Mechanisms of vGPCR mediated Cytomegalovirus Growth in the Salivary Gland

vGPCR 介导巨细胞病毒在唾液腺中生长的机制

基本信息

批准号：
10180884
负责人：
WILLIAM E MILLER
金额：
$ 40.11万
依托单位：
UNIVERSITY OF CINCINNATI
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-07-01 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10180884
关键词：
Adult Affect American Animal Model Antiviral Agents Area Biological Models Biology Cell model Cell physiology Cells Complement Coupled Cytomegalovirus Cytomegalovirus Infections Data Development Disease Ensure Epithelial Epithelial Cells Fetal Development Future G-Protein Signaling Pathway G-Protein-Coupled Receptors Gene Expression Gene Expression Profile Genes Genetic study Gland Glean Goals Growth Homologous Gene Horizontal Disease Transmission Human Immunocompromised Host Implant In Vitro Individual Infection Intervention Investigation Knock-out Lead Lentivirus Leukocytes Life Liquid substance Mediating Medical Molecular Movement Murid herpesvirus 1 Mus Neurologic Olfactory Mucosa Oral Organ Organoids Parotid Gland Pathogenesis Pathologic Pharmacology Physiological Physiological Adaptation Physiological Processes Play Population Positioning Attribute Process Property Public Health Publishing Regulation Role Saliva Salivary Salivary Glands Science Severities Signal Pathway Signal Transduction Sorting - Cell Movement Species Specificity Spleen Study models Submandibular gland System Testing Tissues Transplant Recipients United States Viral Viral Pathogenesis Viral Proteins Virus Virus Replication base cellular transduction design experimental study in vivo innovation insight knockout animal mouse model mutant novel prevent salivary acinar cell salivary cell salivary gene trafficking transcriptome sequencing transmission process viral transmission

项目摘要

The human cytomegalovirus (HCMV) is a significant public health concern in the United States. Most important are the effects of the virus on developing fetuses and immunocompromised individuals where it causes a variety of pathological conditions ranging in severity from mild to life-threatening. Since HCMV is present in a persistent or latent form in 50-90% of the world’s adult population, the identification of viral gene products that contribute to viral trafficking, persistence, and horizontal transmission is an intense and important area of investigation. Interestingly, HCMV encodes 4 genes that are homologous to cellular G-protein coupled receptors (GPCRs). The HCMV GPCRs are not essential for viral replication in vitro, however their homology to cellular GPCRs suggests that they may profoundly affect cellular physiology to ensure replication of the viruses in organs important for pathogenesis. The strict species specificity of HCMV has precluded an analysis of the function of HCMV GPCRs in vivo. However, as the biology of the related murine cytomegalovirus (MCMV) is similar to that of HCMV, the murine virus has served as a useful model for studying how the vGPCRs affect cytomegalovirus pathogenesis in vivo. Interestingly, the MCMV encoded M33 GPCR is essential for replication within the salivary gland of mice, suggesting that M33 may have a direct impact on persistence or horizontal transmission of virus. Based on our preliminary data, we hypothesize that human and murine CMV encoded vGPCRs activate similar Gaq-dependent signaling pathways to alter salivary acinar cell physiology and facilitate efficient amplification within the salivary epithelium. The proposed studies are highly significant as little is known about the viral and cellular properties that facilitate viral amplification within the gland and promote movement of virus into the saliva. In aim 1, we will use knockout and pharmacological approaches to test the hypothesis that Gaq/Ga11 is the proximal signaling pathway used by M33 for MCMV growth within the salivary gland in vivo. In aim 2, we will use novel animal models to examine cellular gene expression in salivary acinar epithelial cells infected in vivo and determine how MCMV vGPCR-induced changes provide physiological adaptations critical for optimal cytomegalovirus growth in the gland. In aim 3, we will use primary salisphere-derived organoids and test whether salivary epithelial cells similarly require CMV vGPCR activity to promote viral growth in vitro and in vivo. In aim 4 we will examine signaling properties of the HCMV and MCMV vGPCRs in the organoid systems. The innovative experiments proposed in this application will lead to important insight into the function of cytomegalovirus vGPCRs in vivo and define mechanisms by which cytomegaloviruses persist and gain access to fluids important for horizontal transmission. Defining the essential roles for cytomegalovirus vGPCRs in promoting salivary gland replication and spread could ultimately lead to the development of unique antivirals designed to prevent cytomegalovirus transmission via saliva.

在美国，人类巨细胞病毒（HCMV）是一个重大的公共卫生问题。最重要的是该病毒对发育中的胎儿和免疫功能低下的个体的影响，在这种情况下会导致各种病理状况，从而严重程度从轻度到威胁生命。由于HCMV以持续或潜在的形式存在于全球成年人口的50-90％中，因此鉴定了导致病毒贩运，持久性和水平传播的病毒基因产品的鉴定是一个强烈而重要的投资领域。有趣的是，HCMV编码与细胞G蛋白偶联受体（GPCR）同源的4个基因。 HCMV GPCR对于体外病毒复制并不是必不可少的，但是它们与细胞GPCR的同源性表明，它们可能会深刻影响细胞生理学，以确保器官中病毒的复制对发病机理重要。 HCMV的严格规格特异性排除了对体内HCMV GPCR功能的分析。然而，由于相关鼠巨细胞病毒（MCMV）的生物学类似于HCMV，因此鼠病毒已成为研究VGPCR如何影响巨细胞病毒发病机理的有用模型。有趣的是，MCMV编码的M33 GPCR对于在小鼠的唾液腺内复制至关重要，这表明M33可能直接影响病毒的持久性或水平传播。基于我们的初步数据，我们假设人和鼠CMV编码的VGPCR激活了类似的GAQ依赖性信号通路，以改变唾液腺泡细胞生理，并促进唾液上皮中的有效扩增。拟议的研究非常重要，因为对促进腺体内病毒扩增并促进病毒向唾液的病毒和细胞特性知之甚少。在AIM 1中，我们将使用基因敲除和药物方法来测试以下假设：GAQ/GA11是M33用于体内唾液腺内MCMV生长的近端信号传导途径。 In aim 2, we will use novel animal models to examine cellular gene expression in salivary acinar epithelial cells infected in vivo and determine how MCMV vGPCR-induced changes provide physical adaptations critical for optimal In aim 3, we will use primary salisphere-derived organoids and test whether salivariate epithelial cells similarly require CMV vGPCR activity to promote viral growth in vitro and in vivo.在AIM 4中，我们将检查器官系统中HCMV和MCMV VGPCR的信号传导特性。本应用程序中提出的创新实验将导致对体内巨细胞病毒VGPCR的功能的重要洞察力，并定义机制，通过该机制，巨细胞病毒持续存在并获得对水平传播重要的FLU的机制。定义巨细胞病毒VGPCR在促进唾液腺复制和扩散中的基本作用最终可能导致开发旨在防止通过唾液传播的独特抗病毒毒药。