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DNA Hypomethylation and Histone Variant macroH2A1 Synergistically Attenuate Chemotherapy-Induced Senescence to Promote Hepatocellular Carcinoma Progression.

ABSTRACT: Aging is a major risk factor for progression of liver diseases to hepatocellular carcinoma (HCC). Cellular senescence contributes to age-related tissue dysfunction, but the epigenetic basis underlying drug-induced senescence remains unclear. macroH2A1, a variant of histone H2A, is a marker of senescence-associated heterochromatic foci that synergizes with DNA methylation to silence tumor-suppressor genes in human fibroblasts. In this study, we investigated the relationship between macroH2A1 splice variants, macroH2A1.1 and macroH2A1.2, and liver carcinogenesis. We found that protein levels of both macroH2A1 isoforms were increased in the livers of very elderly rodents and humans, and were robust immunohistochemical markers of human cirrhosis and HCC. In response to the chemotherapeutic and DNA-demethylating agent 5-aza-deoxycytidine (5-aza-dC), transgenic expression of macroH2A1 isoforms in HCC cell lines prevented the emergence of a senescent-like phenotype and induced synergistic global DNA hypomethylation. Conversely, macroH2A1 depletion amplified the antiproliferative effects of 5-aza-dC in HCC cells, but failed to enhance senescence. Senescence-associated secretory phenotype and whole-transcriptome analyses implicated the p38 MAPK/IL8 pathway in mediating macroH2A1-dependent escape of HCC cells from chemotherapy-induced senescence. Furthermore, chromatin immunoprecipitation sequencing revealed that this hepatic antisenescence state also required active transcription that could not be attributed to genomic occupancy of these histones. Collectively, our findings reveal a new mechanism by which drug-induced senescence is epigenetically regulated by macroH2A1 and DNA methylation and suggest macroH2A1 as a novel biomarker of hepatic senescence that could potentially predict prognosis and disease progression.

SUBMITTER: Borghesan M

PROVIDER: S-EPMC4785885 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature

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DNA Hypomethylation and Histone Variant macroH2A1 Synergistically Attenuate Chemotherapy-Induced Senescence to Promote Hepatocellular Carcinoma Progression.

Borghesan Michela M Fusilli Caterina C Rappa Francesca F Panebianco Concetta C Rizzo Giovanni G Oben Jude A JA Mazzoccoli Gianluigi G Faulkes Chris C Pata Illar I Agodi Antonella A Rezaee Farhad F Minogue Shane S Warren Alessandra A Peterson Abigail A Sedivy John M JM Douet Julien J Buschbeck Marcus M Cappello Francesco F Mazza Tommaso T Pazienza Valerio V Vinciguerra Manlio M

Cancer research 20160115 3

Aging is a major risk factor for progression of liver diseases to hepatocellular carcinoma (HCC). Cellular senescence contributes to age-related tissue dysfunction, but the epigenetic basis underlying drug-induced senescence remains unclear. macroH2A1, a variant of histone H2A, is a marker of senescence-associated heterochromatic foci that synergizes with DNA methylation to silence tumor-suppressor genes in human fibroblasts. In this study, we investigated the relationship between macroH2A1 spli ...[more]

PMID: 26772755

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Project description:BackgroundProstate cancer (PCa), a major cause of cancer-related morbidity and mortality worldwide and mostly asymptomatic at earliest stages, is characterized by disruption of genetic and epigenetic balance. A better understanding of how those mechanisms orchestrate disease might improve diagnostic and prognostic tools, allowing for improvements in treatment efficacy. Replacement of canonical histones, an epigenetic mechanism, is highly conserved among species and altered expression of histones variants (e.g., MacroH2A1) has been associated with tumorigenesis. H2AFY gene encodes two isoforms of H2A histone variant MacroH2A1: MacroH2A1.1 and MacroH2A1.2. Specifically, MacroH2A1.1 isoform inhibits cell proliferation and promotes cellular differentiation. Because the contribution of this histone variant to carcinogenesis has been reported in several cancer types, but not for PCa, we aimed to investigate the contribution of MacroH2A1 for prostate carcinogenesis.MethodsMacroH2A1, MacroH2A1.1 and MacroH2A1.2 isoforms and the corresponding splicing regulators transcript levels were evaluated by RT-qPCR, in a tissue cohort composed by PCa, prostatic intraepithelial neoplasia (PIN) and normal prostate cases. Knockdown for MacroH2A1 and MacroH2A1.1 was performed through lentiviral transduction in DU145 cells, and MacroH2A1.1 overexpression was achieved in LNCaP cells by plasmid transfection, followed by functional assays. Biological and/or experimental replicates were performed when necessary, and specific statistical tests were applied to perform data analysis.ResultsMacroH2A1.1 transcript levels were downregulated in PIN and primary PCa compared to normal prostate tissues. The same was found for QKI, a MacroH2A1.1's splicing regulator. Moreover, lower MacroH2A1.1 and QKI expression levels associated with less differentiated tumors (Gleason score ≥ 7). Interestingly, MacroH2A1.1, but more impressively DDX17 (AUC = 0.93; p < 0.0001) and QKI (AUC = 0.94; p < 0.0001), accurately discriminated cancerous from noncancerous prostate tissues. Furthermore, in PCa cell lines, total MacroH2A1 knockdown augmented malignant features, whereas MacroH2A1.1 overexpression impressively attenuated the malignant phenotype.ConclusionsOverall, our data, derived from primary PCa tissues and cell lines, anticipate a tumor suppressive role for MacroH2A1, particularly for the MacroH2A1.1 isoform, in prostate carcinogenesis.

Project description:Background: Gene expression in eukaryotic cells can be governed by histone variants, which replace replication-coupled histones, conferring unique chromatin properties. MacroH2A1 is a histone H2A variant containing a domain highly similar to H2A and a large non-histone (macro) domain. MacroH2A1, in turn, is present in two alternatively exon-spliced isoforms: macroH2A1.1 and macroH2A1.2, which regulate cell plasticity and proliferation in a remarkably distinct manner. The N-terminal and the C-terminal tails of H2A histones stem from the nucleosome core structure and can be target sites for several post-translational modifications (PTMs). MacroH2A1.1 and macroH2A1.2 isoforms differ only in a few amino acids and their ability to bind NAD-derived metabolites, a property allegedly conferring their different functions in vivo. Some of the modifications on the macroH2A1 variant have been identified, such as phosphorylation (T129, S138) and methylation (K18, K123, K239). However, no study to our knowledge has analyzed extensively, and in parallel, the PTM pattern of macroH2A1.1 and macroH2A1.2 in the same experimental setting, which could facilitate the understanding of their distinct biological functions in health and disease. Methods: We used a mass spectrometry-based approach to identify the sites for phosphorylation, acetylation, and methylation in green fluorescent protein (GFP)-tagged macroH2A1.1 and macroH2A1.2 expressed in human hepatoma cells. The impact of selected PTMs on macroH2A1.1 and macroH2A1.2 structure and function are demonstrated using computational analyses. Results: We identified K7 as a new acetylation site in both macroH2A1 isoforms. Quantitative comparison of histone marks between the two isoforms revealed significant differences in the levels of phosphorylated T129 and S170. Our computational analysis provided evidence that the phosphorylation status in the intrinsically disordered linker region in macroH2A1 isoforms might represent a key regulatory element contributing to their distinct biological responses. Conclusions: Taken together, our results report different PTMs on the two macroH2A1 splicing isoforms as responsible for their distinct features and distribution in the cell.

Dataset Information

DNA Hypomethylation and Histone Variant macroH2A1 Synergistically Attenuate Chemotherapy-Induced Senescence to Promote Hepatocellular Carcinoma Progression.

Publications

DNA Hypomethylation and Histone Variant macroH2A1 Synergistically Attenuate Chemotherapy-Induced Senescence to Promote Hepatocellular Carcinoma Progression.

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