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The role of host and viral translation factors during HCMV infection

宿主和病毒翻译因子在 HCMV 感染过程中的作用

基本信息

批准号：
10456099
负责人：
Nathaniel J Moorman
金额：
$ 38.46万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-12-01 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10456099
关键词：
5&apos Untranslated Regions Adult Antiviral Agents Binding Cells Child Complement Complex Cytomegalovirus Data Disease Drug Targeting Drug toxicity Ensure Eukaryotic Initiation Factor-4F Event Genetic Transcription Genetic Translation Goals Human Immunocompromised Host Infection Knowledge Lead Messenger RNA Molecular Pathway interactions Peptide Initiation Factors Pharmaceutical Preparations Phosphotransferases Protein Biosynthesis Proteins Publishing RNA RNA Helicase RNA Sequences RNA helicase A RNA-Binding Proteins Resistance Ribosomes Role Series Structure Transcript Translating Translation Initiation Translations Viral Viral Proteins Virus Virus Replication adaptive immune response antagonist base experimental study human pathogen innovation insight mRNA capping novel pathogen prevent protein expression public health relevance recruit translation factor translation to humans translational approach translational barrier

项目摘要

Abstract No virus encodes a ribosome, thus all viruses require the host translation machinery for the synthesis of viral proteins. This fundamental host:pathogen interface creates competition for the host translation machinery, which is a critical barrier to human cytomegalovirus (HCMV) mRNA translation. Many viruses prevent competition for the translation machinery by inhibiting host translation, however host translation is maintained during HCMV infection. How HCMV mRNAs successfully compete for access to host translation machinery to ensure efficient viral protein synthesis is poorly understood, highlighting a critical gap in our knowledge of the mechanisms controlling HCMV mRNA translation. How might HCMV mRNAs compete with host mRNAs for access to ribosomes? Based on our data we hypothesize that HCMV avoids competition by using a unique complement of host and viral translation initiation factors to facilitate viral mRNA translation. Formation of the eIF4F translation initiation complex is the limiting step in host mRNA translation, thus competition between host and viral mRNAs would be most intense for eIF4F components. We showed that the eIF4F complex is necessary for the continued translation of host mRNAs during infection; however HCMV mRNAs translate efficiently when the eIF4F complex is disrupted. We found that translation of the HCMV IE1 and IE2 mRNAs, which encode critical regulators of virus replication, is resistant to eIF4F disruption, depletion or inhibition. How HCMV mRNAs translate in the absence of eIF4F is unknown. As translation initiation factors assemble on the mRNA 5' untranslated region (5'UTR), in Aim1 we determine how RNA sequence and structure in the shared 5'UTR of the IE1 and IE2 mRNAs recruit cellular factors to allow for their eIF4F-independent translation. We also determine how these factors contribute to the widespread eIF4F-independent translation of HCMV mRNAs as a whole. Aim 2 builds on our published results suggesting that the HCMV TRS1 protein (pTRS1) acts as a viral translation initiation factor to facilitate viral protein synthesis. While pTRS1 antagonizes the antiviral kinase PKR, which inhibits protein synthesis, we showed that pTRS1 stimulates translation in PKR deficient cells. Further, we found that pTRS1 binds the mRNA cap independent of the eIF4F complex and co- purifies with 40S ribosomal subunits, suggesting a role in translation initiation. We now show that pTRS1 interacts with the DHX29 RNA helicase, a translation initiation factor that can recruit ribosomes to mRNAs in the absence of the eIF4F complex, and that DHX29 is necessary for efficient IE1/2 mRNA translation and virus replication. In Aim 2 we role of pTRS1 and DHX29 in translation initiation complex on HCMV mRNAs, and the role of their interaction in the ability of pTRS1 to enhance translation independent of PKR antagonism. We further determine how pTRS1 binding to DHX29 impacts virus replication and the translation of HCMV mRNAs in the absence of the eIF4F complex. Together these aims will identify regulatory events unique to HCMV mRNA translation that could be targeted to limit HCMV disease.

摘要没有病毒编码核糖体，因此所有病毒都需要宿主翻译机器来合成病毒的核糖体。 proteins.这种基本的宿主：病原体界面为宿主翻译机制创造了竞争，这是人巨细胞病毒（HCMV）mRNA翻译的关键障碍。许多病毒会阻止通过抑制宿主翻译来竞争翻译机制，然而宿主翻译被维持在HCMV感染期间。HCMV mRNAs如何成功地竞争进入宿主翻译机器，确保有效的病毒蛋白质合成是知之甚少，突出了我们的知识的关键差距，控制HCMV mRNA翻译的机制。HCMV mRNA如何与宿主mRNA竞争进入核糖体基于我们的数据，我们假设HCMV通过使用独特的宿主和病毒翻译起始因子的互补以促进病毒mRNA翻译。形成 eIF 4F翻译起始复合物是宿主mRNA翻译的限制性步骤，因此宿主之间存在竞争而病毒mRNA对eIF 4F组分的表达最强。我们发现eIF 4F复合物是在感染过程中宿主mRNA的持续翻译所必需的;然而HCMV mRNA翻译当eIF 4F复合物被破坏时，我们发现HCMV IE 1和IE 2 mRNA的翻译，其编码病毒复制的关键调节因子，对eIF 4F破坏、耗尽或抑制具有抗性。如何 HCMV mRNAs在eIF 4F缺失的情况下的翻译是未知的。当翻译启动因子聚集在 mRNA 5'非翻译区（5' UTR），在Aim 1中，我们确定了RNA序列和结构如何在共享的 IE 1和IE 2 mRNA的5 'UTR募集细胞因子以允许它们的eIF 4F非依赖性翻译。我们还确定了这些因素如何促进HCMV广泛的eIF 4F非依赖性翻译 mRNAs作为一个整体目的2建立在我们发表的结果表明，HCMV TRS 1蛋白（pTRS 1）作为病毒翻译起始因子促进病毒蛋白质合成。虽然pTRS 1拮抗抗病毒激酶PKR抑制蛋白质合成，我们发现pTRS 1刺激PKR的翻译缺陷细胞此外，我们发现pTRS 1不依赖于eIF 4F复合物与mRNA帽结合，纯化40 S核糖体亚基，表明在翻译起始中的作用。我们现在发现pTRS 1 与DHX 29 RNA解旋酶相互作用，DHX 29 RNA解旋酶是一种翻译起始因子，可以将核糖体募集到mRNA中， eIF 4F复合物的缺乏，以及DHX 29对于有效的IE 1/2 mRNA翻译和病毒复制的目的2：研究pTRS 1和DHX 29在HCMV mRNAs翻译起始复合物中的作用，它们的相互作用在pTRS 1不依赖于PKR拮抗作用而增强翻译的能力中的作用。我们进一步确定pTRS 1与DHX 29的结合如何影响病毒复制和HCMV mRNA的翻译在不存在eIF 4F复合物的情况下。这些目标将共同确定HCMV特有的调控事件可以有针对性地限制HCMV疾病的mRNA翻译。