Prothymosin-Alpha- A Novel Antiviral Restriction Factor

胸腺素-α-一种新型抗病毒限制因子

• 批准号: 7418426
• 负责人: Mary E. Klotman
• 金额: $ 25.43万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-25 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7418426
• 关键词: Accounting; Address; Amino Acids; Antiviral Agents; Biological Assay; CD8-Positive T-Lymphocytes; CD8B1 gene; CREB1 gene; Cell Line; Cell Nucleus; Cell Proliferation; Cell Surface Receptors; Cell surface; Cells; Culture Media; Cyclic AMP; Dactinomycin; Data; Dendritic Cells; Development; Down-Regulation; Endocytosis; Event; Experimental Designs; Gene Expression; Genes; Genetic Transcription; Genome; HIV; HIV-1; Histones; Human; Immune system; In Vitro; Investigation; Label; Lead; Life Cycle Stages; Localized; MAP Kinase Gene; MAPK14 gene; Mediating; Mediator of activation protein; Northern Blotting; Nuclear; Nuclear Extract; Nuclear Localization Signal; Pathway interactions; Peptides; Pharmaceutical Preparations; Phosphorylation; Production; Proteins; RNA; RNA chemical synthesis; Recombinants; Reporting; Role; Running; Signal Pathway; Signal Transduction Pathway; Specificity; System; T-Lymphocyte; Therapeutic; Transcriptional Activation; Viral; Viral Genome; Virus Shedding; Work; chemokine; design; inhibitor/antagonist; macrophage; novel; particle; prothymosin alpha; therapeutic target; transcription factor; uptake

DESCRIPTION (provided by applicant): Supernatants derived from primary as well as transformed CD8+ cells have potent HIV inhibitory activity. Not all of that activity can be accounted for by known active components including 2-chemokines. Using a strategy designed to screen specifically for novel factors that inhibit HIV in primary macrophages following viral entry we identified and have reported that prothymosin alpha (ProT1), a protein found in the cell-culture media of the HVS-transformed CD8+ T-cell-line, K#1 50K, has potent HIV-1 inhibitory activity (41). Depletion of native ProT1 from an HIV-1 inhibitory fraction of CD8+ cell supernatants removes the inhibitory activity, supporting its role in inhibition via soluble mediators. ProT1 is an abundant, acidic peptide that has been reported to be localized in the nucleus and associated with cell proliferation and activation of transcription. ProT1 suppresses HIV-1 replication, its activity is target-cell specific and inhibition predominantly occurs following viral integration (41). Native and recombinant ProT1 protein potently inhibit HIV-1 LTR-driven gene expression in macrophages and dendritic cells. The mechanism(s) of ProT1 -mediated suppression of integrated HIV-1 is not yet known. Our hypothesis is that exogenous ProT1 interacts with a cell surface receptor triggering signal transduction pathways that inhibit HIV-1 gene expression. The cell specificity of the effect is due to the involvement of cell specific transcription factors in HIV replication. In order to design an approach to understanding the mechanism of the post integrational suppression of HIV-1 by ProT1 some initial questions need to be addressed as outlined in this R-21 proposal. These include the relationship between activity and cellular uptake/nuclear localization, the role of major signaling pathways in the observed inhibition and the post-integration step in the HIV life cycle that is inhibited. Answers to these basic questions will allow the rational design of an experimental approach to determine the mechanism of action of this host restriction pathway. This initial approach will take advantage of what has been determined in other systems regarding function and functional domains of ProT1. Understanding the mechanism of suppression of proviral HIV-1 by ProT1 and identification of the active anti-HIV-1 domain(s) of this molecule will open new avenues for developing therapeutics targeting integrated HIV-1.After the human immunodeficiency virus type-1 (HIV-1) infects cells of the immune system the viral genome becomes integrated into the host genome. Viral replication at the post-integration step of the viral life cycle leads to production of new viral particles that are shed from the infected cell and go on to infect other cells. This is of particular importance with infected macrophages which can shed viral particles for up to 2-3 month. Post-integration events in HIV-1 life cycle are difficult to interrupt because there are very few drugs able to interfere with this part of the viral life cycle. We have recently shown that the human protein prothymosin alpha (ProT1) very effectively interferes with HIV-1 at the post-integration step of its life cycle leading to decrease of viral production. Investigation of the mechanism of action of ProT1 in HIV-1 suppression will lead to the development of therapeutics that can block this important step of the viral life cycle.

描述（由申请人提供）：源自原发性和转化的CD8+细胞的上清液具有有效的HIV抑制活性。并非所有这些活动都可以由已知的活性组件（包括2个化学因子）来解释。使用旨在筛查的策略，专门针对病毒进入后抑制原发性巨噬细胞中HIV的新因素，我们确定并报告说，在HVS转换的CD8+ T-Cell-line的细胞培养培养基中发现了蛋白质（Prot1），K＃1 50K具有有效的HIV-1 50K，具有有效的HIV-1抑制活性（41）。从CD8+细胞上清液的HIV-1抑制分数中对天然PROT1的耗竭可消除抑制活性，从而支持其通过可溶性介质在抑制中的作用。 Prot1是一种丰富的酸性肽，据报道已定位在细胞核中，与细胞增殖和转录激活有关。 PROT1抑制HIV-1复制，其活性是靶细胞特异性和抑制作用，主要发生在病毒整合后（41）。天然和重组Prot1蛋白在巨噬细胞和树突状细胞中有效抑制HIV-1 LTR驱动的基因表达。 Prot1介导的对综合HIV -1的抑制的机制尚不清楚。我们的假设是，外源ProT1与抑制HIV-1基因表达的信号转导途径的细胞表面受体相互作用。该作用的细胞特异性是由于细胞特异性转录因子在HIV复制中的参与所致。为了设计一种方法来理解ProT1对HIV-1的整合抑制的机制，需要解决此R-21建议中概述的一些初始问题。其中包括活性与细胞摄取/核定位之间的关系，主要信号通路在观察到的抑制和抑制HIV生命周期中的整体步骤中的作用。这些基本问题的答案将允许实验方法的合理设计来确定该宿主限制途径的作用机理。这种初始方法将利用其他系统中有关Prot1功能和功能域的确定的优势。了解通过ProT1抑制病毒HIV-1的机理，并鉴定该分子的活性抗HIV-1结构域将开辟新的途径，以开发靶向综合HIV-1的疗法。随后，人类免疫缺陷病毒类型-1型（HIV-1）感染免疫系统的细胞，即免疫基因系统，病毒基因的病毒基因将成为宿主基因组成的基因组。病毒生命周期的整合性步骤的病毒复制导致产生从感染细胞中脱离的新病毒颗粒并继续感染其他细胞。这对于感染的巨噬细胞尤其重要，可以将病毒颗粒释放长达2-3个月。 HIV-1生命周期中的整合后事件很难中断，因为很少有药物能够干扰病毒生命周期的这一部分。我们最近表明，在其生命周期的整合性步骤中，人类蛋白蛋白α（ProT1）非常有效地干扰了HIV-1，导致病毒产生的降低。对HIV-1抑制中ProT1作用机理的研究将导致可以阻止病毒生命周期的这一重要步骤的治疗剂的发展。