Department of Gastroenterology, Zhengzhou University People's Hospital, Henan Provincial People's Hospital, Zhengzhou, 450003, Henan, China.
The department of infectious diseases, The Second Affiliated Hospital of Zhengzhou University, Zhengzhou, 450000, Henan, China.
Arch Virol. 2024 Jan 24;169(2):36. doi: 10.1007/s00705-023-05949-6.
Current therapies for hepatitis B virus (HBV) infection can slow disease progression but cannot cure the infection, as it is difficult to eliminate or permanently silence HBV covalently closed circular DNA (cccDNA). The interaction between host factors and cccDNA is essential for their formation, stability, and transcriptional activity. Here, we focused on the regulatory role of the host factor ENPP1 and its interacting transcription factor LMNB1 in HBV replication and transcription to better understand the network of host factors that regulate HBV, which may facilitate the development of new antiviral drugs. Overexpression of ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) in Huh7 cells decreased HBV pregenomic RNA (pgRNA) and hepatitis B core antigen (HBcAg) expression levels, whereas knockdown of ENPP1 increased them. A series of HBV promoter and mutant plasmids were constructed, and a luciferase reporter assay showed that overexpression of ENPP1 caused inhibition of the HBV promoter and its mutants. A DNA pull-down assay showed that lamin B1 (LMNB1), but not ENPP1, interacts directly with the HBV enhancer II/ basic core promoter (EnhII/BCP). ZDOCK and PyMOL software were used to predict the interaction of ENPP1 with LMNB1. Overexpression of LMNB1 inhibited the activity of the HBV promoter and its mutant. The acetylation levels at the amino acids 111K, 261K, and 483K of LMNB1 were reduced compared to the control, and an LMNB1 acetylation mutant containing 111R, 261Q, 261R, 483Q, and 483R showed increased promoter activity. In summary, ENPP1 together with LMNB1 increased the acetylation level at 111K and 261K, and LMNB1 inhibited the activity of HBV promoter and downregulated the expression of pregenomic RNA and HBcAg. Our follow-up studies will investigate the expression, clinical significance, and relevance of ENPP1 and LMNB1 in HBV patient tissues, explore the effect of LMNB1 on post-transcriptional progression, and examine whether ENPP1 can reduce cccDNA levels in the nucleus.
当前治疗乙型肝炎病毒（HBV）感染的方法可以减缓疾病进展，但不能治愈感染，因为很难消除或永久沉默 HBV 共价闭合环状 DNA（cccDNA）。宿主因素与 cccDNA 的相互作用对于它们的形成、稳定性和转录活性至关重要。在这里，我们专注于宿主因素 ENPP1 及其相互作用的转录因子 LMNB1 在 HBV 复制和转录中的调节作用，以更好地了解调节 HBV 的宿主因素网络，这可能有助于开发新的抗病毒药物。在 Huh7 细胞中过表达核苷酸磷酸二酯酶 1（ENPP1）可降低 HBV 前基因组 RNA（pgRNA）和乙型肝炎核心抗原（HBcAg）的表达水平，而敲低 ENPP1 则增加了它们的表达。构建了一系列 HBV 启动子和突变质粒，并进行了荧光素酶报告基因检测，结果显示过表达 ENPP1 抑制了 HBV 启动子及其突变体的活性。DNA 下拉实验显示，核纤层蛋白 B1（LMNB1），而不是 ENPP1，与 HBV 增强子 II/基本核心启动子（EnhII/BCP）直接相互作用。使用 ZDOCK 和 PyMOL 软件预测了 ENPP1 与 LMNB1 的相互作用。过表达 LMNB1 抑制了 HBV 启动子及其突变体的活性。与对照相比，LMNB1 的氨基酸 111K、261K 和 483K 的乙酰化水平降低，并且包含 111R、261Q、261R、483Q 和 483R 的 LMNB1 乙酰化突变体显示出增强的启动子活性。总之，ENPP1 与 LMNB1 一起增加了 111K 和 261K 的乙酰化水平，而 LMNB1 抑制了 HBV 启动子的活性并下调了前基因组 RNA 和 HBcAg 的表达。我们的后续研究将调查 ENPP1 和 LMNB1 在 HBV 患者组织中的表达、临床意义和相关性，探索 LMNB1 对转录后进展的影响，并检查 ENPP1 是否可以降低核内 cccDNA 水平。
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