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.

项目概要 我们的长期目标是了解轮状病毒 (RV) 如何利用自噬等细胞途径 膜、钙稳态和脂滴 (LD) 形成，以增强其复制并引起 尽管引入了 RV 疫苗，但 RV 仍然是重要的人类病原体。 复制是独特的，但与其他病毒广泛相关，例如细胞质细胞器（病毒质，VI） 由病毒和细胞蛋白和 LD 组成，形成有效病毒复制的物理平台 LD 是参与脂质储存和代谢的动态多功能细胞内细胞器。 以及信号转导、膜运输以及免疫和炎症反应的调节。 LD 在多种病毒和细胞内细菌感染中发挥着重要作用，并且在许多方面都很重要 健康和疾病（新陈代谢、糖尿病、肥胖、心脏病）的机制信息。 脂质积累与这些病原体和疾病之间的相互作用还远未完成。 对 LD 和 VI 的研究建立在我们最近的工作基础上，而已知病毒蛋白 NSP2 和 NSP5 是。 VI 形成所需的，这两种蛋白质如何相互结合的分子机制 与其他病毒和细胞蛋白以及形成 VI/LD 的 LD 成分一样，我们仍有待阐明。 发现了两种形式的 NSP2 与 NSP5 的不同亚型相互作用：分散的 (dNSP2) 形式相互作用 低磷酸化的 NSP5 和先前识别的 VI (vNSP2) 形式与超磷酸化相互作用 我们阐明了 VI 形成的一种新的磷酸化依赖性机制，其中 普遍存在的、组成型活性细胞蛋白激酶 CK1α 通过以下方式部分控制 RV VI 的组装： 磷酸化 NSP2 以触发 NSP2 八聚体-八聚体晶格形成。 自动激酶并预测 NSP2 可能会磷酸化其他病毒或细胞蛋白以进行 VI 组装和 RV 我们研究了 RV 和细胞蛋白在特定微域中的相互作用。 内质网成核并诱导病毒复制必需的 VI/LD，影响内质网的组成 LD 相关蛋白并导致以前未被认识的 RV 诱导发病机制。 (1) NSP4、NSP2和特化微结构域在 ER 导致 VI/LD 成核？ (2) 磷酸化如何协调 VI 形成和转化 dNSP2 到 vNSP2 启动 VI/LD 形成和随后的 VI/LD 成熟？ 降解导致 LD 形成，PLIN1 和 PLIN3 LD 在 RV 感染和感染中的具体作用是什么？ 这些研究意义重大，因为病毒对宿主信号传导和代谢的干扰 涉及 LD 的途径对于多种病原体至关重要，因为 RV 在小肠细胞中复制。 肠道是体内脂肪吸收的主要部位，了解 RV 感染对 LD 生物学的影响 有可能揭示病毒感染对宿主代谢影响的新见解。