Dissection of the Functions of Herpes Simplex Virus ICPO

单纯疱疹病毒 ICPO 功能剖析

• 批准号: 7834052
• 负责人: Bernard Roizman
• 金额: $ 58.16万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2011-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7834052
• 关键词: Acceleration; Adaptor Signaling Protein; Address; Adult; Afferent Neurons; Antiviral Agents; Biology; Budgets; Cell Nucleus; Cells; Complement; Complex; Data; Development; Disease; Dissection; Drug Controls; Drug Design; Equipment; Event; Funding; Gene Expression; Gene Targeting; Genes; Genome; Grant; HDAC1 gene; HDAC2 gene; Health; Histones; Human; Human Resources; Individual; Infection; Integration Host Factors; Interferons; Investigation; Knowledge; Lesion; Letters; Lysine; Maintenance; Maps; Medical; MicroRNAs; Mus; Neuroglia; Neurons; Newborn Infant; Occupations; Persons; Pharmaceutical Preparations; Play; Positioning Attribute; Process; Protein Kinase; Proteins; Reagent; Recruitment Activity; Reporting; Research; Research Proposals; Resources; Role; Simplexvirus; Staging; Structure; Structure of trigeminal ganglion; Technology; Testing; Time; Transferase; Trauma; Viral; Viral Genes; Viral Genome; Viral Proteins; Virus; Wages; Work; attenuation; base; cost; design; drug resistant virus; genetic regulatory protein; immunosuppressed; in vivo; inhibitor/antagonist; interest; latent infection; mutant; public health relevance; research study; small molecule; ubiquitin ligase; viral DNA

DESCRIPTION (provided by applicant): Diseases and suffering caused by reactivations of latent Herpes Simplex Viruses (HSV) remain a major medical problem notwithstanding effective attenuation of disease by currently available antiviral drugs. The major problem facing development of specific treatment of latency is that in the course of the establishment of latency and its maintenance most of the viral genome is silenced by cellular proteins. What we do not know is the mechanism by which the infected neuron silences the viral genome. If we knew the mechanisms by which the virus is silenced, we could devise screens for small molecule drugs that either enhance the silent state and preclude reactivation or reactivate viral replication in the presence of antiviral drugs. This revision of the R37 CA78766 grant takes advantage of accrued knowledge to address this issue. The R37 CA78766 grant focuses entirely on ICP0, a herpes simplex virus regulatory protein that is emerging as a key regulator of viral gene expression in both productive infections and in the establishment and maintenance of latency. In brief, in the past 3 years we have made a fundamental discovery that ICP0 plays a key role early in productive infection in suppressing the cellular machinery whose main objective is to silence viral DNA. The machinery employed by the cells in its attempt to silence HSV DNA consists of a complex containing HDAC-1 or -2, CoREST, REST and LSD1. Of particular interest is the fact that CoREST and REST are known as repressors of neuronal genes in non neuronal cells. LSD1 - the lysine specific demethylase along with HDAC1 or HDAC2 play a key role in this process. This complex is disassembled by ICP0, phosphorylated by viral protein kinases and expelled from the nucleus. The discovery of the role of the HDAC1 or -2/CoREST/REST/LSD1 in suppressing viral gene expression in productively infected cells raises the interesting question whether components of this complex also act as repressors of viral gene expression during latency. This hypothesis is tested in Aim 1 of the Competitive Revision Application. The objective of the second aim is to define the role of a set of viral micro RNAs that target the genes encoding ICP0, ICP4 and ICP34.5. It has been suggested, but not proven, that these micro RNAs play a role in silencing key genes to enable the maintenance of the silent state of the viral genome during latency. There is, however, an alternative hypothesis based on observations that HSV down regulates the synthesis and function of regulatory proteins including ICP0 in the course of productive infection. The available data also support the hypothesis that the viral micro RNAs are components of this regulatory network. These aims are as follows: Aim 1. To test the hypothesis that components of the HDAC-1 or -2, CoREST, REST and LSD1 complex enable establishment of latency by silencing the HSV genome upon entry into the sensory neurons of trigeminal ganglia in vivo. Neuronal cells contain components of the complex described above. We plan to construct viruses that disrupt the complex in the same way that we have shown that in non neuronal cells disruption of this complex complements ?ICP0 minus viruses. In carefully controlled experiments we plan to determine whether disruption of the suppressor complex precludes or diminishes the establishment of latency. Aim 2; To determine whether the micro RNAs reported as potential candidates for suppression of ICP0 and ICP34.5 play a role in establishment of latency or whether they suppress the synthesis or function of ? ( immediate early) regulatory proteins ICP0, ICP22 and ICP4 or both. We have a large number of mutants in the domain of the HSV genome in which the micro RNAs map. It is relatively easy for us to construct viruses that fail to express the micro RNAs. These mutants and the restored wild-type viruses will be tested with respect to their ability to establish and maintain latent infections. The studies will be completed during the two year grant period. We anticipate that these studies will identify targets for development of drugs that control latency studies and contribute to the design of screens for small molecule inhibitors. PUBLIC HEALTH RELEVANCE: This revision of the R37 CA78766 grant takes advantage of accrued knowledge to ablate the diseases and suffering caused by reactivations of latent Herpes Simplex Viruses (HSV). The major problem facing development of specific treatment of latency is that in the course of the establishment of latency and its maintenance most of the viral genome is silenced by cellular proteins. If we could substantiate the proposed mechanisms by which the virus is silenced, we could devise screens for small molecule drugs that either enhance the silent state and preclude reactivation or reactivate viral replication in the presence of antiviral drugs.

描述（由申请人提供）：尽管目前可用的抗病毒药物有效地减弱了疾病，但由潜伏的单纯疱疹病毒（HSV）再激活引起的疾病和痛苦仍然是一个主要的医学问题。 潜伏期特异性治疗的发展面临的主要问题是，在潜伏期的建立和维持过程中，大多数病毒基因组被细胞蛋白质沉默。 我们不知道的是受感染的神经元使病毒基因组沉默的机制。如果我们知道病毒沉默的机制，我们就可以设计出筛选小分子药物的方法，这些药物可以增强沉默状态并阻止病毒的重新激活，或者在抗病毒药物存在的情况下重新激活病毒复制。R37 CA 78766补助金的这一修订利用了累积的知识来解决这个问题。 R37 CA 78766资助完全集中在ICP 0上，ICP 0是一种单纯疱疹病毒调节蛋白，正在成为生产性感染以及潜伏期建立和维持中病毒基因表达的关键调节因子。 简而言之，在过去的3年里，我们已经取得了一个基本的发现，即ICP 0在生产性感染的早期抑制细胞机制中起着关键作用，其主要目的是沉默病毒DNA。细胞试图沉默HSV DNA的机制由含有HDAC-1或HDAC-2、CoREST、REST和LSD 1的复合物组成。特别令人感兴趣的是，CoREST和REST被认为是非神经元细胞中神经元基因的阻遏物。LSD 1-赖氨酸特异性脱甲基酶沿着HDAC 1或HDAC 2在该过程中起关键作用。 该复合物被ICP 0分解，被病毒蛋白激酶磷酸化并从细胞核排出。 HDAC 1或-2/CoREST/REST/LSD 1在抑制生产性感染细胞中病毒基因表达中的作用的发现提出了一个有趣的问题，即该复合物的组分是否也在潜伏期期间作为病毒基因表达的阻遏物。 在竞争修订申请的目标1中对该假设进行了检验。 第二个目标是确定一组靶向编码ICP 0、ICP 4和ICP34.5的基因的病毒微小RNA的作用。 有人提出，但尚未证明，这些微小RNA在沉默关键基因中发挥作用，以使病毒基因组在潜伏期期间保持沉默状态。 然而，有一个替代假设，根据观察，HSV下调的合成和功能的调节蛋白，包括ICP 0在生产性感染的过程中。 现有的数据也支持这样的假设，即病毒microRNA是这个调控网络的组成部分。 这些目标如下：目标1。为了检验以下假设：HDAC-1或HDAC-2、CoREST、REST和LSD 1复合物的组分能够通过在体内进入三叉神经节的感觉神经元后沉默HSV基因组来建立潜伏期。神经元细胞含有上述复合物的成分。我们计划构建病毒，以同样的方式破坏复合物，我们已经表明，在非神经元细胞中，这种复合物的破坏补充？ICP 0减去病毒。在精心控制的实验中，我们计划确定抑制复合物的破坏是否会排除或减少潜伏期的建立。 目标2;为了确定报告为抑制ICP 0和ICP34.5的潜在候选者的微小RNA是否在建立潜伏期中发挥作用，或者它们是否抑制ICP 0和ICP34.5的合成或功能？ （立即早期）调节蛋白ICP 0、ICP 22和ICP 4或两者。 我们在HSV基因组的微RNA图谱中发现了大量的突变体。对我们来说，构建不表达微小RNA的病毒相对容易。将检测这些突变体和恢复的野生型病毒建立和维持潜伏感染的能力。 这些研究将在两年的资助期内完成。我们预计，这些研究将确定控制潜伏期研究的药物开发的目标，并有助于小分子抑制剂的筛选设计。 公共卫生相关性：R37 CA 78766补助金的这一修订利用了累积的知识来消融 由潜伏的单纯疱疹病毒（HSV）再激活引起的疾病和痛苦。 潜伏期特异性治疗的发展面临的主要问题是，在潜伏期的建立和维持过程中，大多数病毒基因组被细胞蛋白质沉默。 如果我们能够证实病毒沉默的机制，我们就可以设计小分子药物的筛选，这些药物可以增强沉默状态并阻止病毒的再激活，或者在抗病毒药物存在的情况下重新激活病毒复制。