由衣壳接头序列和抑制剂调节的核衣壳中区域特异性乙型肝

StephenW

由衣壳接头序列和抑制剂调节的核衣壳中区域特异性乙型肝炎病毒基因组暴露：脱壳的意义
作者：姬曦、崔秀姬、刘宽成、刘海涛、王若瑟、胡建明 https://orcid.org/0000-0002-3967-2133 [email protected] Info & Affiliations
DOI：
https://doi.org/10.1128/jvi.00399-22
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合资公司
第 96 卷，第 8 期
2022 年 4 月 27 日

    抽象的
    参考

抽象的
乙型肝炎病毒 (HBV) 包含在宿主细胞质中的核衣壳 (NC) 内合成的部分双链、松弛的环状 (RC) DNA 基因组。 RC DNA 从 NC 释放到细胞核中是一个定义不明确的过程，称为脱壳，这是其转化为共价闭合环状 (CCC) DNA 所必需的，该病毒附加体是所有病毒 RNA 的转录模板。复制，因此对于建立和维持病毒感染至关重要。为了更好地了解脱壳，我们分析了 HBV 核心 (HBc) 突变体，这些突变体显示出不同水平的核 CCC DNA，但几乎没有细胞质 RC DNA。我们发现这些突变体可以在细胞外合成 RC DNA，但与野生型 (wt) 相比，突变体 NCs 无法保护 RC DNA 免受细胞裂解物中的内源性核酸酶或外源性 DNase 的消化.亚细胞分离表明主要的 RC DNA 降解活性与膜相关。用序列特异性和非特异性 DNA 酶消化揭示了突变 NC 中 RC DNA 特定区域的暴露。类似地，用已知通过影响脱壳增加 CCC DNA 的核心抑制剂处理 wt NCs 也导致 RC DNA 的区域特异性暴露。此外，未经处理的野生型 (wt) 成熟 NCs 的亚群也显示出 RC DNA 的位点特异性暴露。因此，在 NC 脱壳过程中 RC DNA 降解及其转化为 CCC DNA 之间的竞争可能在 HBV 感染的建立和持续存在中起重要作用，并且对衣壳靶向抗病毒药物的开发具有重要意义。
重要性 乙型肝炎病毒 (HBV) 核衣壳 (NC) 的解体以释放其基因组 DNA，这是一个被称为脱壳的不为人知的过程，是在宿主细胞核中形成病毒核附加体所必需的，这是一种病毒 DNA，对于建立和释放其基因组 DNA 是必不可少的。持续感染乙肝病毒。消除 HBV 核附加体仍然是开发 HBV 治愈方法的圣杯。我们在此报告，HBV 基因组 DNA 在脱壳过程中以区域特异性方式暴露，这通过衣壳蛋白和衣壳靶向抗病毒化合物的突变而增强。病毒基因组的暴露可以导致其快速降解，或者可以增强核附加体的形成，从而对 HBV 感染和持久性产生重大影响。因此，这些结果对于了解 HBV 复制和持续存在的基本机制以及对 HBV 治愈的持续追求非常重要。

StephenW

Region-Specific Hepatitis B Virus Genome Exposure from Nucleocapsid Modulated by Capsid Linker Sequence and Inhibitor: Implications for Uncoating
Authors: Ji Xi, Xiuji Cui, Kuancheng Liu, Haitao Liu, Joseph Wang, Jianming Hu https://orcid.org/0000-0002-3967-2133 [email protected] Info & Affiliations
DOI:
https://doi.org/10.1128/jvi.00399-22
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JVI
Volume 96, Number 8
27 April 2022

    ABSTRACT
    REFERENCES

ABSTRACT
Hepatitis B virus (HBV) contains a partially double-stranded, relaxed circular (RC) DNA genome synthesized within a nucleocapsid (NC) in the host cell cytoplasm. The release of RC DNA from the NC, in an ill-defined process called uncoating, to the nucleus is required for its conversion to the covalently closed circular (CCC) DNA, the viral episome serving as the transcriptional template for all viral RNAs necessary for replication and, thus, essential for establishing and sustaining viral infection. In efforts to better understand uncoating, we analyzed HBV core (HBc) mutants that show various levels of nuclear CCC DNA but little to no cytoplasmic RC DNA. We found that RC DNA could be synthesized by these mutants outside the cell, but in contrast to the wild type (wt), the mutant NCs were unable to protect RC DNA from digestion by the endogenous nuclease(s) in cellular lysates or exogenous DNase. Subcellular fractionation suggested that the major RC DNA-degrading activity was membrane associated. Digestion with sequence-specific and nonspecific DNases revealed the exposure of specific regions of RC DNA from the mutant NC. Similarly, treatment of wt NCs with a core inhibitor known to increase CCC DNA by affecting uncoating also led to region-specific exposure of RC DNA. Furthermore, a subpopulation of untreated wild type (wt) mature NCs showed site-specific exposure of RC DNA as well. Competition between RC DNA degradation and its conversion to CCC DNA during NC uncoating thus likely plays an important role in the establishment and persistence of HBV infection and has implications for the development of capsid-targeted antivirals.
IMPORTANCE Disassembly of the hepatitis B virus (HBV) nucleocapsid (NC) to release its genomic DNA, in an ill-understood process called uncoating, is required to form the viral nuclear episome in the host cell nucleus, a viral DNA essential for establishing and sustaining HBV infection. The elimination of the HBV nuclear episome remains the holy grail for the development of an HBV cure. We report here that the HBV genomic DNA is exposed in a region-specific manner during uncoating, which is enhanced by mutations of the capsid protein and a capsid-targeted antiviral compound. The exposure of the viral genome can result in its rapid degradation or, alternatively, can enhance the formation of the nuclear episome, thus having a major impact on HBV infection and persistence. These results are thus important for understanding fundamental mechanisms of HBV replication and persistence and for the ongoing pursuit of an HBV cure.