Targeting histone demethylases JMJD3 and UTX: selenium as a potential therapeutic agent for cervical cancer.

BACKGROUND: The intriguing connection between selenium and cancer resembles a captivating puzzle that keeps researchers engaged and curious. While selenium has shown promise in reducing cancer risks through supplementation, its interaction with epigenetics in cervical cancer remains a fascinating yet largely unexplored realm. Unraveling the intricacies of selenium's role and its interaction with epigenetic factors could unlock valuable insights in the battle against this complex disease. RESULT: Selenium has shown remarkable inhibitory effects on cervical cancer cells in various ways. In in vitro studies, it effectively inhibits the proliferation, migration, and invasion of cervical cancer cells, while promoting apoptosis. Selenium also demonstrates significant inhibitory effects on human cervical cancer-derived organoids. Furthermore, in an in vivo study, the administration of selenium dioxide solution effectively suppresses the growth of cervical cancer tumors in mice. One of the mechanisms behind selenium's inhibitory effects is its ability to inhibit histone demethylases, specifically JMJD3 and UTX. This inhibition is observed both in vitro and in vivo. Notably, when JMJD3 and UTX are inhibited with GSK-J4, similar biological effects are observed in both in vitro and in vivo models, effectively inhibiting organoid models derived from cervical cancer patients. Inhibiting JMJD3 and UTX also induces G2/M phase arrest, promotes cellular apoptosis, and reverses epithelial-mesenchymal transition (EMT). ChIP-qPCR analysis confirms that JMJD3 and UTX inhibition increases the recruitment of a specific histone modification, H3K27me3, to the transcription start sites (TSS) of target genes in cervical cancer cells (HeLa and SiHa cells). Furthermore, the expressions of JMJD3 and UTX are found to be significantly higher in cervical cancer tissues compared to adjacent normal cervical tissues, suggesting their potential as therapeutic targets. CONCLUSIONS: Our study highlights the significant inhibitory effects of selenium on the growth, migration, and invasion of cervical cancer cells, promoting apoptosis and displaying promising potential as a therapeutic agent. We identified the histone demethylases JMJD3 and UTX as specific targets of selenium, and their inhibition replicates the observed effects on cancer cell behavior. These findings suggest that JMJD3 and UTX could be valuable targets for selenium-based treatments of cervical cancer.

[Mechanism of Puerariae Lobatae Radix against lung cancer by inhibiting histone demethylase LSD1].

Histone lysine-specific demethylase 1(LSD1) has become a promising molecular target for lung cancer therapy. Upon the screening platform for LSD1 activity, some Chinese herbal extracts were screened for LSD1 activity inhibition, and the underlying mechanism was preliminarily investigated at both molecular and cellular levels. The results of LSD1 inhibition showed that Puerariae Lobatae Radix extract can effectively reduce LSD1 expression to elevate the expression of H3 K4 me2 and H3 K9 me2 substrates in H1975 and H1299 cells. Furthermore, Puerariae Lobatae Radix was evaluated for its anti-lung cancer activity. It had a potent inhibitory ability against the proliferation and colony formation of both H1975 and H1299 cells. Flow cytometry and DAPI staining assays indicated that Puerariae Lobatae Radix can induce the apoptosis of lung cancer cells. In addition, it can significantly suppress the migration and reverse the epithelial-mesenchymal transition(EMT) process of lung cancer cells by activating E-cadherin and suppressing the expression of N-cadherin, slug and vimentin. To sum up, Puerariae Lobatae Radix displayed a robust inhibitory activity against lung cancer, and the mechanism may be related to the down-regulation of LSD1 expression to induce the cell apoptosis and suppress the cell migration and EMT process. These findings will provide new insights into the action of Puerariae Lobatae Radix as an anti-lung cancer agent and offer new ideas for the study on the anti-cancer action of Chinese medicine based on the epigenetic modification.

[Expression pattern of the histone lysine demethylase family and its potential role in bladder cancer: a multi-omics analysis].

OBJECTIVE: To investigate the expression patterns of 19 histone lysine demethylases (KDMs) and their role in bladder cancer. METHODS: In this study, UALCAN and GSCALite were used to analyze the transcriptional expression, methylation level and somatic variation of KDMs in bladder cancer samples from TCGA. Kaplan Meier-Plotter and Assistant for clinical bioinformatics were used to investigate the effect of KDMs expression on the prognosis of BLCA samples. The immune infiltration and drug sensitivity of KDMs in bladder cancer were analyzed by Timer and GSCALite. RESULTS: The KDMs did not show consistent expressions patterns in bladder cancer, where the expressions of KDM1A/1B/2B/4A/4B/5B/5C were significantly upregulated while those of KDM3B/6B/7C were significantly downregulated. Methylation data analysis showed that methylation levels of KDM1A/3B/4A/4B/4C/5A/5B/5C/7B were significantly downregulated and that of KDM7C was upregulated. The transcription levels of 14 KDMs had significant negative correlations with their methylation levels, and among them KDM1A showed the strongest correlation. Mutation analysis revealed that KDM6A had the highest frequency of nonsynonymous mutations with the largest variety, and these mutations were complementary to nonsynonymous mutations of the other KDMs. Survival analysis showed that KDM3A/4C/5D/6A/7B were protective for OS while KDM3B/5B/5C adversely affected RFS of BLCA patients. Further comprehensive prognostic modeling confirmed that KDM4C/6A/7B were potential prognostic biomarkers of bladder cancer, and their expressions were positively correlated with immune infiltration in BLCA patients. KDM2B/3B/4B/4C/5A were negatively correlated with the sensitivity to most anticancer drugs, while KDM2B/4B were positively correlated with the sensitivity to 4 anticancer drugs. CONCLUSION: The expression patterns of the KDMs in bladder cancer highlight a high mutation complementarity and a negative correlation between over-expression and hypomethylation level closely related with the prognosis, immune infiltration and drug sensitivity.

X-linked histone H3K27 demethylase Kdm6a regulates sexually dimorphic differentiation of hypothalamic neurons.

Several X-linked genes are involved in neuronal differentiation and may contribute to the generation of sex dimorphisms in the brain. Previous results showed that XX hypothalamic neurons grow faster, have longer axons, and exhibit higher expression of the neuritogenic gene neurogenin 3 (Ngn3) than XY before perinatal masculinization. Here we evaluated the participation of candidate X-linked genes in the development of these sex differences, focusing mainly on Kdm6a, a gene encoding for an H3K27 demethylase with functions controlling gene expression genome-wide. We established hypothalamic neuronal cultures from wild-type or transgenic Four Core Genotypes mice, a model that allows evaluating the effect of sex chromosomes independently of gonadal type. X-linked genes Kdm6a, Eif2s3x and Ddx3x showed higher expression in XX compared to XY neurons, regardless of gonadal sex. Moreover, Kdm6a expression pattern with higher mRNA levels in XX than XY did not change with age at E14, P0, and P60 in hypothalamus or under 17ß-estradiol treatment in culture. Kdm6a pharmacological blockade by GSK-J4 reduced axonal length only in female neurons and decreased the expression of neuritogenic genes Neurod1, Neurod2 and Cdk5r1 in both sexes equally, while a sex-specific effect was observed in Ngn3. Finally, Kdm6a downregulation using siRNA reduced axonal length and Ngn3 expression only in female neurons, abolishing the sex differences observed in control conditions. Altogether, these results point to Kdm6a as a key mediator of the higher axogenesis and Ngn3 expression observed in XX neurons before the critical period of brain masculinization.

KDM6A promotes imatinib resistance through YY1-mediated transcriptional upregulation of TRKA independently of its demethylase activity in chronic myelogenous leukemia.

Rationale: Despite landmark therapy of chronic myelogenous leukemia (CML) with tyrosine kinase inhibitors (TKIs), drug resistance remains problematic. Cancer pathogenesis involves epigenetic dysregulation and in particular, histone lysine demethylases (KDMs) have been implicated in TKI resistance. We sought to identify KDMs with altered expression in CML and define their contribution to imatinib resistance. Methods: Bioinformatics screening compared KDM expression in CML versus normal bone marrow with shRNA knockdown and flow cytometry used to measure effects on imatinib-induced apoptosis in K562 cells. Transcriptomic analyses were performed against KDM6A CRISPR knockout/shRNA knockdown K562 cells along with gene rescue experiments using wildtype and mutant demethylase-dead KDM6A constructs. Co-immunoprecipitation, luciferase reporter and ChIP were employed to elucidate mechanisms of KDM6A-dependent resistance. Results: Amongst five KDMs upregulated in CML, only KDM6A depletion sensitized CML cells to imatinib-induced apoptosis. Re-introduction of demethylase-dead KDM6A as well as wild-type KDM6A restored imatinib resistance. RNA-seq identified NTRK1 gene downregulation after depletion of KDM6A. Moreover, NTRK1 expression positively correlated with KDM6A in a subset of clinical CML samples and KDM6A knockdown in fresh CML isolates decreased NTRK1 encoded protein (TRKA) expression. Mechanistically, KDM6A was recruited to the NTRK1 promoter by the transcription factor YY1 with subsequent TRKA upregulation activating down-stream survival pathways to invoke imatinib resistance. Conclusion: Contrary to its reported role as a tumor suppressor and independent of its demethylase function, KDM6A promotes imatinib-resistance in CML cells. The identification of the KDM6A/YY1/TRKA axis as a novel imatinib-resistance mechanism represents an unexplored avenue to overcome TKI resistance in CML.

Epigallocatechin-3-gallate (EGCG) Alters Histone Acetylation and Methylation and Impacts Chromatin Architecture Profile in Human Endothelial Cells.

Epigallocatechin gallate (EGCG), the main green tea polyphenol, exerts a wide variety of biological actions. Epigenetically, the catechin has been classified as a DNMTs inhibitor, however, its impact on histone modifications and chromatin structure is still poorly understood. The purpose of this study was to find the impact of EGCG on the histone posttranslational modifications machinery and chromatin remodeling in human endothelial cells of both microvascular (HMEC-1) and vein (HUVECs) origin. We analyzed the methylation and acetylation status of histones (Western blotting), as well as assessed the activity (fluorometric assay kit) and gene expression (qPCR) of the enzymes playing a prominent role in shaping the human epigenome. The performed analyses showed that EGCG increases histone acetylation (H3K9/14ac, H3ac), and methylation of both active (H3K4me3) and repressive (H3K9me3) chromatin marks. We also found that the catechin acts as an HDAC inhibitor in cellular and cell-free models. Additionally, we observed that EGCG affects chromatin architecture by reducing the expression of heterochromatin binding proteins: HP1α, HP1γ. Our results indicate that EGCG promotes chromatin relaxation in human endothelial cells and presents a broad epigenetic potential affecting expression and activity of epigenome modulators including HDAC5 and 7, p300, CREBP, LSD1 or KMT2A.

Inhibition of lysine-specific histone demethylase 1A results in meiotic aberration during oocyte maturation in vitro in goats.

Histone methylation is associated with oocyte maturation in several species and is also expected in goat oocytes, while the mechanism is still unclear. Therefore, single-cell RNA sequencing (scRNA-seq) was performed on goat germinal vesicle (GV) and metaphase II (MII) oocytes, and the functions of lysine-specific histone demethylase 1A (LSD1), one of the differentially expressed genes (DEGs) were investigated during in vitro maturation (IVM) of goat oocytes. Through scRNA-seq, 4516 DEGs were identified from GV oocytes and MII oocytes in goats, among which there were 16 histone methyltransferase and demethylase DEGs (including LSD1). The functions of LSD1 during IVM of goat oocytes were investigated through its inhibitor, GSK-LSD1. We found that the first polar body extrusion rate of goat oocytes significantly reduced with an increase in GSK-LSD1 concentration supplemented into IVM medium (0 µM: 58.84 ± 0.95%; 2.5 µM: 52.14 ± 0.51%, P < 0.01; 50 µM: 41.22 ± 0.42%, P < 0.001; 100 µM: 29.78 ± 1.78%, P < 0.001). Moreover, compared with the control group, the level of H3K4me2 methylation and p-H2AX in goat oocytes significantly increased (P < 0.001 and P < 0.01, respectively) upon 50-µM GSK-LSD1 treatment for 12 h. Furthermore, abnormalities in spindle assembly (25.94 ± 1.02% vs. 71.15 ± 3.32%; P < 0.01) and chromosome alignment (22.93 ± 1.11% vs. 76.03 ± 3.25%; P < 0.01) were observed, and cytoskeletal organization (15.31 ± 1.60% vs. 67.50 ± 3.09%; P < 0.001) was disrupted upon treatment with 50-µM GSK-LSD1 for 12 h, which compared with that in the control group. Additionally, the ratio of BCL2:BAX significantly higher (P < 0.01) in oocytes with 50-µM GSK-LSD1 treatment than that in control group. Collectively, these results indicate the important role of LSD1 in meiotic maturation of goat oocytes. Our data not only clarify dynamic changes in mRNA during oocyte maturation but also provide a theoretical basis and technical means for further studies of meiotic maturation of goat oocytes.

Methylation of H3K27 and H3K4 in key gene promoter regions of thymus in RA mice is involved in the abnormal development and differentiation of iNKT cells.

Epigenetic modifications have been shown to be important for immune cell differentiation by regulating gene transcription. However, the role and mechanism of histone methylation in the development and differentiation of iNKT cells in rheumatoid arthritis (RA) mice have yet to be deciphered. The DBA/1 mouse RA model was established by using a modified GPI mixed peptide. We demonstrated that total peripheral blood, thymus, and spleen iNKT cells in RA mice decreased significantly, while iNKT1 in the thymus and spleen was increased significantly. PLZF protein and PLZF mRNA levels were significantly decreased in thymus DP T cells, while T-bet protein and mRNA were significantly increased in thymus iNKT cells. We found a marked accumulation in H3K27me3 around the promoter regions of the signature gene Zbtb16 in RA mice thymus DP T cells, and an accumulation of H3K4me3 around the promoters of the Tbx21 gene in iNKT cells. The expression levels of UTX in the thymus of RA mice were significantly reduced. The changes in the above indicators were particularly significant in the progressive phase of inflammation (11 days after modeling) and the peak phase of inflammation (14 days after modeling) in RA mice. Developmental and differentiation defects of iNKT cells in RA mice were associated with abnormal methylation levels (H3K27me3 and H3K4me3) in the promoters of key genes Zbtb16 (encoding PLZF) and Tbx21 (encoding T-bet). Decreased UTX of thymus histone demethylase levels resulted in the accumulation of H3K27me3 modification.

Soybean (Glycine max) prevents the progression of breast cancer cells by downregulating the level of histone demethylase JMJD5.

BACKGROUND: Breast cancer is the first noticeable disease in female patients. Long-term use of soybean (Glycine max) may prevent the progression of cancer. However, the molecular mechanism for the functions of soybean remains unclear. Histone demethylase JMJD5, an important epigenetic molecule, is overexpressed in the progression of breast cancer suggesting that soybean may ameliorate cancer by affecting the expression of JMJD5. MATERIALS AND METHODS: To test the hypothesis, human breast cancer cell lines MCF-7 and MDA-MB-231 were treated with different concentrations of soybean and/or transfected with the plasmids pcDNA3.1-JMJD5 and pTZU6 + 1-shRNA-JMJD5. The growth rate was measured using xCELLigence real-time cell analysis. The level of JMJD5 was measured by using quantitative reverse transcription-polymerase chain reaction and Western blot. RESULTS: Soybean showed significant inhibitory effects on the growth rates ofMCF-7 and MDA-MB-231 cells in a concentration-dependent way (P < 0.05). Meanwhile, the levels of JMJD5 were reduced with the increase of soybean concentration (P < 0.05). JMJD5 transfection increased the growth rates of MCF-7 and MDA-MB-231 by 25% and 40%. In contrast, the growth rates of MCF-7 and MDA-MB-231 cells were decreased by 17% and 23% after being transfected with JMJD5 shRNA. Soybean inhibited the growth rate of MCF-7 and MDA-MB-231 cells when they were transfected by JMJD5 gene but no for the cells transfected with JMJD5 shRNA. CONCLUSION: The complicated compositions of soybean will be beneficial to the therapy of breast cancer since its causes may be involved in multiple aspects. Soybean represses breast cancer development by downregulating the level of JMJD5.

Exosomes from Suxiao Jiuxin pill-treated cardiac mesenchymal stem cells decrease H3K27 demethylase UTX expression in mouse cardiomyocytes in vitro.

Suxiao Jiuxin pill (SJP) is a traditional Chinese medicine for the treatment of acute coronary syndrome in China, which contains two principal components, tetramethylpyrazine (TMP) and borneol (BOR). Thus far, however, the molecular mechanisms underlying the beneficial effects of SJP on the cardiac microenvironment are unknown. Cardiac mesenchymal stem cells (C-MSCs) communicate with cardiomyocytes (CMs) through the release of microvesicles (exosomes) to restore cardiac homeostasis and elicit repair, in part through epigenetic regulatory mechanisms. In this study, we examined whether SJP treatment altered C-MSC-derived exosomes (SJP-Exos) to cause epigenetic chromatic remodeling in recipient CMs. C-MSC isolated from mouse hearts were pretreated with SJP (SJP-Exos), TMP (TMP-Exos) or BOR (BOR-Exos). Then, HL-1 cells, a mouse cardiomyocyte line, were treated with exosomes from control C-MSCs (Ctrl-Exos), SJP-Exos, TMP-Exos or BOR-Exos. Treatment with SJP-Exos significantly increased the protein levels of histone 3 lysine 27 trimethylation (H3K27me3), a key epigenetic chromatin marker for cardiac transcriptional suppression, in the HL-1 cells. To further explore the mechanisms of SJP-Exo-mediated H3K27me3 upregulation, we assessed the mRNA expression levels of key histone methylases (EZH1, EZH2 and EED) and demethylases (JMJD3 and UTX) in the exosome-treated HL-1 cells. Treatment with SJP-Exo selectively suppressed UTX expression in the recipient HL-1 cells. Furthermore, PCNA, an endogenous marker of cell replication, was significantly higher in SJP-Exo-treated HL-1 cells than in Ctrl-Exo-treated HL-1 cells. These results show that SJP-Exos increase cardiomyocyte proliferation and demonstrate that SJP can modulate C-MSC-derived exosomes to cause epigenetic chromatin remodeling in recipient cardiomyocytes; consequently, SJP-Exos might be used to promote cardiomyocyte proliferation.

Lysine-specific demethylase 1 activation by vitamin B2 attenuates efficacy of apatinib for proliferation and migration of gastric cancer cell MGC-803.

B vitamins play an essential role in the biosynthesis of nucleotides, replication of DNA, supply of methyl-groups, growth and repair of cells, aberrancies of which have all been implicated in carcinogenesis. Although the potential role of vitamin B in relation to the risk of cancer, including breast, and colorectal cancer, has been investigated in several observational studies, the mechanism of action is still unclear. In this study, vitamin B2 exhibited efficient activation of LSD1 by occupying the active sites where FAD stands. Interestingly, vitamin B2 significantly downregulated expression of CD86, a sensitive surrogate biomarker of LSD1 inhibition, and showed marked activation of gastric cancer cell migration and invasion. Meanwhile, vitamin B2 induced activation of LSD1 may attenuate the proliferation inhibition, and anti-migration effects of apatinib in gastric cancer cells. These findings suggested that vitamin B supplementation may interfere with the efficacy of apatinib in patients with gastric cancer.

Mechanisms of anti-cancer effects of ascorbate: Cytotoxic activity and epigenetic modulation.

Vitamin C (Vit C or Ascorbate) is essential for many fundamental biochemical processes. Vit C is an essential nutrient with redox functions at normal physiologic concentrations. The main physiologic function of this vitamin is related to its capacity to act as a co-factor for a large family of enzymes, collectively known as Fe and 2-oxoglutarate-dependent dioxygenases. It also modulates epigenetic gene expression through the control of TET enzymes activity. Vit C also has several biological properties allowing to restore the deregulated epigenetic response observed in many tumors. High-dose Vit C has been investigated as a treatment for cancer patients since the 1969. Pharmacologic ascorbate acts as a pro-drug for hydrogen peroxide formation (H2O2) and, through this mechanism, kills cancer cells. To achieve high in vivo concentrations, Ascorbate must be injected by i.v. route. Initial clinical studies of Ascorbate cancer treatment have provided encouraging results, not confirmed in subsequent studies. Recent clinical studies using i.v. injection of high-dose Ascorbate have renewed the interest in the field, showing that significant anti-tumor activity. Pre-clinical studies have led to identify tumors sensitive to Ascorbate that could potentially benefit from this treatment either through an epigenetic modulator effect or through tumor killing by oxidative stress.

Crosstalk between epigenetics and metabolism-Yin and Yang of histone demethylases and methyltransferases in cancer.

Histone methylation is an epigenetic modification of chromatin undergoing dynamic changes and balancing tissue-specific demands of proliferation and differentiation. In cancer, aberrant histone methylation can facilitate oncogenic and tumor suppression programs by modulating gene expression. Histone remodelers such as lysine methyltransferases and lysine demethylases are seemingly opposite or contrary forces but may be part of an interconnected network complementing each other. We identify several layers of molecular communication where epigenetic master regulators engage in crosstalk between tumor metabolism and histone remodeling. Epigenetic master regulators have the ability to cooperate with members of the transcriptional machinery, DNA methyltransferases, as well as other histone modifiers. High-throughput sequencing and omics data in combination with cancer systems biology analysis have the power to prioritize regulatory events epigenome-wide.

Yin-yang actions of histone methylation regulatory complexes in the brain.

Dysregulation of histone methylation has emerged as a major driver of neurodevelopmental disorders including intellectual disabilities and autism spectrum disorders. Histone methyl writer and eraser enzymes generally act within multisubunit complexes rather than in isolation. However, it remains largely elusive how such complexes cooperate to achieve the precise spatiotemporal gene expression in the developing brain. Histone H3K4 methylation (H3K4me) is a chromatin signature associated with active gene-regulatory elements. We review a body of literature that supports a model in which the RAI1-containing H3K4me writer complex counterbalances the LSD1-containing H3K4me eraser complex to ensure normal brain development. This model predicts H3K4me as the nexus of previously unrelated neurodevelopmental disorders.

Lysine-specific demethylase 5C promotes hepatocellular carcinoma cell invasion through inhibition BMP7 expression.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most common type of tumor and is associated with high morbidity and mortality rates. Patients with HCC routinely undergo surgery followed by adjuvant radiation therapy and chemotherapy. Despite such aggressive treatment approaches, median survival times remain under 1 year in most cases. KDM5C is a member of the family of JmjC domain-containing proteins that removes methyl residues from methylated lysine 4 on histone H3 lysine 4 (H3K4). KDM5C has been proposed as an oncogene in many types of tumors; however, its role and underlying mechanisms in HCC remain unclear. METHODS: Expression level of KDM5C was examined by RT-PCR, and IHC. Forced expression of KDM5C was mediated by retroviruses, and KDM5C was downregulated by shRNAs expressing lentiviruses. Migration and invasion of HCC cells was measured by wound healing, Transwell and Matrigel assays respectively. RESULTS: In this study, we report that KDM5C is abundantly expressed in invasive human HCC cells. Cellular depletion of KDM5C by shRNA inhibited HCC cell migration, invasion and epithelial-mesenchymal transition in vitro, and markedly decreased the metastasis capacity of invasive HCC cells in the liver and lung. Furthermore, ectopic expression of KDM5C in HCC cells promoted cell migration, invasion and epithelial-mesenchymal transition via the inactivation of BMP7. Knockdown of BMP7 significantly promotes shKDM5C-induced cell migration inhibition. CONCLUSIONS: Taken together, these data suggest that KDM5C-mediated BMP7 inactivation is essential for HCC cell invasion.

Cell-based assays to support the profiling of small molecules with histone methyltransferase and demethylase modulatory activity.

Histone methylation is a prevalent and dynamic chromatin modification, executed by the action of histone methyltransferases (HMTs) and demethylases (HDMs). Aberrant activity of many of these enzymes is associated with human disease, hence, there is a growing interest in identifying corresponding small molecule inhibitors with therapeutic potential. To date, most of the technologies supporting the identification of these inhibitors constitute in vitro biochemical assays which, although robust and sensitive, do not study HMTs and HDMs in their native cellular state nor provide information of inhibitor's cell permeability and toxicity. The evident need for complementary cellular approaches has recently propelled the development of cell-based assays that enable screening of HMT and HDM enzymes in a more relevant environment. Here, we highlight current cellular methodologies for HMT and HDM drug discovery support. We anticipate that implementation of these cell-based assays will positively impact the discovery of pharmacologically potent HMT and HDM inhibitors.

Histone lysine trimethylation or acetylation can be modulated by phytoestrogen, estrogen or anti-HDAC in breast cancer cell lines.

AIM: The isoflavones genistein, daidzein and equol (daidzein metabolite) have been reported to interact with epigenetic modifications, specifically hypermethylation of tumor suppressor genes. The objective of this study was to analyze and understand the mechanisms by which phytoestrogens act on chromatin in breast cancer cell lines. MATERIALS & METHODS: Two breast cancer cell lines, MCF-7 and MDA-MB 231, were treated with genistein (18.5 µM), daidzein (78.5 µM), equol (12.8 µM), 17ß-estradiol (10 nM) and suberoylanilide hydroxamic acid (1 µM) for 48 h. A control with untreated cells was performed. 17ß-estradiol and an anti-HDAC were used to compare their actions with phytoestrogens. The chromatin immunoprecipitation coupled with quantitative PCR was used to follow soy phytoestrogen effects on H3 and H4 histones on H3K27me3, H3K9me3, H3K4me3, H4K8ac and H3K4ac marks, and we selected six genes (EZH2, BRCA1, ERα, ERß, SRC3 and P300) for analysis. RESULTS: Soy phytoestrogens induced a decrease in trimethylated marks and an increase in acetylating marks studied at six selected genes. CONCLUSION: We demonstrated that soy phytoestrogens tend to modify transcription through the demethylation and acetylation of histones in breast cancer cell lines.

The H3K27 demethylase Utx regulates somatic and germ cell epigenetic reprogramming.

Induced pluripotent stem cells (iPSCs) can be derived from somatic cells by ectopic expression of different transcription factors, classically Oct4 (also known as Pou5f1), Sox2, Klf4 and Myc (abbreviated as OSKM). This process is accompanied by genome-wide epigenetic changes, but how these chromatin modifications are biochemically determined requires further investigation. Here we show in mice and humans that the histone H3 methylated Lys 27 (H3K27) demethylase Utx (also known as Kdm6a) regulates the efficient induction, rather than maintenance, of pluripotency. Murine embryonic stem cells lacking Utx can execute lineage commitment and contribute to adult chimaeric animals; however, somatic cells lacking Utx fail to robustly reprogram back to the ground state of pluripotency. Utx directly partners with OSK reprogramming factors and uses its histone demethylase catalytic activity to facilitate iPSC formation. Genomic analysis indicates that Utx depletion results in aberrant dynamics of H3K27me3 repressive chromatin demethylation in somatic cells undergoing reprogramming. The latter directly hampers the derepression of potent pluripotency promoting gene modules (including Sall1, Sall4 and Utf1), which can cooperatively substitute for exogenous OSK supplementation in iPSC formation. Remarkably, Utx safeguards the timely execution of H3K27me3 demethylation observed in embryonic day 10.5-11 primordial germ cells (PGCs), and Utx-deficient PGCs show cell-autonomous aberrant epigenetic reprogramming dynamics during their embryonic maturation in vivo. Subsequently, this disrupts PGC development by embryonic day 12.5, and leads to diminished germline transmission in mouse chimaeras generated from Utx-knockout pluripotent cells. Thus, we identify Utx as a novel mediator with distinct functions during the re-establishment of pluripotency and germ cell development. Furthermore, our findings highlight the principle that molecular regulators mediating loss of repressive chromatin during in vivo germ cell reprogramming can be co-opted during in vitro reprogramming towards ground state pluripotency.

Cytotoxic effects produced by arecoline correlated to epigenetic regulation in human K-562 cells.

Epigenetic effects are considered heritable but may also be modified by environmental factors. Arecoline (ARC), a major component of areca nut alkaloids, is an important environmental risk factor for oral cancer and hepatocellular carcinomain Taiwan. The aim of this study was to determine the influence of ARC on the epigenome. The mRNA expression of histone methyltransferases, acetyltransferases, and demethylases were assessed in K-562 cells following exposure to ARC. Results demonstrated that ARC produced changes in the expressions of several genes catalyzing histone methylation (Mll, Setdb1, and Suv39h2), acetylation (Atf2), and demethylation (JMJD6). Since H3K9 methylation is involved in maintaining the stability of heterochromatin structures and inactivating euchromatic gene expressions, data suggest that the ARC-induced epigenetic changes play a role in the mechanisms underlying chemical-mediated cytotoxicity and genotoxicity.