BACKGROUND & AIMS Hepatitis B virus (HBV) has a DNA genome but replicates within the nucleus by reverse transcription of an RNA pregenome, which is converted to DNA in cytoplasmic capsids. Capsids in this compartment are correlated with inflammation and epitopes of the capsid protein core (Cp) are a major target for T cell-mediated immune responses. We investigated the mechanism of cytoplasmic capsid transport, which is important for infection but also for cytosolic capsid removal. METHODS We used virion-derived capsids containing mature rcDNA (matC) and empty capsids (empC). RNA-containing capsids (rnaC) were used as a control. The investigations comprised pull-down assays for identification of cellular interaction partners, immune fluorescence microscopy for their colocalization and electron microscopy after microinjection to determine their biological significance. RESULTS matC and empC underwent active transport through the cytoplasm towards the nucleus, while rnaC was poorly transported. We identified the dynein light chain LL1 as a functional interaction partner linking capsids to the dynein motor complex and showed that there is no compensatory transport pathway. Using capsid and dynein LL1 mutants we characterized the required domains on the capsid and LL1. CONCLUSIONS This is the first investigation on the detailed molecular mechanism of how matC pass the cytoplasm upon infection and how empC can be actively removed from the cytoplasm into the nucleus. Considering that hepatocytes with cytoplasmic capsids are better recognized by the T cells, we hypothesize that targeting capsid DynLL1-interaction will not only block HBV infection but also stimulate elimination of infected cells. LAY SUMMARY In this study, we identified the molecular details of HBV translocation through the cytoplasm. Our evidence offers a new drug target which could not only inhibit infection but also stimulate immune clearance of HBV infected cells.

中文翻译：

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分子伴侣动力蛋白LL1介导空的和成熟的乙型肝炎病毒衣壳的细胞质运输

背景与目的 乙型肝炎病毒 (HBV) 具有 DNA 基因组，但通过 RNA 前基因组的逆转录在细胞核内复制，该前基因组在细胞质衣壳中转化为 DNA。该隔室中的衣壳与炎症相关，衣壳蛋白核心 (Cp) 的表位是 T 细胞介导的免疫反应的主要目标。我们研究了细胞质衣壳运输的机制，这对于感染和胞质衣壳去除都很重要。方法我们使用含有成熟 rcDNA (matC) 和空衣壳 (empC) 的病毒粒子衍生衣壳。含有 RNA 的衣壳 (rnaC) 用作对照。调查包括用于鉴定细胞相互作用伙伴的下拉分析，免疫荧光显微镜检查它们的共定位和显微注射后的电子显微镜检查以确定它们的生物学意义。结果 matC 和 empC 通过细胞质向细胞核进行主动转运，而 rnaC 转运较差。我们将动力蛋白轻链 LL1 鉴定为将衣壳连接到动力蛋白运动复合体的功能相互作用伙伴，并表明没有补偿性运输途径。使用衣壳和动力蛋白 LL1 突变体，我们表征了衣壳和 LL1 上所需的结构域。结论 这是对 matC 如何在感染时通过细胞质以及 empC 如何从细胞质中主动移入细胞核的详细分子机制的首次研究。考虑到有胞质衣壳的肝细胞更容易被 T 细胞识别，我们假设靶向衣壳 DynLL1 相互作用不仅会阻止 HBV 感染，还会刺激感染细胞的清除。概述 在这项研究中，我们确定了 HBV 通过细胞质易位的分子细节。我们的证据提供了一种新的药物靶点，它不仅可以抑制感染，还可以刺激 HBV 感染细胞的免疫清除。