Suppression of HIV-1 Infection by APOBEC3 Proteins in Primary Human CD4+ T Cells Is Associated with Inhibition of Processive Reverse Transcription as Well as Excessive Cytidine Deamination
Suppression of HIV-1 Infection by APOBEC3 Proteins in Primary Human CD4+ T Cells Is Associated with Inhibition of Processive Reverse Transcription as Well as Excessive Cytidine Deamination
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DOI：
10.1128/jvi.02587-12
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发表时间：
2013-02-01
影响因子：
5.4
通讯作者：
Malim, Michael H.
中科院分区：
文献类型：
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作者：
Gillick, Kieran;Pollpeter, Darja;Malim, Michael H.
人类免疫缺陷病毒1型（HIV-1）的Vif蛋白通过下调细胞编码的抗病毒APOBEC 3蛋白来促进病毒复制。这些蛋白质通过抑制病毒逆转录以及诱导新生病毒cDNA内的胞苷脱氨基作用来发挥其抑制作用。重要的是，这两种作用在人类CD 4（+）T细胞中尚未详细描述，导致对它们在HIV-1复制的天然细胞靶点中对病毒抑制的可能贡献存在争议。在这里，我们使用来自多个供体的CD 4（+）T细胞的野生型和Vif缺陷型病毒来检查APOBEC 3蛋白在自然表达水平下功能的后果。我们证明，APOBEC 3蛋白质赋予一个深刻的缺陷，逆转录从cDNA合成的初始阶段，以及过量的胞苷脱氨基（超突变）的DNA合成。使用来自转染细胞的病毒的实验和用于绘制cDNA的3'末端的新方法表明，逆转录的抑制并不限于几个特定位点，认为APOBEC 3蛋白阻碍酶促合成能力。对病毒cDNA突变谱的详细分析强烈暗示，一种特定的APOBEC 3蛋白APOBEC 3G提供了CD 4（+）T细胞中的大部分抗病毒表型，而APOBEC 3F和APOBEC 3D的作用不太显著。综上所述，我们得出结论，APOBEC 3蛋白的双重作用机制结合联合收割机提供比任何一种功能本身更有效的HIV-1限制。
The Vif protein of human immunodeficiency virus type 1 (HIV-1) promotes viral replication by downregulation of the cell-encoded, antiviral APOBEC3 proteins. These proteins exert their suppressive effects through the inhibition of viral reverse transcription as well as the induction of cytidine deamination within nascent viral cDNA. Importantly, these two effects have not been characterized in detail in human CD4(+) T cells, leading to controversies over their possible contributions to viral inhibition in the natural cell targets of HIV-1 replication. Here we use wild-type and Vif-deficient viruses derived from the CD4(+) T cells of multiple donors to examine the consequences of APOBEC3 protein function at natural levels of expression. We demonstrate that APOBEC3 proteins impart a profound deficiency to reverse transcription from the initial stages of cDNA synthesis, as well as excessive cytidine deamination (hypermutation) of the DNAs that are synthesized. Experiments using viruses from transfected cells and a novel method for mapping the 3' termini of cDNAs indicate that the inhibition of reverse transcription is not limited to a few specific sites, arguing that APOBEC3 proteins impede enzymatic processivity. Detailed analyses of mutation spectra in viral cDNA strongly imply that one particular APOBEC3 protein, APOBEC3G, provides the bulk of the antiviral phenotype in CD4(+) T cells, with the effects of APOBEC3F and APOBEC3D being less significant. Taken together, we conclude that the dual mechanisms of action of APOBEC3 proteins combine to deliver more effective restriction of HIV-1 than either function would by itself.