CpG dinucleotide enrichment in the influenza A virus genome as a live attenuated vaccine development strategy.
CpG dinucleotide enrichment in the influenza A virus genome as a live attenuated vaccine development strategy.
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CpG流感病毒基因组中的CPG二核苷酸富集是一种活疫苗开发策略。
DOI：
10.1371/journal.ppat.1011357
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发表时间：
2023-05
期刊：
影响因子：
6.7
通讯作者：
中科院分区：
文献类型：
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作者：
RNA病毒基因组的同义重编码是产生减毒病毒用作疫苗的有前途的方法。有问题的是，重新编码通常会阻碍病毒生长，但这可以使用CpG二核苷酸富集来纠正。CpG被细胞锌指抗病毒蛋白（ZAP）识别，因此原则上，从病毒繁殖系统中去除ZAP传感将逆转富含CpG的病毒的减毒，从而实现疫苗病毒的高滴度产量。我们使用基因组片段1中CpG含量增加的A型流感病毒（IAV）疫苗株进行了测试。病毒减毒由ZAP的短同种型介导，与添加的CpG的数量相关，并且通过病毒转录物的周转来制定。富含CpG的病毒在小鼠中被强烈减毒，但传达了对野生型病毒的潜在致死攻击剂量的保护。对于疫苗开发重要的是，CpG富集的病毒在连续传代期间是遗传稳定的。出乎意料的是，在用于繁殖减毒活流感疫苗的MDCK细胞和含胚鸡蛋中，ZAP敏感性病毒具有完全复制能力。因此，在人类系统中有缺陷的ZAP敏感性CpG富集病毒可在疫苗繁殖系统中产生高滴度，从而提供现实的、经济上可行的平台来增强现有的活减毒疫苗。CpG二核苷酸在脊椎动物基因组中代表不足，其中CpG构象中的胞嘧啶被甲基化以调节转录。甲基化胞嘧啶易于脱氨基，导致TpG二核苷酸取代CpG。由此产生的CpG抑制提供了一种途径，通过该途径，脊椎动物细胞可以使用细胞锌指抗病毒蛋白（ZAP）作为CpG传感器识别来自入侵病原体的RNA。感染脊椎动物的RNA病毒也在基因组上抑制CpG，据信这是一种进化的性状，以逃避ZAP的检测。在这里，我们设计了一种CpG含量升高的甲型流感病毒（IAV），并描述了这如何影响病毒复制。添加CpG导致病毒减毒，由ZAP活性介导。CpG抑制在狗和鸡基因组中是保守的（与活的减毒IAV疫苗繁殖相关），并且合乎逻辑地预测ZAP介导的CpG传感在这些物种中也是保守的。然而，当我们在同源培养系统中繁殖ZAP敏感的IAV时，我们没有看到复制缺陷。这一意想不到的结果提出了关于为什么感染这些物种的病毒在其基因组中抑制CpG的问题，并且重要的是提供了一种新的、易于处理的方法来增强合理的活减毒IAV疫苗设计。
Synonymous recoding of RNA virus genomes is a promising approach for generating attenuated viruses to use as vaccines. Problematically, recoding typically hinders virus growth, but this may be rectified using CpG dinucleotide enrichment. CpGs are recognised by cellular zinc-finger antiviral protein (ZAP), and so in principle, removing ZAP sensing from a virus propagation system will reverse attenuation of a CpG-enriched virus, enabling high titre yield of a vaccine virus. We tested this using a vaccine strain of influenza A virus (IAV) engineered for increased CpG content in genome segment 1. Virus attenuation was mediated by the short isoform of ZAP, correlated with the number of CpGs added, and was enacted via turnover of viral transcripts. The CpG-enriched virus was strongly attenuated in mice, yet conveyed protection from a potentially lethal challenge dose of wildtype virus. Importantly for vaccine development, CpG-enriched viruses were genetically stable during serial passage. Unexpectedly, in both MDCK cells and embryonated hens’ eggs that are used to propagate live attenuated influenza vaccines, the ZAP-sensitive virus was fully replication competent. Thus, ZAP-sensitive CpG enriched viruses that are defective in human systems can yield high titre in vaccine propagation systems, providing a realistic, economically viable platform to augment existing live attenuated vaccines. CpG dinucleotides are under-represented in vertebrate genomes, wherein cytosines in the CpG conformation are methylated to regulate transcription. Methylated cytosines are prone to deamination, resulting in TpG dinucleotides replacing CpGs. The resultant CpG suppression has provided a route by which vertebrate cells can recognise RNA from invading pathogens, using cellular Zinc-finger Antiviral Protein (ZAP) as a CpG sensor. Vertebrate-infecting RNA viruses also genomically suppress CpGs, and it is believed that this is an evolved trait to evade detection by ZAP. Here, we engineered an influenza A virus (IAV) with elevated CpG content and characterised how this impacts viral replication. CpG addition resulted in viral attenuation, mediated by ZAP activity. CpG suppression is conserved in dog and chicken genomes (relevant for live attenuated IAV vaccine propagation), and it is logical to predict that ZAP-mediated CpG sensing would also be conserved in these species. However, when we propagated ZAP-sensitive IAV in cognate culture systems, we saw no replication defect. This unexpected result raises questions about why viruses infecting these species suppress CpG in their genomes, and importantly delivers a new, tractable approach to augment rational live attenuated IAV vaccine design.
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DOI：
10.1002/wrna.1679
发表时间：
2022-03
期刊：
Wiley interdisciplinary reviews. RNA
影响因子：
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作者：
Gaunt ER;Digard P
通讯作者：
Digard P
影响因子：
6.7
作者：
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通讯作者：
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影响因子：
16.8
作者：
Chen, Shoudeng;Xu, Yihui;Liu, Yingfang
通讯作者：
Liu, Yingfang
影响因子：
2.9
作者：
Goossens, Kate E.;Karpala, Adam J.;Bean, Andrew G. D.
通讯作者：
Bean, Andrew G. D.
影响因子：
56.9
作者：
Goll, MG;Kirpekar, F;Bestor, TH
通讯作者：
Bestor, TH