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（由申请方提供）：源自原代和转化CD 8+细胞的上清液具有强效HIV抑制活性。不是所有的活性都可以由已知的活性成分包括β 2-趋化因子来解释。使用设计用于特异性筛选在病毒进入后抑制原代巨噬细胞中的HIV的新因子的策略，我们鉴定并报道了在HVS转化的CD 8 + T细胞系K#1 50 K的细胞培养基中发现的蛋白质胸腺素原α（ProT 1）具有强效的HIV-1抑制活性（41）。从CD 8+细胞上清液的HIV-1抑制级分中消耗天然ProT 1可消除抑制活性，支持其通过可溶性介质的抑制作用。ProT 1是一种丰富的酸性肽，据报道其定位于细胞核中并与细胞增殖和转录激活相关。ProT 1抑制HIV-1复制，其活性具有靶细胞特异性，抑制作用主要发生在病毒整合后（41）。天然和重组ProT 1蛋白有效抑制巨噬细胞和树突状细胞中HIV-1 LTR驱动的基因表达。ProT 1介导的抑制整合HIV-1的机制尚不清楚。我们的假设是外源性ProT 1与细胞表面受体相互作用，触发抑制HIV-1基因表达的信号转导途径。这种效应的细胞特异性是由于细胞特异性转录因子参与了HIV复制。为了设计一种方法来理解整合后ProT 1抑制HIV-1的机制，需要解决一些初步问题，如本R-21提案所述。这些包括活性和细胞摄取/核定位之间的关系，在观察到的抑制和HIV生命周期中的后整合步骤被抑制的主要信号传导途径的作用。这些基本问题的答案将允许实验方法的合理设计，以确定该宿主限制性途径的作用机制。这种初始方法将利用在其他系统中确定的关于ProT 1的功能和功能结构域的信息。了解ProT 1抑制HIV-1前病毒的机制并鉴定其抗HIV-1活性结构域将为开发针对整合HIV-1的治疗药物开辟新的途径。HIV-1感染免疫系统细胞后，病毒基因组整合到宿主基因组中。在病毒生命周期的整合后步骤的病毒复制导致产生新的病毒颗粒，所述病毒颗粒从受感染的细胞脱落并继续感染其他细胞。这对于感染的巨噬细胞特别重要，巨噬细胞可以排出病毒颗粒长达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。我们最近已经表明，人类蛋白质胸腺素原α（ProT 1）非常有效地干扰HIV-1在其生命周期的整合后步骤，导致病毒生产的减少。研究ProT 1在HIV-1抑制中的作用机制将导致开发可以阻断病毒生命周期这一重要步骤的治疗方法。