Background & Aims

Our understanding of the interactions between HBV and its host cells is still quite limited. Spliceosome associated factor 1 (SART1) has recently been found to restrict HCV. Thus, we aimed to dissect its role in HBV infection.

Methods

SART1 was knocked down by RNA interference and over-expressed by lentiviral or adeno-associated virus (AAV) vectors in HBV-infected cell cultures and in vivo in HBV-infected mice. Luciferase reporter assays were used to determine viral or host factor promoter activities, and chromatin immunoprecipitation (ChIP) was used to investigate protein-DNA interactions.

Results

In HBV-infected cell cultures, downregulation of SART1 did not affect covalently closed circular HBV DNA but resulted in markedly enhanced HBV RNA, antigen expression and progeny virus production. On the other hand, HBV transcription and replication were significantly inhibited by overexpression of SART1. Similar results were observed in AAV-HBV-infected mice persistently replicating HBV. Inhibition of Janus kinases had no effect on SART1-mediated inhibition of HBV replication. HBV promoter assays revealed that SART1 reduced HBV core promoter activity. By screening known HBV transcription factors, we found that SART1 specifically suppressed the expression of hepatocyte nuclear factor 4α (HNF4α). Luciferase reporter and ChIP assays demonstrated a direct downregulation of HNF4α expression by association of SART1 with the HNF4α proximal P1 promoter element.

Conclusions

We identify SART1 as a novel host factor suppressing HBV cccDNA transcription. Besides its effect on interferon-stimulated genes, SART1 exerts an anti-HBV activity by suppressing HNF4α expression, which is essential for transcription of HBV cccDNA.

Lay summary

Hepatitis B virus (HBV) infects hepatocytes and persists in the form of covalently closed circular DNA (cccDNA), which remains a major obstacle to successful antiviral treatment. In this study, using various HBV models, we demonstrate that the protein SART1 restricts HBV cccDNA transcription by suppressing a key transcription factor, HNF4α.

中文翻译：

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SART1 在 HNF4α 转录调控中的新功能有助于其在 HBV 感染期间的抗病毒作用

背景与目标

我们对 HBV 与其宿主细胞之间相互作用的理解仍然非常有限。最近发现剪接体相关因子 1 (SART1) 可以限制 HCV。因此，我们旨在剖析其在 HBV 感染中的作用。

方法

SART1 被 RNA 干扰击倒，并在 HBV 感染的细胞培养物中和HBV 感染的小鼠体内被慢病毒或腺相关病毒 (AAV) 载体过度表达。荧光素酶报告基因检测用于确定病毒或宿主因子启动子活性，染色质免疫沉淀 (ChIP) 用于研究蛋白质-DNA 相互作用。

结果

在 HBV 感染的细胞培养物中，SART1 的下调不影响共价闭合环状 HBV DNA，但会导致 HBV RNA、抗原表达和后代病毒产生显着增强。另一方面，SART1的过表达显着抑制了HBV的转录和复制。在持续复制 HBV 的 AAV-HBV 感染小鼠中观察到类似的结果。抑制 Janus 激酶对 SART1 介导的 HBV 复制抑制没有影响。HBV 启动子检测显示 SART1 降低了 HBV 核心启动子的活性。通过筛选已知的HBV转录因子，我们发现SART1特异性抑制肝细胞核因子4α（HNF4α）的表达。

结论

我们将 SART1 鉴定为抑制 HBV cccDNA 转录的新型宿主因子。除了对干扰素刺激基因的影响外，SART1 通过抑制 HNF4α 表达发挥抗 HBV 活性，这对于 HBV cccDNA 的转录至关重要。

总结

乙型肝炎病毒 (HBV) 感染肝细胞并以共价闭合环状 DNA (cccDNA) 的形式持续存在，这仍然是成功抗病毒治疗的主要障碍。在这项研究中，使用各种 HBV 模型，我们证明 SART1 蛋白通过抑制关键转录因子 HNF4α 来限制 HBV cccDNA 转录。