穿透 哺乳动物呼肠孤病毒作为实验系统。我们将继续使用NCMI的设施来 进一步研究呼肠孤病毒解体和膜穿透的结构基础 确定呼肠孤病毒顺序分解中间产物的结构，以及呼肠孤病毒 在持续呼肠孤病毒感染期间选择的呼肠孤病毒突变体(PI病毒)， 展示了在存在的情况下增强的体外拆解和生长动力学 内体酸化的抑制剂(氯化铵)4和蛋白分解。 流感病毒NS1蛋白：最近出现的高致病性禽流感(H5N1) 流感病毒，其流行性和泛发性及其与致死性的联系 人类感染引发了严重的全球健康问题。几项研究已经 强调非结构蛋白NS1在提高致病性中的重要性 以及这些菌株的毒力。NS1是一种抗病毒的I型干扰素拮抗剂。 主机中的响应。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Major thrust of our research projects is to understand structure-function relationships in medically important viruses such as rotavirus (NIH AI36040), caliciviruses (NIH P01 AI57788-01 in collaboration with Dr. M.K. Estes), orthoreoviruses (NIH RO1 AI32539 in collaboration with Dr. T. Dermody), and influenza viruses (in collaboration with Dr. Andrew Rice, grant submission in progress). We have been using NCRR funded NCMI facility over years, and we will continue to use this excellent facility. The world-class high resolution imaging and image processing facilities are excellently suited for our projects. Rotavirus: Rotavirus is the major cause of severe, life-threatening gastroenteritis in young children and animals. Rotaviruses are large (1000 ¿ ), complex, icosahedral assemblies. This virus has been the subject of extensive biochemical, genetic and structural studies because of its medical relevance, intriguing structural complexity, and unique strategies of replication and morphogenesis. We will continue further understanding of the structural basis of rotavirus cell entry, endogenous transcription, viroplasm formation, genome replication and packaging at sub-nanometer resolution using the NCMI cryo-EM facility. Our focus will be particularly on human strains. While some of these studies involve rotavirus particles (1000 ¿ diameter), others that are related to viroplasm formation, genome replication/packaging involving rotavirus non-structural proteins such as NSP2, NSP5, VP1, VP2 and NSP6. In infected cells, these proteins work in concert during viroplasm formation, genome replication/encapsidation. To obtain structural insights into how these rotavirus proteins function, high-resolution cryo-EM analysis is perhaps the only option. Caliciviruses: Caliciviruses, grouped into four genera, are important human and veterinary pathogens with a potential for zoonosis2. In these ssRNA viruses, capsid related functions such as assembly, antigenicity, and receptor interactions are encoded in a single protein that forms an icosahedral capsid. We are interested in obtaining structural information about members of the Caliciviridae. We will continue to use NCMI facility to investigate calicivirus-receptor, and calicivirus-antibody interactions. In addition, we plan to use these resources available at NCMI for characterizing norovirus p41 protein that exhibits helicase/NTPase activities. Noroviruses are causative agents of epidemic diarrhea in humans 3. Orthoreoviruses: The goal of the proposed research is to determine mechanisms of non- enveloped virus internalization, disassembly, and membrane penetration using mammalian reovirus as an experimental system. We will continue to use NCMI facilities to further investigate structural basis for reovirus disassembly and membrane penetration by determining structures of sequential reovirus disassembly intermediates, and that of reovirus mutants selected during persistent reovirus infection (PI viruses), which demonstrate enhanced in vitro kinetics of disassembly and grow in the presence of inhibitors of endosomal acidification (ammonium chloride)4 and proteolysis. Influenza virus NS1 protein: Recent emergence of highly pathogenic avian (H5N1) influenza viruses, their epizootic and panzootic nature, and their association with lethal human infections have raised significant global health concerns. Several studies have underscored the importance of non-structural protein NS1 in the increased pathogenicity and virulence of these strains. NS1, is an antagonist of antiviral type-I interferon (IFN) response in the host.