We have previously reported that carcinogenic nickel compounds decreased global histone H4 acetylation and silenced the gpt transgene in G12 Chinese hamster cells. However, the nature of this silencing is still not clear. Here, we report that nickel ion exposure increases global H3K9 mono-and dimethylation, both of which are critical marks for DNA methylation and long-term gene silencing. In contrast to the up-regulation of global H3K9 dimethylation, nickel ions decreased the expression and activity of histone H3K9 specific methyltransferase G9a. Further investigation demonstrated that nickel ions interfered with the removal of histone methylation in vivo and directly decreased the activity of a Fe(II)-2-oxoglutarate-dependent histone H3K9 demethylase in nuclear extract in vitro. These results are the first to show a histone H3K9 demethylase activity dependent on both iron and 2-oxoglutarate. Exposure to nickel ions also increased H3K9 dimethylation at the gpt locus in G12 cells and repressed the expression of the gpt transgene. An extended nickel ion exposure led to increased frequency of the gpt transgene silencing, which was readily reversed by treatment with DNAdemethylating agent 5-aza-2-deoxycytidine. Collectively, our data strongly indicate that nickel ions induce transgene silencing by increasing histone H3K9 dimethylation, and this effect is mediated by the inhibition of H3K9 demethylation.Posttranslational modifications of histone N-terminal tails are important in chromatin organization, gene transcription, and DNA replication and repair (19). To date, a diverse array of histone modifications has been identified, including acetylation, methylation, phosphorylation, and ubiquitination (28). Among them, methylation of histone H3 lysine 9 (H3K9) is one of the best-studied modifications. H3K9 may be mono-, di-, or trimethylated without changing the positive charge of the lysine residue. Trimethylated H3K9 is typically connected with constitutive heterochromatin, while mono-and dimethylated H3K9 are mainly located in euchromatin and generally linked to repressed promoter regions (29). Suv39h family enzymes are responsible for trimethylation of H3K9 in vivo (27,29), while G9a and GLP/EuHMTase 1 are two major histone methyltransferases responsible for H3K9 dimethylation in vivo (35,36). Genetic ablation of either G9a or GLP/EuHMTase 1 dramatically diminished global H3K9 dimethylation in mouse embryonic stem cells (35,36).Methylation of histone lysines had long been thought of as a "permanent" modification since there was no known enzyme to demethylate. However, this dogma was challenged by the recent discoveries of histone H3 lysine 4 (H3K4) demethylase LSD1 and H3 lysine 36 (H3K36) demethylase JHDM1 (JmjC domain-containing histone demethylase 1) (32, 39). Although both LSD1 and JHDM1 can remove the methyl group from lysine residues on histone H3, they utilize different mechanisms to demethylate. LSD1 is a flavin-dependent amine oxidase and removes the methyl group from mono-or dimethyl H3K4 by catalyzing the...

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Histone H3 lysine 4 (H3K4) trimethylation (H3K4me3) at the promoter region of genes has been linked to transcriptional activation. In the present study, we found that hypoxia (1% oxygen) increased H3K4me3 in both normal human bronchial epithelial Beas-2B cells and human lung carcinoma A549 cells. The increase of H3K4me3 from hypoxia was likely caused by the inhibition of H3K4 demethylating activity, as hypoxia still increased H3K4me3 in methionine-deficient medium. Furthermore, an in vitro histone demethylation assay showed that 1% oxygen decreased the activity of H3K4 demethylases in Beas-2B nuclear extracts because ambient oxygen tensions were required for the demethylation reaction to proceed. Hypoxia only minimally increased H3K4me3 in the BEAS-2B cells with knockdown of JARID1A, which is the major histone H3K4 demethylase in this cell line. However, the mRNA and protein levels of JARID1A were not affected by hypoxia. GeneChip and pathway analysis in JARID1A knockdown Beas-2B cells revealed that JARID1A regulates the expression of hundreds of genes involved in different cellular functions, including tumorigenesis. Knocking down of JARID1A increased H3K4me3 at the promoters of HMOX1 and DAF genes. Thus, these results indicate that hypoxia might target JARID1A activity, which in turn increases H3K4me3 at both the global and genespecific levels, leading to the altered programs of gene expression and tumor progression.