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A pre-neoplastic epigenetic field defect in HCV-infected liver at transcription factor binding sites and polycomb targets

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ABSTRACT

The predisposition of patients with Hepatitis C virus (HCV) infection to hepatocellular carcinoma (HCC) involves components of viral infection, inflammation and time. The development of multifocal, genetically distinct tumors is suggestive of a field defect affecting the entire liver. The molecular susceptibility mediating such a field defect is not understood. One potential mediator of long-term cellular reprogramming is heritable (epigenetic) regulation of transcription, exemplified by DNA methylation. We studied epigenetic and transcriptional changes in HCV-infected livers in comparison with control, uninfected livers and HCC, allowing us to identify pre-neoplastic epigenetic and transcriptional events.

We find the HCV-infected liver to have a pattern of acquisition of DNA methylation targeted to candidate enhancers active in liver cells, enriched for the binding sites of the FOXA1, FOXA2 and HNF4A transcription factors. These enhancers can be subdivided into those proximal to genes implicated in liver cancer or to genes involved in stem cell development, the latter distinguished by increased CG dinucleotide density and polycomb-mediated repression, manifested by the additional acquisition of histone H3 lysine 27 trimethylation (H3K27me3). Transcriptional studies on our samples showed that the increased DNA methylation at enhancers was associated with decreased local gene expression, results validated in independent samples from The Cancer Genome Atlas (TCGA). Pharmacological depletion of H3K27me3 using the EZH2 inhibitor GSK343 in HepG2 cells suppressed cell growth and also revealed that local acquired DNA methylation was not dependent upon the presence of polycomb-mediated repression.

The results support a model of HCV infection influencing the binding of transcription factors to cognate sites in the genome, with consequent local acquisition of DNA methylation, and the added repressive influence of polycomb at a subset of CG-dense cis-regulatory sequences. These epigenetic events occur before neoplastic transformation, resulting in what may be a pharmacologically-reversible epigenetic field defect in HCV-infected liver.

No MeSH data available.


Related in: MedlinePlus

Identification of subsets if loci with distinctive progressive changes in DNA methylation. Panel (a) shows the comparison between malignant and control samples, with the majority of loci losing DNA methylation (green) with malignant transformation. In panel (b), k-means clustering shows the majority of loci gaining DNA methylation to do so in the infected stage (groups III and IV), but the loci losing DNA methylation only have this occur in the transition from infected to cancer (groups V and VI).
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Figure 2: Identification of subsets if loci with distinctive progressive changes in DNA methylation. Panel (a) shows the comparison between malignant and control samples, with the majority of loci losing DNA methylation (green) with malignant transformation. In panel (b), k-means clustering shows the majority of loci gaining DNA methylation to do so in the infected stage (groups III and IV), but the loci losing DNA methylation only have this occur in the transition from infected to cancer (groups V and VI).

Mentions: This analysis confirmed the impression of the heat map of Figure 1, that the malignant samples have a greater number of loci with significant loss (16,921) than gain (6,894) of DNA methylation relative to control samples (Figure 2a). Linking the identified K-means clustering groups with this restricted set of loci shows the loss of DNA methylation to be a late event, mostly occurring in the transition from the infected to malignant states (groups V-VI, Figure 2b), whereas the smaller subset of loci gaining DNA methylation does so at the earlier, infected stage (groups III-IV, Figure 2b).


A pre-neoplastic epigenetic field defect in HCV-infected liver at transcription factor binding sites and polycomb targets
Identification of subsets if loci with distinctive progressive changes in DNA methylation. Panel (a) shows the comparison between malignant and control samples, with the majority of loci losing DNA methylation (green) with malignant transformation. In panel (b), k-means clustering shows the majority of loci gaining DNA methylation to do so in the infected stage (groups III and IV), but the loci losing DNA methylation only have this occur in the transition from infected to cancer (groups V and VI).
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Related In: Results  -  Collection

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Figure 2: Identification of subsets if loci with distinctive progressive changes in DNA methylation. Panel (a) shows the comparison between malignant and control samples, with the majority of loci losing DNA methylation (green) with malignant transformation. In panel (b), k-means clustering shows the majority of loci gaining DNA methylation to do so in the infected stage (groups III and IV), but the loci losing DNA methylation only have this occur in the transition from infected to cancer (groups V and VI).
Mentions: This analysis confirmed the impression of the heat map of Figure 1, that the malignant samples have a greater number of loci with significant loss (16,921) than gain (6,894) of DNA methylation relative to control samples (Figure 2a). Linking the identified K-means clustering groups with this restricted set of loci shows the loss of DNA methylation to be a late event, mostly occurring in the transition from the infected to malignant states (groups V-VI, Figure 2b), whereas the smaller subset of loci gaining DNA methylation does so at the earlier, infected stage (groups III-IV, Figure 2b).

View Article: PubMed Central - PubMed

ABSTRACT

The predisposition of patients with Hepatitis C virus (HCV) infection to hepatocellular carcinoma (HCC) involves components of viral infection, inflammation and time. The development of multifocal, genetically distinct tumors is suggestive of a field defect affecting the entire liver. The molecular susceptibility mediating such a field defect is not understood. One potential mediator of long-term cellular reprogramming is heritable (epigenetic) regulation of transcription, exemplified by DNA methylation. We studied epigenetic and transcriptional changes in HCV-infected livers in comparison with control, uninfected livers and HCC, allowing us to identify pre-neoplastic epigenetic and transcriptional events.

We find the HCV-infected liver to have a pattern of acquisition of DNA methylation targeted to candidate enhancers active in liver cells, enriched for the binding sites of the FOXA1, FOXA2 and HNF4A transcription factors. These enhancers can be subdivided into those proximal to genes implicated in liver cancer or to genes involved in stem cell development, the latter distinguished by increased CG dinucleotide density and polycomb-mediated repression, manifested by the additional acquisition of histone H3 lysine 27 trimethylation (H3K27me3). Transcriptional studies on our samples showed that the increased DNA methylation at enhancers was associated with decreased local gene expression, results validated in independent samples from The Cancer Genome Atlas (TCGA). Pharmacological depletion of H3K27me3 using the EZH2 inhibitor GSK343 in HepG2 cells suppressed cell growth and also revealed that local acquired DNA methylation was not dependent upon the presence of polycomb-mediated repression.

The results support a model of HCV infection influencing the binding of transcription factors to cognate sites in the genome, with consequent local acquisition of DNA methylation, and the added repressive influence of polycomb at a subset of CG-dense cis-regulatory sequences. These epigenetic events occur before neoplastic transformation, resulting in what may be a pharmacologically-reversible epigenetic field defect in HCV-infected liver.

No MeSH data available.


Related in: MedlinePlus