nackednaviruses病毒中HBV RNA元素ε的保守性揭示了蛋白质引发的

StephenW

Conservation of the HBV RNA element epsilon in nackednaviruses reveals ancient origin of protein-primed reverse transcription
View ORCID ProfileJürgen Beck, View ORCID ProfileStefan Seitz, View ORCID ProfileChris Lauber, and View ORCID ProfileMichael Nassal

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PNAS March 30, 2021 118 (13) e2022373118; https://doi.org/10.1073/pnas.2022373118

    Edited by Stephen P. Goff, Columbia University Medical Center, New York, NY, and approved February 7, 2021 (received for review October 26, 2020)

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Significance

Hepadnaviruses, including hepatitis B virus (HBV) as a major human pathogen, are small, enveloped DNA viruses, which replicate by reverse transcription of an RNA intermediate. Unlike retroviruses, which use tRNA as primer for reverse transcription, hepadnaviruses employ a unique protein-priming mechanism involving de novo synthesis of a DNA primer covalently attached to the viral polymerase. Here, we show that this mechanism is highly conserved on the molecular level in distantly related non-enveloped fish viruses which diverged from ancestral hepadnaviruses more than 400 Mya. The exceptional level of conservation and the absence of known homologous cellular mechanisms renders HBV protein priming a promising, yet unexplored, antiviral target for the development of novel therapeutics against this highly relevant pathogen.
Abstract

Hepadnaviruses, with the human hepatitis B virus as prototype, are small, enveloped hepatotropic DNA viruses which replicate by reverse transcription of an RNA intermediate. Replication is initiated by a unique protein-priming mechanism whereby a hydroxy amino acid side chain of the terminal protein (TP) domain of the viral polymerase (P) is extended into a short DNA oligonucleotide, which subsequently serves as primer for first-strand synthesis. A key component in the priming of reverse transcription is the viral RNA element epsilon, which contains the replication origin and serves as a template for DNA primer synthesis. Here, we show that recently discovered non-enveloped fish viruses, termed nackednaviruses [C. Lauber et al., Cell Host Microbe 22, 387–399 (2017)], employ a fundamentally similar replication mechanism despite their huge phylogenetic distance and major differences in genome organization and viral lifestyle. In vitro cross-priming studies revealed that few strategic nucleotide substitutions in epsilon enable site-specific protein priming by heterologous P proteins, demonstrating that epsilon is functionally conserved since the two virus families diverged more than 400 Mya. In addition, other cis elements crucial for the hepadnavirus-typical replication of pregenomic RNA into relaxed circular double-stranded DNA were identified at conserved positions in the nackednavirus genomes. Hence, the replication mode of both hepadnaviruses and nackednaviruses was already established in their Paleozoic common ancestor, making it a truly ancient and evolutionary robust principle of genome replication that is more widespread than previously thought.

    protein priming initiation of reverse transcription HBV replication mechanism HBV long-term evolution-paleo virology

StephenW

nackednaviruses病毒中HBV RNA元素ε的保守性揭示了蛋白质引发的逆转录的古老起源
查看ORCID配置文件JürgenBeck，查看ORCID配置文件Stefan Seitz，查看ORCID配置文件Christ Lauber和查看ORCID配置文件Michael Nassal

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PNAS 2021年3月30日118（13）e2022373118; https://doi.org/10.1073/pnas.2022373118

    由纽约州哥伦比亚大学医学中心的斯蒂芬·高夫（Stephen P.Goff）编辑，并于2021年2月7日获得批准（2020年10月26日接受审查）

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意义

肝炎病毒，包括作为主要人类病原体的乙型肝炎病毒（HBV），是包膜的小型DNA病毒，可通过RNA中间体的逆转录进行复制。与逆转录病毒不同，逆转录病毒使用tRNA作为逆转录引物，而肝炎病毒则采用独特的蛋白质引发机制，该机制涉及从头合成与病毒聚合酶共价连接的DNA引物。在这里，我们显示了这种机制在远缘的非包膜鱼病毒中从分子水平上是高度保守的，该病毒与祖先嗜肝DNA病毒有400多个Mya分支。优异的保守水平和缺乏已知的同源细胞机制使HBV蛋白成为开发针对这种高度相关病原体的新型疗法的有希望但尚未探索的抗病毒靶标。
抽象的

以人类乙型肝炎病毒为原型的肝炎病毒是小型的，包膜的肝细胞性DNA病毒，可通过RNA中间体的逆转录进行复制。复制是通过独特的蛋白质引发机制启动的，由此病毒聚合酶（P）末端蛋白质（TP）域的羟基氨基酸侧链被延伸到短的DNA寡核苷酸中，该寡核苷酸随后用作第一链合成的引物。引发逆转录的关键成分是病毒RNA元素epsilon，它包含复制起点，并用作DNA引物合成的模板。在这里，我们显示了最近发现的非包膜鱼病毒，称为nackednaviruses [C. Lauber等人，Cell Host Microbe 22，387–399（2017）]，尽管具有巨大的系统发育距离以及基因组组织和病毒生活方式的重大差异，但仍采用了基本相似的复制机制。体外交叉引物研究表明，ε中很少有战略性的核苷酸取代能够通过异源P蛋白引发位点特异性蛋白引物，这表明自两个病毒家族分化超过400个Mya以来，ε在功能上是保守的。此外，在棘轮状病毒基因组中的保守位置鉴定出了其他对肝炎病毒典型的前基因组RNA复制到松弛的环状双链DNA中至关重要的顺式元件。因此，已经在古生代共同祖先中确立了嗜肝DNA病毒和纳克病毒的复制模式，这使其成为真正的古老且进化稳健的基因组复制原理，比以前认为的更为广泛。

    蛋白质启动引发反转录HBV复制机制HBV长期进化-古病毒学