Faculty Bibliography

Molecular signalling pathways delineating the induction of matrix metalloproteinases (MMPs) by ultraviolet radiation (UVR) are currently well-defined; however, the effects of UVR on epigenetic mechanisms of MMP induction are not as well understood. In this study, we examined solar-simulated UVR (ssUVR)-induced gene expression changes and alterations to histone methylation in the promoters of MMP1 and MMP3 in primary human dermal fibroblasts (HDF). Gene expression changes, including the increased expression of MMP1 and MMP3, were observed using Affymetrix GeneChip arrays and confirmed by qRT-PCR. Using ChIP-PCR, we showed for the first time that in HDF irradiated with 12 J/cm(2) ssUVR, the H3K4me3 transcriptional activating mark increased and the H3K9me2 transcriptional silencing mark decreased in abundance in promoters, correlating with the observed elevation of MMP1 and MMP3 mRNA levels following ssUVR exposure. Changes in mRNA levels due to a single exposure were transient and decreased 5 days after exposure.

Cadmium is an established human lung carcinogen with weak mutagenicity. However, the mechanisms underlying cadmium-induced carcinogenesis remain obscure. It has been suggested that epigenetic mechanisms may play a role in cadmium-induced carcinogenesis. In this study, we investigated the effects of cadmium on histone methylation and histone demethylases, and the role of histone methylation in transformation of immortalized normal human bronchial epithelial (BEAS-2B) cells. Exposure to 0.625, 1.25, 2.5, and 5.0 muM of cadmium for 6, 24, and 48 h increased global trimethylated histone H3 on lysine 4 (H3K4me3) and dimethylated histone H3 on lysine 9 (H3K9me2) in BEAS-2B cells compared with untreated cells, and most of these changes remained after the removal of cadmium (P < .05 or P < .01 for most modifications). Meanwhile, cadmium inhibited the activities of histone H3 on lysine 4 (H3K4) and histone H3 on lysine 9 (H3K9) demethylases which were detected by histone demethylation assay. However, there was no significant change in the protein levels of the H3K4 demethylase lysine-specific demethylase 5A (KDM5A) and the H3K9 demethylase lysine-specific demethylase 3A (KDM3A). Interestingly, during transformation of BEAS-2B cells by 20 weeks of exposure to 2.0 muM cadmium as assessed by anchorage-independent growth in soft agar, global H3K4me3, and H3K9me2 were significantly increased at 4 weeks (P < .05 or P < .01), whereas no significant change was observed at 8, 12, 16, and 20 weeks compared with control. Our study suggests that cadmium increases global H3K4me3 and H3K9me2 by inhibiting the activities of histone demethylases, and aberrant histone methylation that occurs early (48 h) and at 4 weeks is associated with cadmium-induced transformation of BEAS-2B cells at the early stage.

The JmjC domain-containing histone demethylases can remove histone lysine methylation and thereby regulate gene expression. The JmjC domain uses iron Fe(II) and alpha-ketoglutarate (alphaKG) as cofactors in an oxidative demethylation reaction via hydroxymethyl lysine. We hypothesize that reactive oxygen species will oxidize Fe(II) to Fe(III), thereby attenuating the activity of JmjC domain-containing histone demethylases. To minimize secondary responses from cells, extremely short periods of oxidative stress (3h) were used to investigate this question. Cells that were exposed to hydrogen peroxide (H2O2) for 3h exhibited increases in several histone methylation marks including H3K4me3 and decreases of histone acetylation marks including H3K9ac and H4K8ac; preincubation with ascorbate attenuated these changes. The oxidative stress level was measured by generation of 2',7'-dichlorofluorescein, GSH/GSSG ratio, and protein carbonyl content. A cell-free system indicated that H2O2 inhibited histone demethylase activity where increased Fe(II) rescued this inhibition. TET protein showed a decreased activity under oxidative stress. Cells exposed to a low-dose and long-term (3 weeks) oxidative stress also showed increased global levels of H3K4me3 and H3K27me3. However, these global methylation changes did not persist after washout. The cells exposed to short-term oxidative stress also appeared to have higher activity of class I/II histone deacetylase (HDAC) but not class III HDAC. In conclusion, we have found that oxidative stress transiently alters the epigenetic program process through modulating the activity of enzymes responsible for demethylation and deacetylation of histones.

The special AT-rich sequence-binding proteins 1 and 2 (SATB1/2) are nuclear matrix associated proteins that are transcription factors involved in chromatin remodeling and gene regulation. Expression of the SATB2 gene is tissue-specific, and the only epithelial cells expressing SATB2 are the glandular cells of the lower gastrointestinal tract where its expression is regulated by microRNA-31 (miR-31) and miR-182. SATB2, along with its homolog SATB1, are thought to be involved in various cancers with their roles in this disease being specific to the type of cancer. Colorectal cancer (CRC) provides the largest association of SATB2 with cancer and the roles of SATB2 are better defined and more studied in CRC than in any other cancer type. SATB1 displays a negative association with SATB2 in CRC. The various studies that have investigated the involvement of SATB1 and 2 in CRC have produced consistent findings. Here, we form four major conclusions regarding the role of these proteins in CRC and their potential clinical value: (i) SATB2 is a sensitive marker to distinguish CRC from other cancer types, (ii) Reduced expression of SATB2 in CRC is associated with poor prognosis, (iii) High levels of SATB1 expression facilitate CRC and are associated with poor prognosis and (iv) Overexpression of miR-31 and -182 in CRC leads to more aggressive cancer. This review will describe several of the key investigations that established these conclusions and highlight results that offer opportunities for future research in the treatment and diagnosis of CRC.

MOF was first identified in Drosophila melanogaster as an important component of the dosage compensation complex. As a member of MYST family of histone acetyltransferase, MOF specifically deposits the acetyl groups to histone H4 lysine 16. Throughout evolution, MOF and its mammalian ortholog have retained highly conserved substrate specificity and similar enzymatic activities. MOF plays important roles in dosage compensation, ESC self-renewal, DNA damage and repair, cell survival, and gene expression regulation. Dysregulation of MOF has been implicated in tumor formation and progression of many types of human cancers. This review will discuss the structure and activity of mammalian hMOF as well as its function in H4K16 acetylation, DNA damage response, stem cell pluripotency, and carcinogenesis.

The mechanisms that underlie metal carcinogenesis are the subject of intense investigation; however, data from in vitro and in vivo studies are starting to piece together a story that implicates epigenetics as a key player. Data from our lab has shown that nickel compounds inhibit dioxygenase enzymes by displacing iron in the active site. Arsenic is hypothesized to inhibit these enzymes by diminishing ascorbate levels - an important co-factor for dioxygenases. Inhibition of histone demethylase dioxygenases can increase histone methylation levels, which also may affect gene expression. Recently, our lab conducted a series of investigations in human subjects exposed to high levels of nickel or arsenic compounds. Global levels of histone modifications in peripheral blood mononuclear cells (PBMCs) from exposed subjects were compared to low environmentally exposed controls. Results showed that nickel increased H3K4me3 and decreased H3K9me2 globally. Arsenic increased H3K9me2 and decreased H3K9ac globally. Other histone modifications affected by arsenic were sex-dependent. Nickel affected the expression of 2756 genes in human PBMCs and many of the genes were involved in immune and carcinogenic pathways. This review will describe data from our lab that demonstrates for the first time that nickel and arsenic compounds affect global levels of histone modifications and gene expression in exposed human populations.

Particulate matter (PM) exposures have been linked to mortality, low birth weights, hospital admissions, and diseases associated with metabolic syndrome, including diabetes mellitus, cardiovascular disease, and obesity. In a previous in vitro and in vivo study, data demonstrated that PM10mum collected from Jeddah, Saudi Arabia (PMSA), altered expression of genes involved in lipid and cholesterol metabolism, as well as many other genes associated with metabolic disorders. PMSA contains a relatively high concentration of nickel (Ni), known to be linked to several metabolic disorders. In order to evaluate whether Ni and PM exposures induce similar gene expression profiles, mice were exposed to 100 mug/50 mul PMSA (PM-100), 50 mug/50 mul nickel chloride (Ni-50), or 100 mug/50 mul nickel chloride (Ni-100) twice per week for 4 wk and hepatic gene expression changes were determined. Ultimately, 55 of the same genes were altered in all 3 exposures. However, where the two Ni groups differed markedly was in the regulation (up or down) of these genes. Ni-100 and PM-100 groups displayed similar regulations, whereby 104 of the 107 genes were similarly modulated. Many of the 107 genes are involved in metabolic syndrome and include ALDH4A1, BCO2, CYP1A, CYP2U, TOP2A. In addition, the top affected pathways, such as fatty acid alpha-oxidation, and lipid and carbohydrate metabolism, are involved in metabolic diseases. Most notably, the top diseased outcome affected by these changes in gene expression was cardiovascular disease. Given these data, it appears that Ni and PMSA exposures display similar gene expression profiles, modulating the expression of genes involved in metabolic disorders.

The replication-dependent histone genes are the only metazoan genes whose messenger RNA (mRNA) does not terminate with a poly(A) tail at the 3' end. Instead, the histone mRNAs display a stem-loop structure at their 3' end. Stem-loop binding protein (SLBP) binds the stem-loop and regulates canonical histone mRNA metabolism. Here we report that exposure to arsenic, a carcinogenic metal, decreases cellular levels of SLBP by inducing its proteasomal degradation and inhibiting SLBP transcription via epigenetic mechanisms. Notably, arsenic exposure dramatically increases polyadenylation of canonical histone H3.1 mRNA possibly through downregulation of SLBP expression. The polyadenylated H3.1 mRNA induced by arsenic is not susceptible to normal degradation that occurs at the end of S phase, resulting in continued presence into mitosis, increased total H3.1 mRNA, and increased H3 protein levels. Excess expression of canonical histones has been shown to increase sensitivity to DNA damage, as well as increase the frequency of missing chromosomes and induce genomic instability. Thus, polyadenylation of canonical histone mRNA following arsenic exposure may contribute to arsenic-induced carcinogenesis.