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: 8113340
• 负责人: MAHMUT SAFAK
• 金额: $ 36.75万
• 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/8113340
• 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; DNA; DNA Binding; DNA biosynthesis; Disease; Disease Progression; Elements; Epidemic; Funding; Grant; Health; Human; Immunocompromised Host; Immunosuppressive Agents; In Vitro; Individual; Infection; Intake; JC Virus; Large T Antigen; Life Cycle Stages; Lymphoproliferative Disorders; Mediating; Mitosis; Molecular; Mutagenesis; Myelin; Myeloproliferative disease; Neuraxis; Oligodendroglia; PKC Phosphorylation Site; Patients; Phase Transition; Phosphoric Monoester Hydrolases; Phosphorylation; Play; Polyomavirus; Process; Progressive Multifocal Leukoencephalopathy; Protein Dephosphorylation; Protein Phosphatase 2A Regulatory Subunit PR53; Proteins; Regulation; Research Project Grants; Role; Testing; Therapeutic; Viral; Viral Regulatory Proteins; Virion; 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; 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; DNA; DNA Binding; DNA biosynthesis; Disease; Disease Progression; Elements; Epidemic; Funding; Grant; Health; Human; Immunocompromised Host; Immunosuppressive Agents; In Vitro; Individual; Infection; Intake; JC Virus; Large T Antigen; Life Cycle Stages; Lymphoproliferative Disorders; Mediating; Mitosis; Molecular; Mutagenesis; Myelin; Myeloproliferative disease; Neuraxis; Oligodendroglia; PKC Phosphorylation Site; Patients; Phase Transition; Phosphoric Monoester Hydrolases; Phosphorylation; Play; Polyomavirus; Process; Progressive Multifocal Leukoencephalopathy; Protein Dephosphorylation; Protein Phosphatase 2A Regulatory Subunit PR53; Proteins; Regulation; Research Project Grants; Role; Testing; Therapeutic; Viral; Viral Regulatory Proteins; Virion; 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（由申请人提供）：JC病毒（JCV）是进行性多灶性白血病患病（PML）的病因学药，这是一种致命的中枢神经系统脱发疾病（CNS），在包括潜在的免疫抑制病（包括白血病，淋巴瘤和助理）的患者中经常见到这种疾病。艾滋病时代的PML急剧增加，这使其成为定义条件的艾滋病。 JCV在体内建立了一个次链的潜在感染，但重新激活后，它进入了大脑。并且分别通过裂解和堕胎感染了少突胶质细胞（髓磷脂产生细胞）和星形胶质细胞。这种感染导致中枢神经系统的大量髓磷脂损失，这是PML的特征性组织病理学地标。这是经过修订的竞争性更新R01赠款应用，名为“ Agnoprotein在JC病毒生命周期中的作用”。在此资助期间，我们确定JCV的a蛋白通过与细胞转录因子YB-1和病毒调节蛋白大型T抗原的分子相互作用通过分子相互作用在JCV生命周期中起重要的调节作用。它消除了细胞周期进程，其中毒素阳性细胞在G2/m相变大大积累。我们还通过缺失分析证明了Agnoprotein的编码区域参与JCV生命周期的调节。在这方面，我们表明，抗蛋白编码区包含重要的顺式作用DNA元素，特定转录因子结合并促进病毒生命周期。此外，通过诱变对这种蛋白质的PKC磷酸化位点的分析表明，磷酸化在促蛋白的功能中起着至关重要的作用，因为没有任何磷酸化突变体（Ser7，Ser7，Ser11和Thr21 to Ala to Ala）能够继续由于有限的复制和限量复制而继续进行病毒复制周期。此外，我们最近的发现还表明，磷酸化形式的致动蛋白是由Ser/Thr磷酸酶PP2A靶向的，用于去磷酸化，这可以被JCV小型T抗原抑制，这表明这三种蛋白质之间的功能性三络合物形成。这些发现共同为我们提供了一个有力的理由，可以进一步研究毒素的功能，并使我们假设a蛋白在JCV病毒率生物发生以及因此在PML的进展中起关键的调节作用。因此，对JCV复制涉及的分子机制的理解对于揭示对疾病进展至关重要的分子机制至关重要，因此我们将能够制定针对这种疾病的有效治疗策略。因此，为了进一步研究我们的假设，我们提出了i）研究分子机制，通过这些机制调节病毒素形成和大型T抗原介导的病毒DNA复制； ii）检查在此过程中，PP2A和SM T-AG对毒素功能的影响。公共卫生相关性渐进性多灶性白血病（PML）是中枢神经系统的白色孕妇病，是由人类神经性多瘤病毒（JC病毒（JCV））引起的。该疾病主要影响具有潜在疾病的免疫功能低下患者，例如淋巴增生性和骨髓增生性疾病，并获得了免疫缺陷综合征（AIDS）。在该研究项目中，我们提出了实验，以了解JCV调节蛋白之一，促蛋白在病毒生物发生中的作用，这可能使我们能够设计有效的治疗策略以遏制受影响个体的疾病。