Regulation of Rotavirus Replication

轮状病毒复制的调控

• 批准号: 10597603
• 负责人: Sue Ellen Crawford
• 金额: $ 53.07万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-11 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10597603
• 关键词: Address; Affect; Antiviral Therapy; Autophagocytosis; BCAR3 gene; Bacterial Infections; Biogenesis; Biology; Calcium; Cells; Cellular Membrane; Child; Cholesterol Esters; Chylomicrons; Cytoplasm; Cytoplasmic Organelle; Diabetes Mellitus; Diarrhea; Disease; Endoplasmic Reticulum; Enterocytes; Enzymes; Fatty acid glycerol esters; Gastroenteritis; Goals; Health; Heart Diseases; Homeostasis; Human; Hypertriglyceridemia; Immune response; Infectious Agent; Inflammatory Response; Intestines; Intracellular Membranes; Knowledge; Lead; Life; Lipids; Membrane; Metabolic Pathway; Metabolism; Modeling; Molecular; Nonstructural Protein; Obesity; Organelles; Pathogenesis; Pathway interactions; Phospholipids; Phosphorylation; Play; Protein Isoforms; Protein Kinase; Proteins; Regulation; Research; Role; Rotavirus; Rotavirus Infections; Signal Pathway; Signal Transduction; Site; Small Intestines; Tissues; Triglycerides; Vaccines; Viral; Viral Proteins; Virus; Virus Diseases; Virus Replication; Work; absorption; design; diacylglycerol O-acyltransferase; enteric infection; experimental study; human disease; human pathogen; immunoregulation; insight; interest; model organism; monolayer; multicatalytic endopeptidase complex; novel; pathogen; perilipin; trafficking

Project Summary Our long-term goal is to understand how rotavirus (RV) exploits cellular pathways such as autophagy membranes, calcium homeostasis, and lipid droplet (LD) formation to enhance their replication and cause disease. RVs remain significant human pathogens in spite of the introduction of vaccines. Several aspects of RV replication are unique yet broadly relevant to other viruses, such as cytoplasmic organelles (viroplasms, VI) formed by both viral and cellular proteins and LDs that form a physical platform for efficient viral replication and maturation. LDs are dynamic, multi-functional intracellular organelles involved in lipid storage and metabolism as well as in signal transduction, membrane trafficking and modulation of immune and inflammatory responses. LDs play essential roles in several viral and intracellular bacterial infections and are important in many aspects of health and disease (metabolism, diabetes, obesity, heart disease). However, mechanistic information of the interplay between lipid accumulation and these pathogens, and disease is far from complete. Our proposed studies on LDs and VIs build on our recent work. While the viral proteins NSP2 and NSP5 are known to be required for VI formation, the molecular mechanisms of how these two proteins associate with each other as well as with other viral and cellular proteins and LD components to form VI/LDs remain to be elucidated. We discovered two forms of NSP2 that interact with different isoforms of NSP5: a dispersed (dNSP2) form interacts with hypo-phosphorylated NSP5 and a previously recognized VI (vNSP2) form interacts with hyper- phosphorylated NSP5. We elucidated a novel phosphorylation-dependent mechanism for VI formation, in which the ubiquitous, constitutively active cellular protein kinase CK1α partially controls the assembly of RV VIs by phosphorylating NSP2 to trigger NSP2 octamer-octamer lattice formation. We also discovered that NSP2 is an autokinase and predict that NSP2 may phosphorylate other viral or cellular proteins for VI assembly and RV replication. We hypothesize that interactions of RV and cellular proteins in specialized microdomains of the endoplasmic reticulum nucleate and induce VI/LDs essential for virus replication, affect the composition of the LD-associated proteins and result in previously unrecognized mechanisms of RV-induced pathogenesis. We propose experiments to answer three questions. (1) How do NSP4, NSP2 and specialized microdomains in the ER lead to nucleation of VI/LDs? (2) How does phosphorylation orchestrate VI formation and the conversion of dNSP2 to vNSP2 to initiate VI/LD formation and subsequent VI/LD maturation? (3) How does DGAT1 degradation lead to LD formation and what are the specific roles of PLIN1 and PLIN3 LDs in RV infection and pathogenesis? These studies are significant because viral perturbations of host signaling and metabolic pathways that involve LDs are critical for multiple pathogens. Because RVs replicate in enterocytes in the small intestine, the major site of fat absorption in the body, understanding the effects of RV infection on LD biology has the potential to reveal new insights into the consequences of virus infection on host metabolism.

项目摘要 我们的长期目标是了解轮状病毒（RV）如何利用诸如自噬之类的细胞途径 膜，钙稳态和脂质液滴（LD）形成，以增强其复制和原因 疾病。尽管引入了疫苗，但RV仍然是人类病原体。 RV的几个方面 复制是独特但与其他病毒广泛相关的，例如细胞质细胞器（Viroplasms，vi） 由病毒蛋白和细胞蛋白和LD形成，形成了有效病毒复制和的物理平台 成熟。 LDS是动态的，多功能的细胞内细胞器，涉及脂质储存和代谢 以及在信号转导，膜运输以及免疫和炎症反应的调节中。 LD在几种病毒和细胞内细菌感染中起着重要作用，在许多方面都很重要 健康与疾病（代谢，糖尿病，肥胖，心脏病）。但是，机械信息 脂质积累与这些病原体之间的相互作用以及疾病远非完整。我们提出的 关于LD和VIS的研究基于我们最近的工作。众所周知，病毒蛋白NSP2和NSP5是 VI形成所需的，这两种蛋白质如何相互关联的分子机制 与其他病毒和细胞蛋白和LD成分一样，形成VI/LD的成分仍有待阐明。我们 发现了两种形式的NSP2与NSP5的不同同工型相互作用：分散（DNSP2）形式相互作用 使用低磷酸化的NSP5和先前公认的VI（VNSP2）形式与超级相互作用 磷酸化的NSP5。我们阐明了一种新型的磷酸化依赖性机制，用于VI形成，其中 普遍存在的，组成型活性的细胞蛋白激酶CK1α部分控制着通过 磷酸化NSP2触发NSP2八聚体晶格形成。我们还发现NSP2是 自动激酶并预测NSP2可能会磷酸化其他病毒或细胞蛋白进行VI组装和RV 复制。我们假设RV和细胞蛋白在专门的微域中的相互作用 内质网核并诱导病毒复制必不可少的VI/LD LD相关的蛋白质，并导致先前未识别的RV诱导的发病机制。我们 提案实验回答三个问题。 （1）NSP4，NSP2和专业微域如何 ER导致VI/LDS的成核？ （2）磷酸化如何编排VI形成和转换 DNSP2至VNSP2启动VI/LD形成和随后的VI/LD成熟？ （3）DGAT1如何 降解导致LD形成以及PLIN1和PLIN3 LD在RV感染中的特定作用是什么 发病？这些研究很重要，因为宿主信号和代谢的病毒扰动 涉及LD的途径对于多种病原体至关重要。因为RVS在小的肠球菌中复制 肠道是体内脂肪滥用的主要部位，了解RV感染对LD生物学的影响 有可能揭示对病毒感染对宿主代谢的后果的新见解。