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Vpr

电压

基本信息

批准号：
8705387
负责人：
YONG-HUI ZHENG
金额：
$ 19.19万
依托单位：
MICHIGAN STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-08-01 至 2017-07-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Vpr and vpx are two viral accessory genes that are expressed in primate lentiviruses, which are required for viral replication and disease progression in vivo. Vpr has two well-known activities in vitro: arrest of cell cycle in G2 phase and enhancement of viral replication in monocyte-derived macrophages. Sharing ~25% homology with Vpr, Vpx only has the viral enhancement activity. Although both Vpr and Vpx enhance HIV-1 replication, different mechanisms are involved. While Vpx counteracts the antiretroviral activity of SAMHD1, the mechanism of Vpr enhancement of viral replication is still unknown. Recently, we reported a potent HIV-1 restriction in the human CD4+ T cell line CEM.NKR (NKR), which was naturally isolated from the human T lymphoblastoid cell line CEM. Although NKR cells express both CD4 and CXCR4, HIV-1 replication is severely restricted from the 2nd round of replication. From the original NKR cells, we isolated three types of clones that show different levels of HIV-1 resistance: non-permissive (NP), semi-permissive (SP), and permissive (P). We then compared wild-type (WT) and Vpr-deficient ( Vpr) HIV-1 replication in these cells. In non-permissive cells, both WT and Vpr viruses were unable to replicate. Notably, a treatment with arsenic trioxide (As2O3) increased the WT virus replication by almost 1000-fold, but did not promote the Vpr virus replication. Similarly, although the WT virus could replicate in the semi-permissive and permissive cells, the Vpr virus replication was completely inhibited in the semi-permissive cells and significantly delayed in the permissive cells. These results suggest that Vpr is absolutely required for HIV-1 replication in the non-permissive and semi-permissive cells. Thus, we have identified Vpr-specific HIV-1 non-permissive human CD4+ T cell line, which represents an important progress in the Vpr field. Our objective is to decipher the mechanism of how Vpr enhances HIV-1 replication in NKR cells. Our hypothesis is that NKR cells may express a Vpr-sensitive restriction factor to block viral replication, or lack a Vpr-like positive factor to support viral replication. Our rationale is that these NKR cells provide a relevant model system for studying Vpr function, which will be useful for further characterization of Vpr activity in vivo. We propose three specific aims: 1) Delineate how Vpr enhances HIV-1 replication in NKR cells; 2) Identify the host factor in NKR cells that is responsible for Vpr- dependent HIV-1 replication; 3) Elucidate the relevance of HIV-1 restriction in NKR cells for HIV-1 biology. We will define the enigmatic role of Vpr in viral life cycle. The discovered new mechanism will be likely translated into innovative tools for antiretroviral therapy.

描述（由申请方提供）：Vpr和Vpx是在灵长类慢病毒中表达的两种病毒辅助基因，是体内病毒复制和疾病进展所必需的。Vpr在体外具有两种众所周知的活性：将细胞周期阻滞在G2期和增强单核细胞衍生的巨噬细胞中的病毒复制。Vpx与Vpr具有约25%的同源性，仅具有病毒增强活性。虽然Vpr和Vpx都能增强HIV-1的复制，但涉及不同的机制。虽然Vpx抵消了SAMHD 1的抗逆转录病毒活性，但Vpr增强病毒复制的机制仍不清楚。最近，我们报道了一个有效的HIV-1限制在人类CD 4 + T细胞系CEM.NKR（NKR），这是从人类T淋巴母细胞系CEM自然分离。虽然NKR细胞表达CD 4和CXCR 4，但HIV-1复制从第二轮复制开始就受到严格限制。我们从原始NKR细胞中分离出三种类型的克隆，它们显示出不同水平的HIV-1耐药性：非允许性（NP）、半允许性（SP）和允许性（P）。然后，我们比较了野生型（WT）和Vpr缺陷型（Vpr）HIV-1在这些细胞中的复制。在非允许细胞中，WT和Vpr病毒都不能复制。值得注意的是，用三氧化二砷（As 2 O3）处理使WT病毒复制增加了几乎1000倍，但没有促进Vpr病毒复制。类似地，尽管WT病毒可以在半容许和容许细胞中复制，但Vpr病毒复制在半容许细胞中被完全抑制并且在容许细胞中被显著延迟。这些结果表明，Vpr是HIV-1在非允许和半允许细胞中复制所必需的。因此，我们已经鉴定了Vpr特异性HIV-1非允许性人CD 4 + T细胞系，这代表了Vpr领域的重要进展。我们的目标是破译Vpr如何增强HIV-1在NKR细胞中复制的机制。我们的假设是，NKR细胞可能表达一种Vpr敏感的限制因子来阻断病毒复制，或者缺乏一种类似Vpr的限制因子。支持病毒复制的积极因素。我们的理由是，这些NKR细胞提供了一个相关的模型系统，研究Vpr功能，这将是有用的Vpr活性在体内的进一步表征。我们提出了三个具体目标：1）描述Vpr如何增强HIV-1在NKR细胞中的复制; 2）鉴定NKR细胞中负责Vpr依赖性HIV-1复制的宿主因子; 3）阐明NKR细胞中HIV-1限制与HIV-1生物学的相关性。我们将明确Vpr在病毒生命周期中的神秘作用。发现的新机制将可能转化为抗逆转录病毒疗法的创新工具。