Regulatory Roles of Agnoprotein in Biology of JC virus

Agno蛋白在JC病毒生物学中的调节作用

基本信息

批准号：
8922270
负责人：
MAHMUT SAFAK
金额：
$ 39万
依托单位：
TEMPLE UNIV OF THE COMMONWEALTH
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-04-01 至 2020-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8922270
关键词：
Acquired Immunodeficiency Syndrome Alternative Splicing Amino Acids Amphipathic Alpha Helix Antibodies Applications Grants Attention Autoimmune Diseases Binding Biochemical Biogenesis Biological Biological Assay Biological Process Biology Brain Cell Nucleus Cells Central Nervous System Diseases Clinical Code Crohn&apos s disease Cytoplasm DNA biosynthesis Defect Dimerization Disease Elements Epidemic Event Genetic Goals Grant HIV-1 Immunocompromised Host Immunosuppressive Agents In Vitro Incidence Individual Investigation JC Virus Large T Antigen Leucine Life Cycle Stages Maintenance Maps Mediating Messenger RNA Molecular Multiple Sclerosis Mutagenesis Mutation Myelin Neuraxis Neuroglia Neurologic Manifestations Nuclear Export Oligodendroglia Open Reading Frames Patients Pattern Pharmaceutical Preparations Play Population Process Progressive Multifocal Leukoencephalopathy Protein Export Pathway RNA RNA Recognition Motif RNA Sequences RNA Splicing Regulation Reporting Risk Factors Role Signal Transduction Structure Technology Tertiary Protein Structure Therapeutic Transcript Viral Virus Replication Wit alpha helix antigen binding base experience genetic regulatory protein in vivo inhibitor/antagonist insight interest latent infection leptomycin B leukemia/lymphoma mutant natalizumab novel patient population protein function protein protein interaction public health relevance rev Protein transcriptome sequencing viral DNA viral RNA

项目摘要

DESCRIPTION (provided by applicant): JC virus (JCV) causes a fatal disease in the central nervous system (CNS), known as progressive multifocal leukoencephalopathy (PML) in patients with underlying immunosuppressive conditions, including leukemia, lymphoma and AIDS. PML in the era of the AIDS epidemic dramatically increased and is an AIDS associated illness. This disease is also steadily increasing among patients with autoimmune disorders, such as multiple sclerosis and Crohn's disease, who are treated with antibody-based drugs (natalizumab), which makes JCV as a risk factor for autoimmune disease populations. Our lab has considerable experience in studying the unique biology of JCV for a number of years. In recent years, however, we have focused our attention to the investigation of the functional roles of one of its regulatory proteins, Agnoprotein (Agno) (71 aa long), which is shown to play critical roles in the viral life cycle. In the absence of its expression, JCV is unable to sustain its productive cycle. We have made significant advances in understanding the biological functions of Agno in recent years. For instance, (i) Agno was found to form highly stable dimeric/oligomeric (multimeric) structures in vitro and in vivo and this is mediated by amino acids spanning from 28 to 39 which forms an amphipathic alpha-helix confirmed by our recent NMR studies. (ii) Agno undergoes a rapid degradation process and the viral replication levels significantly decrease if the alpha- helix region is altered by mutagenesis. (iii) Interestingly, the dimerization domain mutants were found to have profound defects in the alternative splicing process of the viral transcripts, suggesting new roles for Agno in regulation of viral RNA splicing. The initial splicing studies wit the dimerization domain mutants even led to the discovery of a novel and previously unpredicted open reading frame (new ORF) for JCV late transcripts, the significance of which needs to be uncovered for the JCV replication cycle. (iv) Moreover, Agno was also found to specifically bind to the JCV RNA and harbors a predicted nuclear export signal (NES). Furthermore, it specifically interacts with CRM1 and accumulates in the nucleus of the infected cells if they are treated with a CRM1 specific inhibitor, Leptomycin B. These findings also suggest novel regulatory roles for Agno in nucleo- cytoplasmic export of the viral RNA in a CRM1-dependent manner. Collectively, these findings provide us with a strong rationale to further investigate the novel regulatory roles of Agno in splicing and the nucleo- cytoplasmic export of JCV transcripts; and allowed us formulate our central hypothesis: Agno and its multimeric forms play important regulatory roles in the alternative splicing of the viral transcrips and their CRM1-dependent nucleo-cytoplasmic export. We propose to examine our hypothesis by thoroughly investigating the molecular mechanism(s) of the regulation of the both events by Agno. 1

描述（由适用提供）：JC病毒（JCV）在中枢神经系统（CNS）中引起致命疾病，称为潜在的免疫抑制疾病的患者，包括白血病，淋巴瘤和AIDS。艾滋病流行时代的PML急剧增加，是艾滋病相关的疾病。自身免疫性疾病患者（例如多发性硬化症和克罗恩病）的这种疾病也在稳步增加，这些疾病接受了基于抗体的药物（Natalizumab），这使JCV成为自身免疫性疾病群体的危险因素。我们的实验室已经考虑了研究JCV独特生物学多年的经验。然而，近年来，我们将注意力集中在研究其调节蛋白之一的功能作用，即agnoprotein（Agno）（71 AA长），该蛋白（71 AA长）在病毒生命周期中起着至关重要的作用。在没有表达的情况下，JCV无法维持其生产周期。近年来，我们在理解Agno的生物学功能方面取得了重大进步。例如，发现（i）Agno在体外和体内形成高度稳定的二聚体/寡聚（多聚体）结构，这是由28至39的氨基酸介导的，该氨基酸跨越至39，这形成了我们最近的NMR研究证实的两亲α-螺旋。（ii）如果诱变改变了α-螺旋区域，则AGNO经历了快速降解过程，病毒复制水平会显着降低。（iii）有趣的是，在病毒转录本的替代剪接过程中发现二聚域突变体具有深刻的缺陷，这表明AGNO在调节病毒RNA剪接方面的新作用。最初的剪接研究具有二聚化结构域突变体，甚至导致了针对JCV后期转录本的新颖且以前未预测的开放阅读框（新的ORF），在JCV复制周期中需要发现其重要性。（iv）此外，还发现Agno特异性结合了JCV RNA，并具有预测的核输出信号（NES）。此外，如果用CRM1特异性抑制剂治疗了感染细胞的细胞核，它与CRM1特别相互作用，并积聚在感染细胞的细胞核中。这些发现还表明，在CRM1依赖性依赖性的情况下，AGNO在病毒RNA的核胞质中的新调节作用。总的来说，这些发现为我们提供了强有力的理由，以进一步研究Agno在剪接和JCV转录本的核质质出口中的新调节作用。并使我们提出了中心假设：AGNO及其多个中等形式在病毒式转纸的替代剪接及其CRM1依赖性核胞质出口中起着重要的调节作用。我们建议通过彻底研究Agno调节两种事件的分子机制来检查我们的假设。 1