Agnoprotein in JC virus virion biogenesis and replication

JC病毒病毒体生物发生和复制中的Agno蛋白

• 批准号: 7554846
• 负责人: MAHMUT SAFAK
• 金额: $ 36.25万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-04-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7554846
• 关键词: Acquired Immunodeficiency Syndrome; Affect; Antigens; Applications Grants; Astrocytes; Binding; Biogenesis; Brain; Capsid; Capsid Proteins; Cell Cycle Progression; Cells; Central Nervous System Diseases; Characteristics; Clinical; Code; Complex; Condition; DNA; DNA Binding; DNA biosynthesis; Disease; Disease Progression; Elements; Epidemic; Funding; Grant; Human; Immunocompromised Host; Immunosuppressive Agents; In Vitro; Individual; Infection; Intake; JC Virus; Large T Antigen; Leukoencephalopathy; Life Cycle Stages; Mediating; Mitosis; Molecular; Mutagenesis; Myelin; Myeloproliferative disease; Neuraxis; Oligodendroglia; PKC Phosphorylation Site; Patients; Phase Transition; Phosphoric Monoester Hydrolases; Phosphorylation; Play; Process; Progressive Multifocal Leukoencephalopathy; Protein Dephosphorylation; Protein Phosphatase 2A Regulatory Subunit PR53; Proteins; Public Health; Regulation; Research Project Grants; Role; Testing; Therapeutic; Viral; Viral Regulatory Proteins; Virion; Virus; Virus Replication; central nervous system demyelinating disorder; deletion analysis; design; genetic regulatory protein; helicase; in vivo; latent infection; leukemia/lymphoma; mutant; neurotropic; research study; transcription factor; viral DNA

DESCRIPTION (provided by applicant): JC virus (JCV) is an etiological agent of Progressive Multifocal Leukoencephalopathy (PML), a fatal demyelinating disease of the Central Nervous System (CNS), which is frequently seen in patients with underlying immunosuppressive conditions, including leukemia, lymphomas and AIDS. PML in the era of AIDS epidemic dramatically increased which makes it an AIDS defining condition. JCV establishes a sub-clinical latent infection in the body but upon reactivation, it enters the brain; and lytically and abortively infects oligodendrocytes (myelin producing cells) and astrocytes respectively. This infection results in an extensive myelin loss in CNS, which is the characteristic histopathological landmark of PML. This is a revised competitive renewal R01 grant application, entitled "Role of agnoprotein in JC virus life cycle". During the course of this funding period, we established that agnoprotein of JCV plays important regulatory roles in JCV life cycle through molecular interactions with a cellular transcription factor, YB-1 and the viral regulatory protein, large T antigen. It deregulates cell cycle progression, in which agnoprotein positive cells largely accumulate at G2/M phase transition. We also demonstrated the involvement of the coding region of agnoprotein in regulation of JCV life cycle by deletion analysis. In this regard, we showed that agnoprotein- coding region contains important cis-acting DNA elements to which specific transcription factors bind and contribute the viral life cycle. Furthermore, analysis of the PKC phosphorylation sites of this protein by mutagenesis revealed the fact that phosphorylation plays a critical role in the function of agnoprotein, because none of the phosphorylation mutants (Ser7, Ser11 and Thr21 to Ala) was able to continue viral replication cycle due to a limited replication and empty capsid formation. Moreover, our recent findings also showed that phosphorylated form of agnoprotein is targeted by a Ser/Thr phosphatase, PP2A, for dephosphorylation and this can be inhibited by JCV small t antigen, suggesting a functional ternary complex formation between these three proteins. These findings collectively have provided us a strong rationale to further study the functions of agnoprotein and led us to hypothesize that agnoprotein plays critical regulatory roles in JCV virion biogenesis and therefore in the progression of PML. As such, the understanding of the molecular mechanisms in which agnoprotein is involved in JCV replication is central to unravel the molecular mechanisms that are critical for the disease progression so that we would be able to develop effective therapeutic strategies against this disease. Thus, to further examine our hypothesis, we propose to i) investigate the molecular mechanisms by which agnoprotein regulates both the virion formation and large T antigen-mediated viral DNA replication; and ii) examine the impact of both PP2A and Sm t-Ag on agnoprotein functions during this process. PUBLIC HEALTH RELEVANCE Progressive multifocal leukoencephalopathy (PML), a white mater disease of the central nervous system, is caused by a human neurotropic polyomavirus, JC virus (JCV). This disease mostly affects immunocompromised patients with underlying disorders such as lymphoproliferative and myeloproliferative diseases and acquired immunodeficiency syndrome, (AIDS). In this research project, we are proposing experiments to understand the role of one of JCV regulatory proteins, agnoprotein in viral biogenesis, which may allow us to design effective therapeutic strategies to curb the disease in affected individuals.

描述（由申请方提供）：JC病毒（JCV）是进行性多灶性白质脑病（PML）的病原体，PML是一种中枢神经系统（CNS）的致死性脱髓鞘疾病，常见于基础免疫抑制疾病（包括白血病、淋巴瘤和AIDS）患者。PML在艾滋病流行的时代急剧增加，使其成为艾滋病的定义条件。JCV在体内建立亚临床潜伏感染，但在再活化后，它进入大脑;并分别裂解和流产感染少突胶质细胞（髓鞘产生细胞）和星形胶质细胞。这种感染导致CNS中广泛的髓鞘丢失，这是PML的特征性组织病理学标志。这是一个修订的竞争性更新R 01赠款申请，题为“在JC病毒生命周期中的作用”。在此资助期间，我们确定了JCV的未知蛋白通过与细胞转录因子YB-1和病毒调节蛋白大T抗原的分子相互作用在JCV生命周期中发挥重要的调节作用。它使细胞周期进程失调，其中未知蛋白阳性细胞大量积聚在G2/M期转换。我们还通过缺失分析证明了未知蛋白编码区参与了JCV生命周期的调控。在这方面，我们表明，未知蛋白编码区含有重要的顺式作用的DNA元件，特异性转录因子结合并有助于病毒的生命周期。此外，通过突变分析该蛋白的PKC磷酸化位点揭示了磷酸化在未知蛋白的功能中起关键作用的事实，因为没有磷酸化突变体（Ser 7，Ser 11和Thr 21至Ala）能够继续病毒复制周期，这是由于有限的复制和空衣壳形成。此外，我们最近的研究结果还表明，磷酸化形式的未知蛋白的丝氨酸/苏氨酸磷酸酶，PP 2A，靶向去磷酸化，这可以抑制JCV小t抗原，这表明这三个蛋白质之间的功能三元复合物的形成。这些发现共同为我们提供了一个强有力的理由，进一步研究的功能，未知蛋白，并使我们假设，未知蛋白在JCV病毒粒子的生物发生，因此在PML的进展中发挥关键的调节作用。因此，对未知蛋白参与JCV复制的分子机制的理解对于解开疾病进展的关键分子机制至关重要，以便我们能够开发针对这种疾病的有效治疗策略。因此，为了进一步验证我们的假设，我们建议i）研究未知蛋白调节病毒粒子形成和大T抗原介导的病毒DNA复制的分子机制; ii）研究PP 2A和Sm t-Ag在此过程中对未知蛋白功能的影响。公共卫生相关性进行性多灶性白质脑病（PML）是一种中枢神经系统的白色脑膜疾病，由人类嗜神经性多瘤病毒JC病毒（JCV）引起。这种疾病主要影响免疫功能低下的患者，这些患者具有潜在的疾病，例如淋巴增生性和骨髓增生性疾病以及获得性免疫缺陷综合征（AIDS）。在这个研究项目中，我们提出了实验来了解JCV调节蛋白之一，即病毒生物发生中的未知蛋白的作用，这可能使我们能够设计有效的治疗策略来抑制受影响个体的疾病。