Assisted evolution enables HIV-1 to overcome a high TRIM5α-imposed genetic barrier to rhesus macaque tropism.
Assisted evolution enables HIV-1 to overcome a high TRIM5α-imposed genetic barrier to rhesus macaque tropism.
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DOI：
10.1371/journal.ppat.1003667
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发表时间：
2013
期刊：
影响因子：
6.7
通讯作者：
Bieniasz PD
中科院分区：
文献类型：
--
作者：
Soll SJ;Wilson SJ;Kutluay SB;Hatziioannou T;Bieniasz PD
宿主进化过程中抗逆转录病毒因子的多样化为跨物种逆转录病毒传播设置了强大的障碍。这种现象可能保护人类免受许多现代逆转录病毒的感染，但它也损害了HIV-1感染的灵长类动物模型的发展。事实上，恒河猴对艾滋病毒-1具有抵抗力，部分原因是TRIM5α蛋白(rhTRIM5α)的限制。最初，我们试图使用两种策略来获得耐rh TRIM5HIV-1α毒株。首先，HIV-1在表达重组人TRIM5α的工程化人类细胞中传代。其次，对随机突变的衣壳蛋白(CA)序列文库进行筛选，以寻找降低重组人TRIM5α敏感性的突变。这两种方法都发现了CA中的几个个体突变，这些突变降低了重组人TRIM5α的敏感性。然而，这两种方法都没有产生完全抗药性的突变，可能是因为突变的位置表明，trim5α识别衣壳表面的多个决定因素。此外，尽管观察到了各种CA突变对HIV-1对重组人TRIM5α的抗性的相加效应，但完全抗性的组合对适合度非常不利。因此，我们采用了一种“辅助进化”的方法，即在包含人工合成CA序列的病毒克隆库中，随机对降低了重组人TRIM5α敏感性的单个CA突变进行分类。随后病毒文库在表达重组人TRIM5α的细胞中的传代导致了对重组人TRIM5α完全匹配和抵抗的单个病毒种类的选择。这些病毒编码CA的五个突变的组合，通过取消对病毒衣壳的识别，完全或接近完全抵抗重组人TRIM5α对传入病毒核心的破坏作用。重要的是，编码这些CA替换的HIV-1变体和SIVmac239 Vif在原代猕猴淋巴细胞中有效复制。这些发现表明，重组人TRIM5α很难避免，但并非不可能避免，这样做应该有助于发展灵长类动物的艾滋病毒-1感染模型。逆转录病毒，如艾滋病毒-1，感染自然宿主以外的物种的能力往往有限。这种现象在一定程度上是由于抗病毒蛋白的存在，这些蛋白可以保护不受尚未适应特定物种的病毒的感染。例如，恒河猴是医学研究中最常用的猴子物种，它们对艾滋病毒-1感染的抵抗力部分归因于艾滋病毒-1对trim5α的脆弱性。猕猴TRIM5α(rh TRIM5α)通过识别病毒衣壳进入细胞后阻止艾滋病毒感染，已被证明很难获得对rh TRIM5α具有耐药性的HIV-1毒株。然而，通过设计一种“辅助进化”的方法，我们确定了使艾滋病毒-1对重组人TRIM5α产生抗药性的特定突变组合。这些突变使艾滋病毒-1能够通过取消对衣壳的识别来逃避重组人TRIM5α。值得注意的是，将rh TRIM5HIV-1抗α衣壳引入HIV-1中，也是为了避免恒河猴APOBEC3抗病毒蛋白，允许艾滋病毒-1在恒河猴淋巴细胞中有效复制。这些发现有可能推动恒河猴HIV-1感染模型的发展。
Diversification of antiretroviral factors during host evolution has erected formidable barriers to cross-species retrovirus transmission. This phenomenon likely protects humans from infection by many modern retroviruses, but it has also impaired the development of primate models of HIV-1 infection. Indeed, rhesus macaques are resistant to HIV-1, in part due to restriction imposed by the TRIM5α protein (rhTRIM5α). Initially, we attempted to derive rhTRIM5α-resistant HIV-1 strains using two strategies. First, HIV-1 was passaged in engineered human cells expressing rhTRIM5α. Second, a library of randomly mutagenized capsid protein (CA) sequences was screened for mutations that reduced rhTRIM5α sensitivity. Both approaches identified several individual mutations in CA that reduced rhTRIM5α sensitivity. However, neither approach yielded mutants that were fully resistant, perhaps because the locations of the mutations suggested that TRIM5α recognizes multiple determinants on the capsid surface. Moreover, even though additive effects of various CA mutations on HIV-1 resistance to rhTRIM5α were observed, combinations that gave full resistance were highly detrimental to fitness. Therefore, we employed an ‘assisted evolution’ approach in which individual CA mutations that reduced rhTRIM5α sensitivity without fitness penalties were randomly assorted in a library of viral clones containing synthetic CA sequences. Subsequent passage of the viral library in rhTRIM5α-expressing cells resulted in the selection of individual viral species that were fully fit and resistant to rhTRIM5α. These viruses encoded combinations of five mutations in CA that conferred complete or near complete resistance to the disruptive effects of rhTRIM5α on incoming viral cores, by abolishing recognition of the viral capsid. Importantly, HIV-1 variants encoding these CA substitutions and SIVmac239 Vif replicated efficiently in primary rhesus macaque lymphocytes. These findings demonstrate that rhTRIM5α is difficult to but not impossible to evade, and doing so should facilitate the development of primate models of HIV-1 infection. Retroviruses such as HIV-1 often exhibit limited capacity to infect species other than their natural hosts. This phenomenon is partly due to the existence of antiviral proteins that protect against infection by viruses that have not adapted to a particular species. For example, the resistance of rhesus macaques, the monkey species most commonly used in medical research, to HIV-1 infection is partly attributable to the vulnerability of HIV-1 to TRIM5α. Rhesus macaque TRIM5α (rhTRIM5α) blocks HIV-1 infection by recognition of the viral capsid following its entry into the cell, and it has proven difficult to derive HIV-1 strains that are resistant to rhTRIM5α. However, by devising an ‘assisted evolution’ approach, we identified particular combinations of mutations that render HIV-1 resistant to rhTRIM5α. These mutations enable HIV-1 to evade rhTRIM5α by abolishing recognition of the capsid. Notably, introduction of rhTRIM5α-resistant capsids into an HIV-1 that was also engineered to avoid the rhesus macaque APOBEC3 antiviral proteins, allowed efficient HIV-1 replication in rhesus macaque lymphocytes. These discoveries have the potential to advance the development of rhesus macaque models of HIV-1 infection.
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影响因子：
32.4
作者：
Blanco-Melo D;Venkatesh S;Bieniasz PD
通讯作者：
Bieniasz PD
影响因子：
6.7
作者：
Kutluay SB;Perez-Caballero D;Bieniasz PD
通讯作者：
Bieniasz PD
影响因子：
5.4
作者：
Hatziioannou, T;Perez-Caballero, D;Bieniasz, PD
通讯作者：
Bieniasz, PD
DOI：
10.1073/pnas.0812587106
发表时间：
2009-03-17
影响因子：
11.1
作者：
Hatziioannou, Theodora;Ambrose, Zandrea;Bieniasz, Paul D.
通讯作者：
Bieniasz, Paul D.
影响因子：
6.7
作者：
McNatt, Matthew W.;Zang, Trinity;Hatziioannou, Theodora;Bartlett, Mackenzie;Ben Fofana, Ismael;Johnson, Welkin E.;Neil, Stuart J. D.;Bieniasz, Paul D.
通讯作者：
Bieniasz, Paul D.