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Multi-Omics Panoramic  Analysis of HBV Integration, Transcriptional Regulation, and Epigenetic  Modifications in PLC/PRF/5 Cell Line        
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  • Guiwen Guan,
  • Abudurexiti Abulaiti,
  • Chia-Chen Chen,
  • Zhiqiang Gu,
  • Jing Yang,
  • Ting Zhang,
  • Xiaojie Chen,
  • Zhao Zhou,
  • Fengmin Lu,
  • Xiangmei Chen
Guiwen Guan
Peking University School of Basic Medical Sciences

Corresponding Author:[email protected]

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Abudurexiti Abulaiti
Peking University School of Basic Medical Sciences
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Chia-Chen Chen
Peking University School of Basic Medical Sciences
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Zhiqiang Gu
Peking University School of Basic Medical Sciences
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Jing Yang
Guangdong Provincial People's Hospital
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Ting Zhang
Peking University School of Basic Medical Sciences
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Xiaojie Chen
Capital Medical University Affiliated Beijing Friendship Hospital
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Zhao Zhou
Peking University School of Basic Medical Sciences
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Fengmin Lu
Peking University School of Basic Medical Sciences

Corresponding Author:[email protected]

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Xiangmei Chen
Peking University School of Basic Medical Sciences

Corresponding Author:[email protected]

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Abstract

Background: The clearance or transcriptional silencing of integrated HBV DNA is crucial for achieving a functional cure in patients with chronic hepatitis B (CHB) and reducing the risk of hepatocellular carcinoma (HCC) development. The PLC/PRF/5 cell line is commonly used as an in vitro model for studying HBV integration. In this study, we employed a range of multi-omics techniques to gain a panoramic understanding of the characteristics of HBV integration in PLC/PRF/5 cells and to reveal the transcriptional regulatory mechanisms of integrated HBV DNA.
Methods: Transcriptome long-read sequencing (ONT) was conducted to analyze and characterize the transcriptional activity of different HBV DNA integration sites in PLC/PRF/5 cells. Additionally, we collected data related to epigenetic regulation, including whole-genome bisulfite sequencing (WGBS), histone chromatin immunoprecipitation sequencing (ChIP-seq), and assays for transposase-accessible chromatin using sequencing (ATAC-seq), to explore the potential mechanisms involved in the transcriptional regulation of integrated HBV DNA.
Result: Long-read RNA sequencing analysis revealed significant transcriptional differences at various integration sites in the PLC/PRF/5 cell line, with higher HBV DNA transcription levels at integration sites on chr11, chr13, and the chr13/chr5 fusion chromosome t (13:5). Combining long-read DNA and RNA sequencing results, we found that transcription of integrated HBV DNA generally starts downstream of the SP1, SP2, or XP promoters. ATAC-seq data confirmed that chromatin accessibility has limited influence on the transcription of integrated HBV DNA in the PLC/PRF/5 cell line. Analysis of whole-genome bisulfite sequencing data showed that the methylation intensity of integrated HBV DNA was highly negatively correlated with its transcription level (r = -0.8929, p = 0.0123). After AzaD treatment, the transcription level of integrated HBV DNA significantly increased, especially for the integration chr17, which had the highest level of methylation. Through ChIP-seq data, we observed the association between histone modification of H3K4me3 and H3K9me3 with the transcription of integrated HBV DNA.
08 Oct 2023Submitted to Journal of Medical Virology
09 Oct 2023Submission Checks Completed
09 Oct 2023Assigned to Editor
09 Oct 2023Review(s) Completed, Editorial Evaluation Pending
22 Oct 2023Editorial Decision: Revise Major
29 Feb 20241st Revision Received
11 Mar 2024Submission Checks Completed
11 Mar 2024Assigned to Editor
11 Mar 2024Reviewer(s) Assigned
18 Mar 2024Review(s) Completed, Editorial Evaluation Pending
20 Mar 2024Editorial Decision: Revise Minor
05 Apr 2024Review(s) Completed, Editorial Evaluation Pending
08 Apr 2024Editorial Decision: Accept