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A phosphorylated subpopulation of the histone variant macroH2A1 is excluded from the inactive X chromosome and enriched during mitosis.

ABSTRACT: Histone variants play an important role in numerous biological processes through changes in nucleosome structure and stability and possibly through mechanisms influenced by posttranslational modifications unique to a histone variant. The family of histone H2A variants includes members such as H2A.Z, the DNA damage-associated H2A.X, macroH2A (mH2A), and H2ABbd (Barr body-deficient). Here, we have undertaken the challenge to decipher the posttranslational modification-mediated "histone code" of mH2A, a variant generally associated with certain forms of condensed chromatin such as the inactive X chromosome in female mammals. By using female human cells as a source of mH2A, endogenous mH2A was purified and analyzed by mass spectrometry. Although mH2A is in low abundance compared with conventional histones, we identified a phosphorylation site, S137ph, which resides within the "hinge" region of mH2A. This lysine-rich hinge is an approximately 30-aa stretch between the H2A and macro domains, proposed to bind nucleic acids. A specific antibody to S137ph was raised; by using this reagent, S137 phosphorylation was found to be present in both male and female cells and on both splice variants of the mH2A1 gene. Although mH2A is generally enriched on the inactive X chromosome in female cells, mH2AS137ph is excluded from this heterochromatic structure. Thus, a phosphorylated subpopulation of mH2A appears to play a unique role in chromatin regulation beyond X inactivation. We provide evidence that S137ph is enriched in mitosis, suggestive of a role in the regulation of mH2A posttranslational modifications throughout the cell cycle.

SUBMITTER: Bernstein E

PROVIDER: S-EPMC2234179 | biostudies-literature | 2008 Feb

REPOSITORIES: biostudies-literature

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A phosphorylated subpopulation of the histone variant macroH2A1 is excluded from the inactive X chromosome and enriched during mitosis.

Bernstein Emily E Muratore-Schroeder Tara L TL Diaz Robert L RL Chow Jennifer C JC Changolkar Lakshmi N LN Shabanowitz Jeffrey J Heard Edith E Pehrson John R JR Hunt Donald F DF Allis C David CD

Proceedings of the National Academy of Sciences of the United States of America 20080128 5

Histone variants play an important role in numerous biological processes through changes in nucleosome structure and stability and possibly through mechanisms influenced by posttranslational modifications unique to a histone variant. The family of histone H2A variants includes members such as H2A.Z, the DNA damage-associated H2A.X, macroH2A (mH2A), and H2ABbd (Barr body-deficient). Here, we have undertaken the challenge to decipher the posttranslational modification-mediated "histone code" of mH ...[more]

PMID: 18227505

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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.

Dataset Information

A phosphorylated subpopulation of the histone variant macroH2A1 is excluded from the inactive X chromosome and enriched during mitosis.

Publications

A phosphorylated subpopulation of the histone variant macroH2A1 is excluded from the inactive X chromosome and enriched during mitosis.

Similar Datasets

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