Role of CMV kinase in regulating infection of ESC-derived neuroprogenitor cells

CMV 激酶在调节 ESC 来源的神经祖细胞感染中的作用

• 批准号: 8256085
• 负责人: Tarin M Bigley
• 金额: $ 4.43万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-02-01 至 2017-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8256085
• 关键词: Antiviral Agents; Apoptosis; Biological Models; Birth; Brain; Cell Cycle; Cell Death; Cell Differentiation process; Cell Line; Cells; Cessation of life; Child; Chromatin; Cyclin-Dependent Kinases; Cytolysis; Cytomegalovirus; Cytomegalovirus Infections; Data; Deacetylase; Development; Diploidy; Disease; Early Promoters; Effectiveness; Embryo; Event; Exhibits; Fellowship; Fibroblasts; Future; Gene Expression; Genome; Goals; Herpesviridae; Histones; Human; Immediate-Early Genes; Infection; Infection prevention; Life; Lytic; Mediating; Modeling; Molecular; Mothers; Nervous System Trauma; Neuraxis; Neuroglia; Neurons; Nuclear Pore Complex; Pathway interactions; Pattern; Pharmacologic Substance; Phosphorylation; Phosphotransferases; Post-Translational Protein Processing; Pregnancy; Process; Protein Kinase; Protein-Serine-Threonine Kinases; Proteomics; Recruitment Activity; Regulation; Risk; Role; Sensorineural Hearing Loss; Specificity; Testing; Time; Toxic effect; United States; Viral; Virus; Virus Diseases; cell motility; congenital cytomegalovirus; congenital infection; developmental disease; embryonic stem cell; fetal; hearing impairment; histone modification; human HDAC1 protein; inhibitor/antagonist; kinase inhibitor; maribavir; neonate; nerve stem cell; neural precursor cell; neuron development; non-genetic; precursor cell; prevent; progenitor; promoter

DESCRIPTION (provided by applicant): Human cytomegalovirus (CMV) is the most common cause of non-hereditary sensorineural hearing loss in the U.S. Current therapies to treat infection during pregnancy or upon birth are unproven and risky. The experimental antiviral compound maribavir is a specific inhibitor of the CMV protein kinase, pUL97, and exhibits low toxicity. The objective of my proposal is to identify the molecular mechanism behind pUL97 kinase-mediated regulation of CMV gene expression and determine the impact of inhibiting kinase activity on CMV infected embryonic stem cell-derived neuronal progenitor cells. I have demonstrated that inhibition of the CMV kinase resulted in decreased expression of viral immediate early (IE) genes. This is a newly identified function of the CMV pUL97 kinase. Our lab previously identified an interaction between the CMV pUL97 kinase and cellular histone deacetylase 1 (HDAC1). HDAC1 acts as a repressor and is recruited to the MIE promoter at early times during infection. Therefore, I hypothesize that the kinase activity of CMV pUL97 influences the histone modification pattern at the viral major immediate early promoter by altering cellular HDAC1 activity. I propose to evaluate the pUL97-dependent changes of histone modifications at the major immediate early promoter (MIEP) during infection. These studies will be initiated using human diploid fibroblasts. In addition, I will identify pUL97-mediated changes in HDAC1 localization within the infected cells and at the MIEP. Other herpesvirus kinases phosphorylate HDAC1 and I propose several approaches to identify potential changes in HDAC1 phosphorylation by pUL97. I will determine the impact of these changes on HDAC1 deacetylase activity. I will further these studies using an embryonic stem cell-derived neural progenitor cell line (ES-NPCs) as a model system for CMV-induced neurological damage due to congenital CMV infection. Expression of IE genes has been demonstrated to alter fetal/neonate NPC differentiation. CMV IE genes regulate cell cycle and apoptosis in human diploid fibroblasts. I will test the role of pUL97 during these early events in the context of ES-NPC's by confirming the major results that were obtained in our fibroblast studies. I will determine the effectiveness of maribavir in preventing the pathogenic effects of HCMV infection on neuronal development. My studies will determine if a pUL97 kinase inhibitor is suitable for treating CMV infected neonates and possibly CMV infected mothers at risk of congenital infection. PUBLIC HEALTH RELEVANCE: Congenital cytomegalovirus (CMV) infection occurs in 1 out of 150 pregnancies each year in the U.S. Infection results in approximately 400 neonate deaths and 5,500 children with developmental disorders of the central nervous system. We will determine the molecular mechanism behind an event that occurs very early during infection and explore possible ways of blocking that event and preventing infection within neuronal progenitor cells.

描述（由申请人提供）：人类巨细胞病毒（CMV）是美国非遗传性感音神经性听力损失的最常见原因。目前治疗怀孕期间或出生时感染的疗法未经证实且存在风险。实验性抗病毒化合物 maribavir 是 CMV 蛋白激酶 pUL97 的特异性抑制剂，具有低毒性。我的提议的目的是确定 pUL97 激酶介导的 CMV 基因表达调节背后的分子机制，并确定抑制激酶活性对 CMV 感染的胚胎干细胞衍生的神经祖细胞的影响。我已经证明，CMV 激酶的抑制会导致病毒立即早期 (IE) 基因的表达减少。这是 CMV pUL97 激酶新发现的功能。我们的实验室之前发现了 CMV pUL97 激酶和细胞组蛋白脱乙酰酶 1 (HDAC1) 之间的相互作用。 HDAC1 充当阻遏蛋白，并在感染早期被招募至 MIE 启动子。因此，我推测 CMV pUL97 的激酶活性通过改变细胞 HDAC1 活性来影响病毒主要立即早期启动子的组蛋白修饰模式。我建议评估感染期间主要立即早期启动子 (MIEP) 处组蛋白修饰的 pUL97 依赖性变化。这些研究将使用人类二倍体成纤维细胞开始。此外，我将确定 pUL97 介导的受感染细胞内和 MIEP 上 HDAC1 定位的变化。其他疱疹病毒激酶也会磷酸化 HDAC1，我提出了几种方法来识别 pUL97 对 HDAC1 磷酸化的潜在变化。我将确定这些变化对 HDAC1 脱乙酰酶活性的影响。我将使用胚胎干细胞衍生的神经祖细胞系（ES-NPC）作为先天性巨细胞病毒感染引起的巨细胞病毒诱导的神经损伤的模型系统来进一步开展这些研究。 IE 基因的表达已被证明可以改变胎儿/新生儿 NPC 的分化。 CMV IE 基因调节人二倍体成纤维细胞的细胞周期和细胞凋亡。我将通过确认我们的成纤维细胞研究中获得的主要结果来测试 pUL97 在 ES-NPC 背景下的这些早期事件中的作用。我将确定马利巴韦在预防 HCMV 感染对神经元发育的致病作用方面的有效性。我的研究将确定 pUL97 激酶抑制剂是否适合治疗 CMV 感染的新生儿以及可能感染 CMV 且有先天性感染风险的母亲。 公共卫生相关性：在美国，每年 150 名孕妇中就有 1 名会发生先天性巨细胞病毒 (CMV) 感染。感染导致约 400 名新生儿死亡，并导致 5,500 名儿童出现中枢神经系统发育障碍。我们将确定感染早期发生的事件背后的分子机制，并探索阻止该事件和预防神经元祖细胞内感染的可能方法。