Restriction of HIV-1 by APOBEC3G is cytidine deaminase-dependent.
Restriction of HIV-1 by APOBEC3G is cytidine deaminase-dependent.
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DOI：
10.1016/j.virol.2009.02.026
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发表时间：
2009-05-10
期刊：
影响因子：
3.7
通讯作者：
Landau, Nathaniel R.
中科院分区：
文献类型：
--
作者：
Browne, Edward P.;Allers, Carolina;Landau, Nathaniel R.
胞苷脱氨基是APOBEC3G抑制HIV-1的主要机制；然而，一些研究报道APOBEC3G也通过一种独立于脱氨基的机制抑制病毒复制。利用活性位点APOBEC3G突变体，我们重新评估了脱氨酶非依赖性APOBEC3G介导的HIV-1限制的生物学相关性。→基因突变的APOBEC3G蛋白在CEM-SS T细胞和293T细胞中以生理水平稳定表达，并检测细胞对Δ病毒复制的支持能力。AS2和AS1/AS2突变体被有效地包装成病毒粒子，但在单周期或多周期HIV-1复制试验中，发现缺乏抗病毒活性。AS1突变体保持了脱氨酶活性，保持了接近野生型的抗病毒功能。为了确定APOBEC3G的抗病毒活性，建立了低水平表达野生型APOBEC3G并产生含有少至1-2个APOBEC3G分子的病毒粒子的细胞系。即使在拷贝数非常低的情况下，APOBEC3G也能显著降低传染性，这表明包装的APOBEC3G单个分子就能灭活病毒。APOBEC3G的高效价与催化限制机制是一致的，在该机制中，单个分子可以诱导一系列突变，但很难与脱氨酶非依赖的非催化机制相协调。对逆转录序列的分析表明，G→A突变是聚集的，可能反映了单个APOBEC3G分子的作用过程。我们得出结论，胞苷脱氨基是APOBEC3G限制HIV-1的机制。
Cytidine deamination is the primary mechanism by which APOBEC3G restricts HIV-1; however, several studies have reported that APOBEC3G also inhibits virus replication via a mechanism that is independent of deamination. Using active site APOBEC3G mutants, we have re-evaluated the biological relevance of deaminase-independent APOBEC3G-mediated restriction of HIV-1. APOBEC3G proteins with Glu→Ala mutations in AS1, AS2 or AS1 and AS2 were stably expressed at physiological levels in CEM-SS T cells and 293T cells and the ability of the cells to support Δvif HIV-1 replication was then tested. The AS2 and AS1/AS2 mutants were packaged efficiently into virions but in single-cycle or multi-cycle HIV-1 replication assays, were found to lack antiviral activity. The AS1 mutant, which retained deaminase activity, maintained near wild-type antiviral function. To determine the potency of APOBEC3G antiviral activity, cell lines were established that that expressed low levels of wild-type APOBEC3G and generated virions that contained as few as 1-2 APOBEC3G molecules. Even at very low copy number, APOBEC3G caused a significant reduction in infectivity, suggesting that a single molecule of packaged APOBEC3G inactivates the virus. The high potency of APOBEC3G is consistent with a catalytic mechanism of restriction in which a single molecule can induce a string of mutations but difficult to reconcile with a deaminase-independent, non-catalytic mechanism. Analysis of the reverse transcript sequences showed that the G→A mutations were clustered, likely reflecting the action of single APOBEC3G molecules acting processively. We conclude that cytidine deamination is the mechanism by which APOBEC3G restricts HIV-1.
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