Virulence Mechanisms of Viral Bcl-2 Homologs

病毒 Bcl-2 同源物的毒力机制

• 批准号: 8026437
• 负责人: JOHN C REED
• 金额: $ 47.75万
• 依托单位: SANFORD BURNHAM PREBYS MEDICAL DISCOVERY INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-12-01 至 2015-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8026437
• 关键词: Address; Adenoviruses; Apoptosis; Apoptotic; Autophagocytosis; BCL2 gene; Binding; Biological Models; Caspase-1; Cells; Cultured Cells; DNA Viruses; Disease; Engineering; Family; Family member; Foundations; Genes; Genome; Goals; Herpesviridae; Homologous Gene; Host Defense; Host Defense Mechanism; Immune response; Inflammation; Inflammatory; Investigation; Knock-in Mouse; Laboratories; Learning; Mediating; Mediator of activation protein; Morbidity - disease rate; Mus; Mutation; Natural Immunity; Pathogenicity; Play; Poxviridae; Process; Protein Family; Proteins; Role; Site; Testing; Therapeutic; Vaccinia virus; Viral; Viral Interference; Viral Proteins; Virulence; Virus; Virus Diseases; base; cytokine; defense response; gammaherpesvirus; human disease; in vivo; inhibition of autophagy; inhibitor/antagonist; member; mortality; mutant; novel; novel strategies; pathogen

DESCRIPTION (provided by applicant): DNA viruses are responsible for extensive morbidity and mortality on a worldwide basis. Viral homologs of anti-apoptotic Bcl-2 family proteins are encoded in the genomes of several classes of DNA viruses. In Vaccinia Virus (VV), a poxvirus-family member that has served as a paradigm for investigations of many aspects of host-pathogen interactions, at least two viral Bcl-2 genes have been identified, F1L and N1L. Neither F1L nor N1L is required for VV infection or replication, but both of these genes make strong contributions to virulence in vivo. Thus, these viral Bcl-2 (vBcl-2) homologs are critically important for in vivo viral pathogenicity of VV. While the anti-apoptotic activity of F1L and N1L is an obvious candidate for explaining their contribution to viral virulence, we have discovered that F1L and N1L have additional functions that include binding to and suppressing the pro-inflammatory actions of NLR-family proteins, important mediators of innate immunity. Viral Bcl-2 proteins are also known to bind Beclin and suppress autophagy, recently recognized as a host defense mechanism against pathogens. We hypothesize the vBcl-2 homologs are multifunctional proteins that utilize 3 discrete mechanisms to thwart host defense mechanisms: (a) suppression of apoptosis; (b) inhibition of autophagy; and (c) interference with NLR-mediated innate immune responses. The hypothesis that we will test is that neutralization by vBcl-2 proteins of each of these 3 classes of host cell targets significantly contributes to virulence. Specifically, we will: (1) Produce site-specific mutations in F1L and N1L that selectively abolish their ability to interact with (a) pro-apoptotic Bcl-2 family proteins [apoptosis]; (b) NLRs [inflammation]; and (c) Beclin [autophagy]; (2) Test the effects of the engineered vBcl-2 proteins on apoptosis, inflammation, and autophagy in cultured cells; (3) Produce recombinant vaccinia viruses with knock-in of F1L and N1L mutants; and (4) Compare the virulence of these recombinant vaccinia viruses in mice. By using VV as a model system, our results will lay a foundation for understanding the role of viral Bcl-2 homologs in viral pathogenicity, thus serving as a paradigm for other DNA viruses that contain vBcl-2 genes and that cause debilitating human diseases. Also, by learning how viruses interfere with apoptosis, autophagy, and inflammation, the information generated may reveal novel strategies for mimicking aspects of vBcl-2 function in therapeutically useful ways for addressing disorders in which excessive apoptosis, autophagy, and inflammation play central roles. PUBLIC HEALTH RELEVANCE: DNA viruses are responsible for extensive morbidity and mortality on a worldwide basis, and yet few therapeutic options are available to counteract these pathogens. Viral homologs of anti-apoptotic Bcl-2 family proteins are encoded in the genomes of several classes of DNA viruses but their roles and mechanism in viral disease are largely unknown. The hypothesis that we will test is that neutralization by viral Bcl-2 proteins of specific classes of host cell proteins significantly contributes to virulence, which will lay a foundation for developing new strategies for viral therapeutics based on disrupting interactions of viral Bcl-2 proteins with their host cell targets.

描述（由申请人提供）：DNA病毒在全球范围内负责广泛的发病率和死亡率。抗凋亡Bcl-2家族蛋白的病毒同源物编码在几类DNA病毒的基因组中。在Vaccinia病毒（VV）中，是一种研究宿主 - 病原体相互作用的许多方面的孢子病毒家庭成员，至少已经确定了两个病毒BCl-2基因，F1L和N1L。 VV感染或复制不需要F1L和N1L，但是这两个基因都对体内的毒力做出了很大的贡献。因此，这些病毒Bcl-2（VBCL-2）同源物对于VV的体内病毒致病性至关重要。尽管F1L和N1L的抗凋亡活性是解释其对病毒毒力贡献的明显候选者，但我们发现F1L和N1L具有其他功能，包括与NLR家庭蛋白质的促炎作用结合并抑制了先天免疫的重要中介体，Insate Impunity的重要中介体。还已知病毒Bcl-2蛋白可以结合Beclin并抑制自噬，最近被认为是针对病原体的宿主防御机制。 我们假设VBCL-2同源物是使用3种离散机制来阻止宿主防御机制的多功能蛋白：（a）抑制凋亡； （b）抑制自噬； （c）干扰NLR介导的先天免疫反应。我们将测试的假设是，这三类宿主细胞靶标的VBCL-2蛋白中和可显着有助于毒力。具体而言，我们将：（1）在F1L和N1L中产生特定位点特异性突变，这些突变有选择地废除了它们与（a）促凋亡的Bcl-2家族蛋白相互作用的能力[凋亡]； （b）NLR [炎症]； （c）Beclin [自噬]； （2）测试工程化的VBCL-2蛋白对培养细胞中细胞凋亡，炎症和自噬的影响； （3）用F1L和N1L突变体敲入重组疫苗病毒； （4）比较这些重组疫苗病毒在小鼠中的毒力。通过使用VV作为模型系统，我们的结果将为了解病毒Bcl-2同源物在病毒致病性中的作用奠定基础，从而充当其他含有VBCL-2基因并导致人类疾病的DNA病毒的范例。同样，通过学习病毒如何干扰凋亡，自噬和炎症，产生的信息可能会揭示模仿VBCL-2功能在治疗有用的方法中，以解决过度凋亡，自动噬菌体和炎症的疾病中的疾病。 公共卫生相关性：DNA病毒在全球范围内负责广泛的发病率和死亡率，但很少有治疗选择可以抵消这些病原体。抗凋亡Bcl-2家族蛋白的病毒同源物在几种DNA病毒的基因组中编码，但它们在病毒疾病中的作用和机制在很大程度上尚不清楚。我们将测试的假说是，特定类型宿主细胞蛋白的病毒BCL-2蛋白中和显着促进毒力，这将为基础开发基于病毒疗法的新策略，基于破坏病毒Bcl-2蛋白与宿主细胞靶标的相互作用。