Three-dimensional and single-cell sequencing of liver cancer reveals comprehensive host-virus interactions in HBV infection

Abstract

BackgroundsHepatitis B virus (HBV) infection is a major risk factor for chronic liver diseases and liver cancer (mainly hepatocellular carcinoma, HCC), while the underlying mechanisms and host-virus interactions are still largely elusive.MethodsWe applied HiC sequencing to HepG2 (HBV-) and HepG2-2.2.15 (HBV+) cell lines and combined them with public HCC single-cell RNA-seq data, HCC bulk RNA-seq data, and both genomic and epigenomic ChIP-seq data to reveal potential disease mechanisms of HBV infection and host-virus interactions reflected by 3D genome organization.ResultsWe found that HBV enhanced overall proximal chromatin interactions (CIs) of liver cells and primarily affected regional CIs on chromosomes 13, 14, 17, and 22. Interestingly, HBV altered the boundaries of many topologically associating domains (TADs), and genes nearby these boundaries showed functional enrichment in cell adhesion which may promote cancer metastasis. Moreover, A/B compartment analysis revealed dramatic changes on chromosomes 9, 13 and 21, with more B compartments (inactive or closed) shifting to A compartments (active or open). The A-to-B regions (closing) harbored enhancers enriched in the regulation of inflammatory responses, whereas B-to-A regions (opening) were enriched for transposable elements (TE). Furthermore, we identified large HBV-induced structural variations (SVs) that disrupted tumor suppressors, NLGN4Y and PROS1. Finally, we examined differentially expressed genes and TEs in single hepatocytes with or without HBV infection, by using single-cell RNA-seq data. Consistent with our HiC sequencing findings, two upregulated genes that promote HBV replication, HNF4A and NR5A2, were located in regions with HBV-enhanced CIs, and five TEs were located in HBV-activated regions. Therefore, HBV may promote liver diseases by affecting the human 3D genome structure.ConclusionOur work promotes mechanistic understanding of HBV infection and host-virus interactions related to liver diseases that affect billions of people worldwide. Our findings may also have implications for novel immunotherapeutic strategies targeting HBV infection.

Multiomics analyses reveal pathological mechanisms of HBV infection and integration in liver cancer

“…Interestingly, we found most of these VIMs (12/14) resided in the boundaries of chromosome TADs in HepG2 liver cancer cells (TAD information downloaded from Wang et al 18 ) (Supporting Information: Figure ). We further found that HBV integration events also tended to happen within TAD boundaries ( p value = 3.9e‐9, odds ratio = 1.45, see Section 2), which was consistent with our recent report 28 …”

“…We further found that HBV integration events also tended to happen within TAD boundaries (p value = 3.9e-9, odds ratio = 1.45, see Section 2), which was consistent with our recent report. 28 Mutations in TAD boundaries were known to cause gene regulatory network rewiring and abnormal expression of oncogenes. 29,30 We further investigated whether any TF binding was enriched in these risk regions by integrating binding peaks from >1100 TFs.…”

Three‐dimensional genome structure and function

“…During this process, there are dynamic changes in the A/B compartments, mainly in the genomic regions on chromosomes 9, 13, and 21. Most genes are moved from transcriptionally repressed B compartments to A compartments, where they are subsequently activated 256 . These regions have many transposable elements that promote viral DNA replication.…”

Human tumor viruses: induction of three-dimensional alterations in the host genome structure

“…Using Hi-C sequencing, Guo et al analysed chromatin interactions in a HBV-infected human hepatoma cell line and its uninfected counterpart ( Guo et al, 2023 ). They observed enhanced chromatin interactions, especially on distinct interphase chromosomes, in HBV-positive cells and recorded characteristic shifts in nuclear subcompartments, accompanied with repression of enhancers regulating inflammatory genes and opening of regions enriched in transposable elements ( Guo et al, 2023 ). These data suggested that reorganization of 3D genome structure may contribute to disease development associated with HBV infection.…”