Selective Degradation of mRNA by Herpes Simplex Virus 1

单纯疱疹病毒 1 对 mRNA 的选择性降解

• 批准号: 8458492
• 负责人: Bernard Roizman
• 金额: $ 30.59万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-06-10 至 2015-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8458492
• 关键词: 3' Untranslated Regions; Actins; Applications Grants; Binding; Biology; Cells; Cleaved cell; Complex; Degradation Pathway; Elements; Endoribonucleases; Enzymes; FOS gene; Genes; Half-Life; Herpes Simplex Virus Protein Vmw65; Herpesvirus 1; Hour; Immune response; Infection; Ligands; Mediating; Messenger RNA; Modeling; Mus; Pancreatic ribonuclease; Pathogenesis; Perception; Poly(A)-Binding Proteins; Polyribosomes; Procedures; Protein Binding; Proteins; RNA; RNA Cap-Binding Proteins; Recruitment Activity; Relative (related person); Reporting; Research Proposals; Residual state; Ribonucleases; Role; Simplexvirus; Specificity; Stress; Structure; TIS11 protein; Testing; Time; Untranslated Regions; VP 16; Viral; Viral Proteins; Virion; Virus; base; biological adaptation to stress; decapping enzyme; eIF-4B; endoribonuclease; follow-up; mRNA Transcript Degradation; mutant; protein function; public health relevance; research study; response; time interval

DESCRIPTION (provided by applicant): Virion host shutoff protein (VHS), the product of UL41 gene of herpes simplex virus 1(HSV-1) is the major viral suppressor of host responses to infection. VHS is brought into the infected cells by the virion and its primary function is to degrade mRNA. Our studies on VHS have completely revolutionized our understanding perceptions of the functions of the protein. Specifically: (i) We have developed a procedure for solubilization and purification of VHS to homogeneity. Purified VHS is an endoribonuclease that cleaves synthetic RNA with the specificity of RNase A in the absence of cellular or viral proteins. (ii) Earlier reports indicated that at mid and late times after infection VHS is "neutralized by the viral protein VP16. We showed that "neutralization" required the formation of a complex that includes VP22 in addition to VP16. (iii) Earlier reports indicated that the degradation of mRNA mediated by VHS is nonselective. We have identified and quantified the VHS- dependent degradation of 3 classes of mRNAs as follows: Class A consists of mRNAs that are normally stable (e.g. actin, GAPDH mRNAs, normal half life >12 h). In infected cells these mRNAs are degraded 5' to 3' within 30 minutes. Class B consists of stress response mRNAs (e.g. c-fos, cox-2, I:B1 and IEX-1 mRNAs) induced immediately after infection. These mRNAs contain AU-rich elements within their 3' UTRs. Typical half life of these mRNAs in mock infected cells is 30 to 45 min. Early after infection, these mRNAs are deadenylated and cleaved 5' to AU-rich elements. The residual portions linger with a half life >3 hrs. At late times (6+ h after infection) degradation of these mRNAs is dependent on ICP27. Both class A and class B mRNAs are degraded in polyribosomes. Class C includes mRNAs that are not degraded. The products of these mRNAs include tristetraprolin, a stress inducible protein that binds to AU-rich elements and sequesters the mRNAs to exosomes for 3' to 5' degradation and GADD452 a transcriptional factor. (iv) Earlier reports indicated that VHS binds 3 components of eIF4F complex, i.e. eIF4H, eIF4B and eIF4AII and VP16. Our studies have shown that VHS also interacts physically with tristetraprolin, ICP27, and through VP16 with VP22. The objective of the research proposal is to test a model of the degradation of the mRNAs in classes A and B. The model proposes that VHS degrades mRNAs in polyribosomes as reported, that it binds to a still to be defined component of eIF4F and mimics the function of the decapping enzymes by cleaving stable mRNAs downstream of the cap structure. The residual portion is then sequestered in P-bodies and rapidly degraded. In the case of class B mRNAs, VHS binds to tristetraprolin bound to AU-rich element mRNA and cleaves the mRNA 5' to the AU-rich elements. In the absence of bound tristetraprolin, the residual mRNA is not recruited promptly for degradation 3' to 5' by exosomes. We also propose to discriminate between 2 alternative potential functions of ICP27 in enabling mRNA degradation late in infection. Finally, given the multiple functions of VHS, the question arises as to the extent to which each function contributes to the pathogenic potential of HSV-1.

描述（由申请方提供）：病毒体宿主关闭蛋白（VHS），单纯疱疹病毒1型（HSV-1）UL 41基因的产物，是宿主对感染反应的主要病毒抑制因子。VHS由病毒体带入感染细胞，其主要功能是降解mRNA。我们对VHS的研究彻底改变了我们对蛋白质功能的理解。具体而言：（i）我们已经开发了用于将VHS溶解和纯化至均质的程序。纯化的VHS是一种核糖核酸内切酶，在不存在细胞或病毒蛋白的情况下，其以RNA酶A的特异性切割合成RNA。(ii)早期的报道指出，在感染后的中期和晚期，VHS被病毒蛋白VP 16中和。我们表明，“中和”需要形成一个复合物，其中包括VP 22除了VP 16。(iii)早期的报道表明VHS介导的mRNA降解是非选择性的。我们已经鉴定并定量了如下3类mRNA的VHS依赖性降解：A类由正常稳定的mRNA组成（例如肌动蛋白、GAPDH mRNA，正常半衰期>12小时）。在感染的细胞中，这些mRNA在30分钟内从5'降解到3'。B类由感染后立即诱导的应激反应mRNA（例如c-fos、考克斯-2、I：B1和IEX-1 mRNA）组成。这些mRNA在其3'UTR内含有富含AU的元件。这些mRNA在模拟感染细胞中的典型半衰期为30至45分钟。感染后早期，这些mRNA被去腺苷化并在5'端切割成富含AU的元件。残留部分的半衰期>3小时。在晚期（感染后6+ h），这些mRNA的降解依赖于ICP 27。A类和B类mRNA都在多聚核糖体中降解。C类包括未降解的mRNA。这些mRNA的产物包括tristetraprolin，一种应激诱导蛋白，其结合富含AU的元件并将mRNA与外来体隔离以进行3'至5'降解，以及GADD 452，一种转录因子。(iv)早期的报道表明VHS结合eIF 4F复合物的3种组分，即eIF 4 H、eIF 4 B和eIF 4AII和VP 16。我们的研究表明，VHS还与tristetraprolin、ICP 27发生物理相互作用，并通过VP 16与VP 22发生相互作用。研究提案的目的是测试A类和B类mRNA降解的模型。该模型提出，VHS降解多聚核糖体中的mRNA，如所报道的，它结合到eIF 4F的仍待定义的组分，并通过切割帽结构下游的稳定mRNA来模拟脱帽酶的功能。残留部分随后被隔离在P-体中并迅速降解。在B类mRNA的情况下，VHS结合与富含AU的元件mRNA结合的三曲脯氨酸，并切割富含AU的元件的5'端的mRNA。在不存在结合的三曲脯氨酸的情况下，残留的mRNA不会被外泌体迅速募集用于3'至5'降解。我们还建议区分ICP 27在感染后期使mRNA降解的2种替代潜在功能。最后，考虑到VHS的多种功能，问题是每种功能在多大程度上有助于HSV-1的致病潜力。