Evidence for Vpr-dependent HIV-1 replication in human CD4+ CEM.NKR T-cells.
Evidence for Vpr-dependent HIV-1 replication in human CD4+ CEM.NKR T-cells.
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DOI：
10.1186/1742-4690-9-93
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发表时间：
2012-11-07
期刊：
影响因子：
3.3
通讯作者：
Zheng YH
中科院分区：
文献类型：
--
作者：
Zhou T;Dang Y;Baker JJ;Zhou J;Zheng YH
Vpr仅在灵长类慢病毒中表达，并有助于体内病毒复制和疾病进展。HIV-1 Vpr在体外具有两种主要活性：将细胞周期阻滞在G2期（G2阻滞）和增强巨噬细胞中的病毒复制。之前，我们报道了在人CD 4 + CEM.NKR（NKR）T细胞中的有效HIV-1限制，其中野生型（WT）HIV-1复制被抑制近1，000倍。从亲本NKR细胞中，我们通过有限稀释分离出8个克隆。这些克隆对WT HIV-1感染表现出三种水平的抗性：非允许性（NP）、半允许性（SP）和允许性（P）。在这里，我们比较了WT，Vif缺陷型，Vpr缺陷型和Vpu缺陷型病毒在这些细胞中的复制。虽然WT和Vpu缺陷型病毒都可以在允许和半允许克隆中复制，但Vif缺陷型和Vpr缺陷型病毒的复制完全受到限制。APOBEC 3G（A3 G）胞苷脱氨酶在NKR细胞中的表达解释了为什么HIV-1复制需要Vif而不是Vpr。当在半容许细胞中将Vpr缺陷型病毒生命周期与WT病毒生命周期进行比较时，发现Vpr缺陷型病毒可以进入细胞并产生含有适当加工的Gag和Env蛋白的病毒体，但这些病毒体在下一轮感染期间显示出低得多的逆转录效率。此外，尽管病毒复制在非允许细胞中受到限制，但用三氧化二砷（As 2 O3）处理可以完全恢复WT，但不能恢复Vpr缺陷型病毒复制。此外，破坏Vpr与其辅因子DCAF 1的结合和/或诱导G2阻滞活性并不破坏Vpr在NKR细胞中增强HIV-1复制的活性。这些结果表明，HIV-1在NKR细胞中的复制是Vpr依赖性的。Vpr可能通过克服病毒复制早期阶段的阻断而促进HIV-1从第2周期开始复制;并且这种活性不需要DCAF 1和G2阻滞。对这一机制的进一步研究将为Vpr在HIV-1生命周期中的功能提供新的认识。
Vpr is exclusively expressed in primate lentiviruses and contributes to viral replication and disease progression in vivo. HIV-1 Vpr has two major activities in vitro: arrest of cell cycle in the G2 phase (G2 arrest), and enhancement of viral replication in macrophages. Previously, we reported a potent HIV-1 restriction in the human CD4+ CEM.NKR (NKR) T cells, where wild-type (WT) HIV-1 replication was inhibited by almost 1,000-fold. From the parental NKR cells, we isolated eight clones by limiting dilution. These clones showed three levels of resistance to the WT HIV-1 infection: non-permissive (NP), semi-permissive (SP), and permissive (P). Here, we compared the replication of WT, Vif-defective, Vpr-defective, and Vpu-defective viruses in these cells. Although both WT and Vpu-defective viruses could replicate in the permissive and semi-permissive clones, the replication of Vif-defective and Vpr-defective viruses was completely restricted. The expression of APOBEC3G (A3G) cytidine deaminase in NKR cells explains why Vif, but not Vpr, was required for HIV-1 replication. When the Vpr-defective virus life cycle was compared with the WT virus life cycle in the semi-permissive cells, it was found that the Vpr-defective virus could enter the cell and produce virions containing properly processed Gag and Env proteins, but these virions showed much less efficiency for reverse transcription during the next-round of infection. In addition, although viral replication was restricted in the non-permissive cells, treatment with arsenic trioxide (As2O3) could completely restore WT, but not Vpr-defective virus replication. Moreover, disruption of Vpr binding to its cofactor DCAF1 and/or induction of G2 arrest activity did not disrupt the Vpr activity in enhancing HIV-1 replication in NKR cells. These results demonstrate that HIV-1 replication in NKR cells is Vpr-dependent. Vpr promotes HIV-1 replication from the 2nd cycle likely by overcoming a block at early stage of viral replication; and this activity does not require DCAF1 and G2 arrest. Further studies of this mechanism should provide new understanding of Vpr function in the HIV-1 life cycle.
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