Perturbing HIV-1 Ribosomal Frameshifting Frequency Reveals a cis Preference for Gag-Pol Incorporation into Assembling Virions
Perturbing HIV-1 Ribosomal Frameshifting Frequency Reveals a cis Preference for Gag-Pol Incorporation into Assembling Virions
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DOI：
10.1128/jvi.01349-21
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发表时间：
2022-01-01
影响因子：
5.4
通讯作者：
Sherer，Nathan M.
中科院分区：
文献类型：
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作者：
Benner，Bayleigh E.;Bruce，James W.;Sherer，Nathan M.
HIV-1病毒体产生由Gag和Gag-Pol（GP）蛋白驱动，其中Gag形成衣壳的大部分并驱动出芽，而GP结合Gag以递送必需的病毒体酶蛋白酶、逆转录酶和整合酶。传统上认为病毒颗粒GP水平反映了GP和Gag在细胞中的相对丰度（101：20），由ingag-polmRNAs发生的a-1程序性核糖体移码（PRF）事件的频率决定。在这里，我们利用了一组PRF突变病毒，以表明除了PRF调节GP掺入病毒体的机制。首先，我们发现GP在病毒体中相对于细胞富集了103倍，在亚生理水平的GP比GP水平太高时更好地保持了病毒感染性。第二，我们报告说，当Gag和GP被切割合成时，GP更有效地掺入病毒体中（即，从sameag-polmRNA）比intranss，这表明Gag/GP翻译和组装是空间耦合的过程。第三，我们发现，令人惊讶的是，病毒体表现出一个强大的上限，以trans-delivered GP掺入;适应，似乎允许病毒回火缺陷GP/Gag切割，可能会产生负面影响逆转录。将这些结果结合在一起，我们提出了一个“加权金发姑娘”方案的HIV-1 GP掺入，其中GP富集和排斥缓冲液病毒粒子感染性在广泛的局部GP浓度的组合机制。这些结果为HIV-1病毒粒子装配途径提供了新的见解，该途径与PRF靶向抗病毒策略的预期功效相关。重要信息HIV-1感染性需要在病毒粒子装配过程中将Gag-Pol（GP）前体多聚蛋白掺入病毒粒子中。决定GP整合到组装病毒体中的机制定义不清，传统上认为病毒体中的GP水平仅反映细胞中表达的Gag和GP的相对水平，由发生在gag-polmRNAs中的-1程序性核糖体移码（PRF）事件的频率决定。在此，我们提供了实验支持的“加权金发姑娘”的情况下，GP掺入，其中病毒利用随机和非随机机制，以缓冲感染性在广泛的GP表达水平。这些机制数据与正在进行的开发针对PRF频率和/或HIV-1病毒粒子成熟的抗病毒策略的努力有关。
HIV-1 virion production is driven by Gag and Gag-Pol (GP) proteins, with Gag forming the bulk of the capsid and driving budding, while GP binds Gag to deliver the essential virion enzymes protease, reverse transcriptase, and integrase. Virion GP levels are traditionally thought to reflect the relative abundances of GP and Gag in cells (∼1:20), dictated by the frequency of a −1 programmed ribosomal frameshifting (PRF) event occurring ingag-polmRNAs. Here, we exploited a panel of PRF mutant viruses to show that mechanisms in addition to PRF regulate GP incorporation into virions. First, we show that GP is enriched ∼3-fold in virions relative to cells, with viral infectivity being better maintained at subphysiological levels of GP than when GP levels are too high. Second, we report that GP is more efficiently incorporated into virions when Gag and GP are synthesized incis(i.e., from the samegag-polmRNA) than intrans, suggesting that Gag/GP translation and assembly are spatially coupled processes. Third, we show that, surprisingly, virions exhibit a strong upper limit totrans-delivered GP incorporation; an adaptation that appears to allow the virus to temper defects to GP/Gag cleavage that may negatively impact reverse transcription. Taking these results together, we propose a “weighted Goldilocks” scenario for HIV-1 GP incorporation, wherein combined mechanisms of GP enrichment and exclusion buffer virion infectivity over a broad range of local GP concentrations. These results provide new insights into the HIV-1 virion assembly pathway relevant to the anticipated efficacy of PRF-targeted antiviral strategies.IMPORTANCEHIV-1 infectivity requires incorporation of the Gag-Pol (GP) precursor polyprotein into virions during the process of virus particle assembly. Mechanisms dictating GP incorporation into assembling virions are poorly defined, with GP levels in virions traditionally thought to solely reflect relative levels of Gag and GP expressed in cells, dictated by the frequency of a −1 programmed ribosomal frameshifting (PRF) event that occurs ingag-polmRNAs. Herein, we provide experimental support for a “weighted Goldilocks” scenario for GP incorporation, wherein the virus exploits both random and nonrandom mechanisms to buffer infectivity over a wide range of GP expression levels. These mechanistic data are relevant to ongoing efforts to develop antiviral strategies targeting PRF frequency and/or HIV-1 virion maturation.