Lower DNA Methylation Research Articles

Association of BER and NER pathway polymorphism haplotypes and micronucleus frequencies with global DNA methylation in benzene-exposed workers of China: Effects of DNA repair genes polymorphisms on genetic damage

ObjectiveThe base excision repair (BER) pathway and nucleotide excision repair (NER) pathway play important roles in the repair of benzene-induced genetic damage, and the effects of polymorphisms in these pathways on genetic damage and global DNA methylation are of great interest. MethodsTen single nucleotide polymorphisms (SNPs) in the BER (XRCC1: rs25489, rs25487; APE1: rs1130409) and NER pathways (XPA: rs1800975; XPC: rs2228000, rs2228002; XPD: rs13181, rs1799793; XPG: rs17655; ERCC1: rs3212986) were analyzed by a Kompetitive allele-specific PCR (KASP) assay to find associations with cytokinesis-block micronucleus (MN) frequency and global DNA methylation in 294 shoe factory workers and 102 control participants. ResultsWorkers who possessed the following genotypes were associated with high MN frequency: rs25487 AA (FR (95% CI): 1.50 (1.16,1.9), p = 0.002, reference GG); rs1130409 GG (FR (95% CI): 1.28 (1.05,1.55), p = 0.010, reference TT); rs17655 GC (FR (95% CI): 1.18 (1.02,1.38), p = 0.038, reference GG); and rs3212986 TT (FR (95% CI): 1.55 (1.31,1.83), p < 0.001, reference GG). Workers with four and three mutant alleles showed 3.72-fold (OR (95% CI): 3.72 (1.34, 10.03), p = 0.009) and 2.48-fold (OR (95% CI): 2.48 (1.27, 4.88), p = 0.008) increased risk of genetic damage compared with workers with no or one mutant allele, and a dose-response relationship was found by the trend test (p = 0.006). The rs1130409 variant allele (GG+GT) was associated with low global DNA methylation (β=-0.20, 95% CI: −0.42, 0.03, p = 0.045). ConclusionIn benzene-exposed workers, BER and NER pathway polymorphism haplotypes are associated with different levels of chromosome damage and had little effect on global DNA methylation.

DNA Methylation and Gene Expression of Matrix Metalloproteinase 9 Gene in Deficit and Non-deficit Schizophrenia.

The biological pathology of deficit schizophrenia (DS) remains unclear. Matrix metalloproteinase 9 (MMP9) might be associated with neural plasticity and glutamate regulation, involved in schizophrenia pathogenesis. This study explores gene expression and DNA methylation of MMP9 in peripheral blood mononuclear cells (PBMCs) and their relationship with clinical symptoms in DS and non-deficit schizophrenia (NDS). Pyrosequencing was used to determine DNA methylation at CpG sites in exon 4 and exon 5 of MMP9 in 51 DS patients, 53 NDS patients and 50 healthy subjects (HC). RT-qPCR was used to detect MMP9 expression. Clinical symptoms were assessed by BPRS, SANS and SAPS scales. MMP9 expression in PBMCs was significantly higher in DS than NDS and HC subjects. Compared to NDS patients, DS patients had significantly lower DNA methylation at individual CpG sites in exon 4 and exon 5 of MMP9. Correlation analysis showed that DNA methylation in exon 4 was negatively correlated with gene expression in DS group. Positive correlation was found between MMP9 expression and negative symptoms in total schizophrenic patients. The social amotivation factor of SANS and negative syndrome of BPRS was negatively correlated with DNA methylation of CpG5-1 in DS patients but not in NDS patients. DS patients showed a specific abnormality of peripheral MMP9 expression and DNA methylation, indicating a pathological mechanism underlying DS as a specific subgroup of schizophrenia.

Perinatal phthalate and high-fat diet exposure induce sex-specific changes in adipocyte size and DNA methylation

Environmental factors such as diet and endocrine-disrupting chemicals have individually been shown to mediate metabolic function. However, the underlying mechanism by which the combination disrupts adipocyte morphology and fat storage remains unknown. The current study evaluated early-life programming by diet and phthalate exposure. During gestation and lactation, pregnant Long–Evans hooded rat dams were fed either a control (C) or high-fat (HF) diet and were orally administered one of three phthalate dosages (0, 200 or 1000 μg/kg/day), yielding six groups of offspring: C-0, C-200, C-1000, HF-0, HF-200 and HF-1000. On postnatal day (PND) 90, gonadal fat pads were collected and analyzed for histology, gene expression and DNA methylation. Differences in body weight were observed only in males. Hematoxylin and eosin staining revealed larger adipocyte size in HF-0 vs. C-0 females. Exposure to 200 or 1000 μg/kg/day phthalates modulated diet-induced changes in adipose morphology. Compared to C-0 females, HF-0 females also had higher expression of the adipogenesis gene Wnt receptor, frizzled 1 (Fzd1) and the triglyceride cleaving enzyme lipoprotein lipase (Lpl). These increases in gene expression were accompanied by lower DNA methylation surrounding the transcription start sites of the two genes. Diet-driven effects were observed in unexposed females but not in phthalate-treated rats. Results suggest a sex-specific association between perinatal HF diet and body weight, adipocyte size and DNA methylation. Perinatal phthalate exposure appears to produce a phenotype that more closely resembles HF-fed animals.

Large DNA Methylation Canyons Anchor Chromatin Loops Maintaining Hematopoietic Stem Cell Identity

Higher order chromatin structure and DNA methylation are implicated in multiple developmental processes, but their relationship to cell state is unknown. In order to understand how the DNA methylation is connected with nuclear architecture and can vary between cell types and during cell differentiation, we began to explore the 3D architecture of human hematopoietic stem and progenitor cells (HSPCs) by performing in situ Hi-C experiments at 5kb resolution. We found that large (~10kb) DNA methylation canyons can form long loops connecting anchor loci that may be dozens of megabases apart. These canyons also can form interchromosomal links (Fig.1a and 1b). We further confirmed these long-range interactions by performing 3D-FISH using two color fluorescent labeled probes that spanned the HOXA locus loop anchor (green) and the SP8 locus loop anchor (red), which are ~7MB apart (Fig. 1c).In order to begin to investigate mechanisms that may regulate these long loops and how they relate to commonly studied loops that are mediated by CTCF-extrusion, we examined their properties systematically. Interestingly, the anchors of long loops exhibited minimal enrichment for CTCF (1.04-fold), and, even when CTCF was bound, they did not obey the convergent rule. The data suggest these loops are formed by phase separation of the interacting loci to form a genomic subcompartment, rather than by CTCF-mediated extrusion. Next, we sought to determine whether other features correlated with these long loops. By aligning DNA methylation profiles with the Hi-C data, we observed that anchors often corresponded to regions of very low DNA methylation, and thus sought to analyze the relationship in detail. We found that the anchor position of the long loops had lower average DNA methylation levels than standard loop anchors and very often overlapped with DNA methylation canyons. Canyons are typically decorated with either active or repressive histone marks. We considered whether a particular group of canyons was associated with the long loops. Our findings further indicate that repressed regions marked by the polycomb-mediated histone modification H3K27me3 at DNA methylation canyons generally mediate the formation of canyon loops.Next, we considered whether the long loops associated with repressive grand canyons that we had annotated in HSPCs were present in other cell types. Using Aggregate Peak Analysis (APA), a computational strategy in which the Hi-C submatrices from the vicinity of multiple putative loops are superimposed, we examined 19 human cell types and 10 murine cell types in which loop-resolution Hi-C maps are available. Interestingly, unlike previously characterized genomic subcompartments, these long-range loops are only present in stem and progenitor cells, but not in differentiated cell types, such as T cells and erythroid progenitors (Fig. 1d).Further, we identified one particular loop anchor that lay at the anchor of a long loop and contained no apparent genes (“geneless” canyon, or “GLS”). The GLS harboring this anchor is 17 kb long, lies 1.4 Mb upstream of the HOXA1 gene, and forms long loops with a 28 kb grand canyon in the HOXA region. In order to understand the role of the GLS region in hematopoietic stem cells (HSCs), we deleted the GLS in HSPCs using Cas9-mediated editing and assayed the edited cells for their ability to form colonies. Strikingly, after deleting the GLS, the number of colonies and their size was greatly reduced in edited cells compared to control experiments using either random guide RNAs or electroporation only (Fig. 1e). After ex vivo culture, the overwhelming majority of GLS-knock out HSPCs acquired the marker CD38, indicating that they were differentiating. Similarly, HOXA gene expression, an indicator of HSPC function, was greatly diminished after GLS deletion compared to control cells. These data indicate that the GLS identified in our study is functionally associated with maintenance of the HSC state. Overall, our work reveals long-range interactions between H3K27me3-marked DNA methylation canyons comprising a novel microcompartment associated with cellular identity. DisclosuresNo relevant conflicts of interest to declare.

Association of maternal prenatal smoking GFI1-locus and cardio-metabolic phenotypes in 18,212 adults.

BackgroundDNA methylation at the GFI1-locus has been repeatedly associated with exposure to smoking from the foetal period onwards. We explored whether DNA methylation may be a mechanism that links exposure to maternal prenatal smoking with offspring's adult cardio-metabolic health.MethodsWe meta-analysed the association between DNA methylation at GFI1-locus with maternal prenatal smoking, adult own smoking, and cardio-metabolic phenotypes in 22 population-based studies from Europe, Australia, and USA (n = 18,212). DNA methylation at the GFI1-locus was measured in whole-blood. Multivariable regression models were fitted to examine its association with exposure to prenatal and own adult smoking. DNA methylation levels were analysed in relation to body mass index (BMI), waist circumference (WC), fasting glucose (FG), high-density lipoprotein cholesterol (HDL—C), triglycerides (TG), diastolic, and systolic blood pressure (BP).FindingsLower DNA methylation at three out of eight GFI1-CpGs was associated with exposure to maternal prenatal smoking, whereas, all eight CpGs were associated with adult own smoking. Lower DNA methylation at cg14179389, the strongest maternal prenatal smoking locus, was associated with increased WC and BP when adjusted for sex, age, and adult smoking with Bonferroni-corrected P < 0·012. In contrast, lower DNA methylation at cg09935388, the strongest adult own smoking locus, was associated with decreased BMI, WC, and BP (adjusted 1 × 10−7 < P < 0.01). Similarly, lower DNA methylation at cg12876356, cg18316974, cg09662411, and cg18146737 was associated with decreased BMI and WC (5 × 10−8 < P < 0.001). Lower DNA methylation at all the CpGs was consistently associated with higher TG levels.InterpretationEpigenetic changes at the GFI1 were linked to smoking exposure in-utero/in-adulthood and robustly associated with cardio-metabolic risk factors.FundEuropean Union's Horizon 2020 research and innovation programme under grant agreement no. 633595 DynaHEALTH.

Tissue-resident memory T cells are epigenetically cytotoxic with signs of exhaustion in human urinary bladder cancer.

SummaryTissue‐resident memory T (TRM) cells are CD8+ T lymphocytes that reside in the tissues, including tumours. This T cell subset possesses a magnitude of cytotoxicity, but its epigenetic regulation has not been studied. Here, we investigate the impact of perforin DNA methylation in TRM cells and correlate it with their functional potential. Fifty‐three urothelial urinary bladder cancer (UBC) patients were recruited prospectively. The DNA methylation status of the perforin gene (PRF1) locus in TRM cells was investigated by pyrosequencing. Flow cytometry with ViSNE analysis and in‐vitro stimulation were used to evaluate TRM cell phenotypes. We discovered that tumour TRM cells have low DNA methylation in the PRF1 locus (32·9% methylation), which corresponds to increased numbers of perforin‐expressing TRM cells. Surprisingly, programmed cell death 1 (PD‐1) expression is high in tumour TRM cells, suggesting exhaustion. Following interleukin‐15 and T cell receptor stimulation, perforin and T‐bet expressions are enhanced, indicating that TRM cells from tumours are not terminally exhausted. Moreover, a high number of TRM cells infiltrating the tumours corresponds to lower tumour stage in patients. In conclusion, TRM cells from UBC tumours are epigenetically cytotoxic with signs of exhaustion. This finding identifies TRM cells as potential new targets for cancer immunotherapy.

Glia-specific APOE epigenetic changes in the Alzheimer’s disease brain

The apolipoprotein E gene (APOE) is the strongest genetic risk factor for developing Alzheimer’s disease (AD). Our recent identification of altered APOE DNA methylation in AD postmortem brain (PMB) prompted this follow-up study. Our goals were to (i) validate the AD-differential methylation of APOE in an independent PMB study cohort and (ii) determine the cellular populations (i.e., neuronal vs. non-neuronal) of AD PMB that contribute to this differential methylation. Here, we obtained an independent cohort of 57 PMB (42 AD and 15 controls) and quantified their APOE methylation levels from frontal lobe and cerebellar tissue. We also applied fluorescence-activated nuclei sorting (FANS) to separate neuronal nuclei from non-neuronal nuclei within the tissue of 15 AD and 14 control subjects. Bisulfite pyrosequencing was used to generate DNA methylation profiles of APOE from both bulk PMB and FANS nuclei. Our results provide independent validation that the APOE CGI holds lower DNA methylation levels in AD compared to control in frontal lobe but not cerebellar tissue. Our data also indicate that the non-neuronal cells of the AD brain, which are mainly composed of glia, are the main contributors to the lower APOE DNA methylation observed in AD PMB. Given that astrocytes are the primary producers of ApoE in the brain our results suggest that alteration of epigenetically regulated APOE expression in glia could be an important part of APOE’s strong effect on AD risk.

An integrated transcriptomic and epigenomic analysis identifies CD44 gene as a potential biomarker for weight loss within an energy-restricted program.

The interindividual variable response to weight-loss treatments requires the search for new predictive biomarkers for improving the success of weight-loss programs. The aim of this study is to identify novel genes that distinguish individual responses to a weight-loss dietary treatment by using the integrative analysis of mRNA expression and DNA methylation arrays. Subjects from Metabolic Syndrome Reduction in Navarra (RESMENA) project were classified as low (LR) or high (HR) responders depending on their weight loss. Transcriptomic (n = 24) and epigenomic (n = 47) patterns were determined by array-based genome-wide technologies in human white blood cells at the baseline of the treatment period. CD44 expression was validated by qRT-PCR and methylation degree of CpGs of the gene was validated by MassARRAY® EpiTYPER™ in a subsample of 47 subjects. CD44 protein levels were measured by ELISA in human plasma. Different expression and DNA methylation profiles were identified in LR in comparison to HR. The integrative analysis of both array data identified four genes: CD44, ITPR1, MTSS1 and FBXW5 that were differently methylated and expressed between groups. CD44 showed higher expression and lower DNA methylation levels in LR than in HR. Although differences in CD44 protein levels between LR and HR were not statistically significant, a positive association was observed between CD44 mRNA expression and protein levels. In summary, the combination of a genome-wide methylation and expression array dataset can be a useful strategy to identify novel genes that might be considered as predictors of the dietary response. CD44 gene transcription and methylation may be a possible candidate biomarker for weight-loss prediction.

Central Nervous System Germ Cell Tumors: A Review of the Literature.

Central nervous system germ cell tumors are rare intracranial tumors that mainly occur in pediatrics with substantial variation in the incidence among different regions and genders. Histologically, central nervous system germ cell tumors can be divided into germinomas and nongerminomatous germ cell tumors. The molecular pathology of central nervous system germ cell tumors, particularly germinomas, is mainly based on the presence of isochromosome 12p, gain-of-function of the KIT gene, and a globally low DNA methylation profile. Diagnoses and differential diagnoses are conducted through imaging, tumor marker detection, surgical biopsy, and cerebrospinal fluid cytology. Germinomas are often treated via whole-ventricular radiotherapy or neoadjuvant chemotherapy combined with reduced-dose whole-ventricular radiotherapy, whereas nongerminomatous germ cell tumors are mainly treated with chemotherapy, surgical resection, and radiotherapy (individually or in combination), depending on tumor composition. Because the main population of patients is pediatric, extending overall survival and reducing treatment side effects should be the main goals of future studies.

Placental DNA Methylation Adaptation to Maternal Glycemic Response in Pregnancy.

Maternal hyperglycemia during pregnancy is associated with excess fetal growth and adverse perinatal and developmental outcomes. Placental epigenetic maladaptation may underlie these associations. We performed an epigenome-wide association study (>850,000 CpG sites) of term placentas and prenatal maternal glycemic response 2-h post oral glucose challenge at 24-30 weeks of gestation among 448 mother-infant pairs. Maternal 2-h glycemia postload was strongly associated with lower DNA methylation of four CpG sites (false discovery rate [FDR] q <0.05) within the phosphodiesterase 4B gene (PDE4B). Additionally, three other individual CpG sites were differentially methylated relative to maternal glucose response within the TNFRSF1B, LDLR, and BLM genes (FDR q <0.05). DNA methylation correlated with expression of its respective genes in placental tissue at three out of four independent identified loci: PDE4B (r = 0.31, P < 0.01), TNFRSF1B (r = -0.24, P = 0.013), and LDLR (r = 0.32, P < 0.001). In an independent replication cohort (N = 65-108 samples), results were consistent in direction but not significantly replicated among tested CpG sites in PDE4B and TNFRSF1B Our study provides evidence that maternal glycemic response during pregnancy is associated with placental DNA methylation of key inflammatory genes whose expression levels are partially under epigenetic control.

Human cis-acting elements regulating escape from X-chromosome inactivation function in mouse.

A long-standing question concerning X-chromosome inactivation (XCI) has been how some genes avoid the otherwise stable chromosome-wide heterochromatinization of the inactive X chromosome. As 20% or more of human X-linked genes escape from inactivation, such genes are an important contributor to sex differences in gene expression. Although both human and mouse have genes that escape from XCI, more genes escape in humans than mice, with human escape genes often clustering in larger domains than the single escape genes of mouse. Mouse models offer a well-characterized and readily manipulated system in which to study XCI, but given the differences in genes that escape it is unclear whether the mechanism of escape gene regulation is conserved. To address conservation of the process and the potential to identify elements by modelling human escape gene regulation using mouse, we integrated a human and a mouse BAC each containing an escape gene and flanking subject genes at the mouse X-linked Hprt gene. Escape-level expression and corresponding low promoter DNA methylation of human genes RPS4X and CITED1 demonstrated that the mouse system is capable of recognizing human elements and therefore can be used as a model for further refinement of critical elements necessary for escape from XCI in humans.

Glucocorticoid receptor gene methylation moderates the association of childhood trauma and cortisol stress reactivity

Exposure to childhood trauma (CT) has been linked to sustained dysregulations of major stress response systems, including findings of both exaggerated and attenuated hypothalamus–pituitary–adrenal (HPA) axis activity. Likewise, CT constitutes a common risk factor for a broad range of psychiatric conditions that involve distinct neuroendocrine profiles. In this study, we investigated the role of epigenetic variability in a stress-related gene as a potential mediator or moderator of such differential trajectories in CT survivors. For this, we screened adult volunteers for CT and recruited a healthy sample of 98 exposed (67 with mild-moderate, 31 with moderate-severe exposure) and 102 control individuals, with an equal number of males and females in each group. DNA methylation (DNAM) levels of the glucocorticoid receptor exon 1F promoter (NR3C1-1F) at functionally relevant sites were analyzed via bisulfite pyrosequencing from whole blood samples. Participants were exposed to a laboratory stressor (Trier Social Stress Test) to assess salivary cortisol stress responses. The major finding of this study indicates that DNAM in a biologically relevant region of NR3C1-1F moderates the specific direction of HPA-axis dysregulation (hypo- vs. hyperreactivity) in adults exposed to moderate-severe CT. Those trauma survivors with increased NR3C1-1F DNAM displayed, on average, 10.4 nmol/l (62.3%) higher peak cortisol levels in response to the TSST compared to those with low DNAM. In contrast, unexposed and mildly-moderately exposed individuals displayed moderately sized cortisol stress responses irrespective of NR3C1-1F DNAM. Contrary to some prior work, however, our data provides no evidence for a direct association of CT and NR3C1-1F DNAM status. According to this study, epigenetic changes of NR3C1-1F may provide a more in-depth understanding of the highly variable neuroendocrine and pathological sequelae of CT.

WITHDRAWN: DNA methylation and gene expression of matrix metalloproteinase-9 gene in Deficit and Non-Deficit Schizophrenia

// Ju Gao 1, 2, * , Hongwei Yi 3, * , Xiaowei Tang 4, * , Xiaotang Feng 5 , Miao Yu 1 , Weiwei Sha 4 , Xiang Wang 6 , Xiaobin Zhang 4 and Xiangrong Zhang 1, 7 1 Department of Geriatric Psychiatry, Nanjing Brain Hospital, Affiliated to Nanjing Medical University, Nanjing, Jiangsu 210029, China 2 Centers of Disease Prevention and Control for Mental Disorders, Shanghai Changning Mental Health Center, Shanghai 200335, China 3 Department of Pharmacology, School of Medicine, Southeast University, Nanjing, Jiangsu 210009, China 4 Department of Psychiatry, Affiliated WuTaiShan Hospital of Medical College of Yangzhou University, Yangzhou, Jiangsu 225003, China 5 Department of Psychiatry, Nanjing Qing Long Mountain Psychiatric Hospital, Nanjing, Jiangsu 211123, China 6 Medical Psychological Institute of the Second Xiangya Hospital, Central South University, Changsha, Hunan 410011, China 7 Department of Neurology, Affiliated ZhongDa Hospital, Neuropsychiatric Institute and Medical School of Southeast University, Nanjing, Jiangsu 210009, China * These authors contributed equally to this work Correspondence to: Xiangrong Zhang, email: drxrz@hotmail.com Keywords: deficit schizophrenia; matrix metalloproteinase-9; DNA methylation; gene expression; negative symptoms Received: July 31, 2017 Accepted: December 05, 2017 Published: January 02, 2018 ABSTRACT Background: The biological pathology of deficit schizophrenia (DS) remains unclear. Matrix metalloproteinase-9 (MMP-9) might be associated with neural plasticity and glutamate regulation, involved in schizophrenia pathogenesis. This study explored gene expression and DNA methylation of MMP-9 in peripheral blood mononuclear cells (PBMCs) and their relationship with clinical symptoms in DS and non-deficit schizophrenia (NDS). Materials And Methods: Pyrosequencing was used to determine DNA methylation at CpG sites in exon 4 and exon 5 of MMP-9 gene in 51 DS patients, 53 NDS patients and 50 healthy subjects (HC). RT-qPCR was used to detect MMP-9 gene expression. Clinical symptoms were assessed by BPRS, SANS and SAPS scales. Results: MMP-9 gene expression in PBMCs was significantly higher in DS than NDS and HC subjects. Compared to NDS patients, DS patients had significantly lower DNA methylation at individual CpG sites in exon 4 and exon 5 of MMP-9 gene. Correlation analysis showed that DNA methylation in exon 4 was negatively correlated with gene expression in DS group. Positive correlation was found between MMP-9 gene expression and negative symptoms in total schizophrenic patients. The social amotivation factor of SANS and negative syndrome of BPRS was negatively correlated with DNA methylation of CpG5-1 in DS patients but not in NDS patients. Conclusions: DS patients showed a specific abnormality of peripheral MMP-9 gene expression and DNA methylation, indicating a pathological mechanism underlying DS as a specific subgroup of schizophrenia.

Maternal low glycemic index dietary intervention and DNA methylation in 5 year old offspring: results from the ROLO Kids Study

Maternal low glycemic index dietary intervention and DNA methylation in 5 year old offspring: results from the ROLO Kids Study

DNA methylation regulates discrimination of enhancers from promoters through a H3K4me1-H3K4me3 seesaw mechanism

BackgroundDNA methylation at promoters is largely correlated with inhibition of gene expression. However, the role of DNA methylation at enhancers is not fully understood, although a crosstalk with chromatin marks is expected. Actually, there exist contradictory reports about positive and negative correlations between DNA methylation and H3K4me1, a chromatin hallmark of enhancers.ResultsWe investigated the relationship between DNA methylation and active chromatin marks through genome-wide correlations, and found anti-correlation between H3K4me1 and H3K4me3 enrichment at low and intermediate DNA methylation loci. We hypothesized “seesaw” dynamics between H3K4me1 and H3K4me3 in the low and intermediate DNA methylation range, in which DNA methylation discriminates between enhancers and promoters, marked by H3K4me1 and H3K4me3, respectively. Low methylated regions are H3K4me3 enriched, while those with intermediate DNA methylation levels are progressively H3K4me1 enriched. Additionally, the enrichment of H3K27ac, distinguishing active from primed enhancers, follows a plateau in the lower range of the intermediate DNA methylation level, corresponding to active enhancers, and decreases linearly in the higher range of the intermediate DNA methylation. Thus, the decrease of the DNA methylation switches smoothly the state of the enhancers from a primed to an active state. We summarize these observations into a rule of thumb of one-out-of-three methylation marks: “In each genomic region only one out of these three methylation marks {DNA methylation, H3K4me1, H3K4me3} is high. If it is the DNA methylation, the region is inactive. If it is H3K4me1, the region is an enhancer, and if it is H3K4me3, the region is a promoter”. To test our model, we used available genome-wide datasets of H3K4 methyltransferases knockouts. Our analysis suggests that CXXC proteins, as readers of non-methylated CpGs would regulate the “seesaw” mechanism that focuses H3K4me3 to unmethylated sites, while being repulsed from H3K4me1 decorated enhancers and CpG island shores.ConclusionsOur results show that DNA methylation discriminates promoters from enhancers through H3K4me1-H3K4me3 seesaw mechanism, and suggest its possible function in the inheritance of chromatin marks after cell division. Our analyses suggest aberrant formation of promoter-like regions and ectopic transcription of hypomethylated regions of DNA. Such mechanism process can have important implications in biological process in where it has been reported abnormal DNA methylation status such as cancer and aging.

DNA methylation and gene expression of TXNIP in adult offspring of women with diabetes in pregnancy.

BackgroundFetal exposure to maternal diabetes increases the risk of type 2 diabetes (T2DM), possibly mediated by epigenetic mechanisms. Low blood TXNIP DNA methylation has been associated with elevated glucose levels and risk of T2DM, and increased skeletal muscle TXNIP gene expression was reported in subjects with impaired glucose metabolism or T2DM. Subcutaneous adipose tissue (SAT) and skeletal muscle play a key role in the control of whole body glucose metabolism and insulin action. The extent to which TXNIP DNA methylation levels are decreased and/or gene expression levels increased in SAT or skeletal muscle of a developmentally programmed at-risk population is unknown.Objective and methodsThe objective of this study was to investigate TXNIP DNA methylation and gene expression in SAT and skeletal muscle, and DNA methylation in blood, from adult offspring of women with gestational diabetes (O-GDM, n = 82) or type 1 diabetes (O-T1DM, n = 67) in pregnancy compared with offspring of women from the background population (O-BP, n = 57).ResultsSAT TXNIP DNA methylation was increased (p = 0.032) and gene expression decreased (p = 0.001) in O-GDM, but these differences were attenuated after adjustment for confounders. Neither blood/muscle TXNIP DNA methylation nor muscle gene expression differed between groups.ConclusionWe found no evidence of decreased TXNIP DNA methylation or increased gene expression in metabolic target tissues of offspring exposed to maternal diabetes. Further studies are needed to confirm and understand the paradoxical SAT TXNIP DNA methylation and gene expression changes in O-GDM subjects.

One-carbon genetic variants and the role of MTHFD1 1958G>A in liver and colon cancer risk according to global DNA methylation.

Several polymorphic gene variants within one-carbon metabolism, an essential pathway for nucleotide synthesis and methylation reactions, are related to cancer risk. An aberrant DNA methylation is a common feature in cancer but whether the link between one-carbon metabolism variants and cancer occurs through an altered DNA methylation is yet unclear. Aims of the study were to evaluate the frequency of one-carbon metabolism gene variants in hepatocellular-carcinoma, cholangiocarcinoma and colon cancer, and their relationship to cancer risk together with global DNA methylation status. Genotyping for BHMT 716A>G, DHFR 19bp ins/del, MTHFD1 1958G>A, MTHFR 677C>T, MTR 2756A>G, MTRR 66A>G, RFC1 80G>A, SHMT1 1420C>T, TCII 776C>G and TS 2rpt-3rpt was performed in 102 cancer patients and 363 cancer-free subjects. Methylcytosine (mCyt) content was measured by LC/MS/MS in peripheral blood mononuclear cells (PBMCs) DNA. The MTHFD1 1958AA genotype was significantly less frequent among cancer patients as compared to controls (p = 0.007) and related to 63% reduction of overall cancer risk (p = 0.003) and 75% of colon cancer risk (p = 0.006). When considering PBMCs mCyt content, carriers of the MTHFD1 1958GG genotype showed a lower DNA methylation as compared to carriers of the A allele (p = 0.048). No differences were highlighted by evaluating a possible relationship between the other polymorphisms analyzed with cancer risk and DNA methylation.The MTHFD1 1958AA genotype is linked to a significantly reduced cancer risk. The 1958GG genotype is associated to PBMCs DNA hypomethylation as compared to the A allele carriership that may exert a protective effect for cancer risk by preserving from DNA hypomethylation.

Regulation of gene transcription in bipolar disorders: Role of DNA methylation in the relationship between prodynorphin and brain derived neurotrophic factor

Bipolar Disorder (BD) is a prevalent and disabling condition, determined by gene-environment interactions, possibly mediated by epigenetic mechanisms. The present study aimed at investigating the transcriptional regulation of BD selected target genes by DNA methylation in peripheral blood mononuclear cells of patients with a DSM-5 diagnosis of type I (BD-I) and type II (BD-II) Bipolar Disorders (n=99), as well as of healthy controls (CT, n=42). The analysis of gene expression revealed prodynorphin (PDYN) mRNA levels significantly reduced in subjects with BD-II but not in those with BD-I, when compared to CT. Other target genes (i.e. catechol-O-methyltransferase (COMT), glutamate decarboxylase (GAD67), serotonin transporter (SERT) mRNA levels remained unaltered. Consistently, an increase in DNA methylation at PDYN gene promoter was observed in BD-II patients vs CT. After stratifying data on the basis of pharmacotherapy, patients on mood-stabilizers (i.e., lithium and anticonvulsants) were found to have lower DNA methylation at PDYN gene promoter. A significantly positive correlation in promoter DNA methylation was observed in all subjects between PDYN and brain derived neurotrophic factor (BDNF), whose methylation status had been previously found altered in BD. Moreover, among key genes relevant for DNA methylation establishment here analysed, an up-regulation of DNA Methyl Transferases 3b (DNMT3b) and of the methyl binding protein MeCP2 (methyl CpG binding protein 2) mRNA levels was also observed again just in BD-II subjects. A clear selective role of DNA methylation involvement in BD-II is shown here, further supporting a role for BDNF and its possible interaction with PDYN. These data might be relevant in the pathophysiology of BD, both in relation to BDNF and for the improvement of available treatments and development of novel ones that modulate epigenetic signatures.

Blood global DNA methylation is decreased in non-severe chronic obstructive pulmonary disease (COPD) patients

BackgroundAlterations in global DNA methylation have been associated with oxidative stress (OS). Since chronic obstructive pulmonary disease (COPD) is characterized by increased oxidative stress we aimed to evaluate the levels of global DNA methylation in this patient group. MethodsWe assessed methylcytosine (mCyt) levels in DNA from blood collected in 43 COPD patients (29 with mild and 14 with moderate disease) and 43 age- and sex-matched healthy controls. ResultsDNA methylation was significantly lower in COPD patients vs. controls (4.20 ± 0.18% mCyt vs. 4.29 ± 0.18% mCyt, p = 0.02). Furthermore, DNA methylation in COPD patients with moderate disease was significantly lower than that in patients with mild disease (4.14 ± 0.15% mCyt vs. 4.23 ± 0.19% mCyt, p < 0.05). Univariate logistic regression analysis showed that lower DNA methylation levels were associated with presence of COPD (crude OR = 0.06, 95% CI 0.00 to 0.67, p = 0.023). This relationship remained significant after adjusting for several confounders (OR 0.03, 95% CI 0.00 to 0.67; p = 0.028). Receiver operating characteristics (ROC) curve analysis demonstrated the area under the curve of mCyt was 0.646, with 46.6% sensitivity and 79.1% specificity for presence of COPD. ConclusionsThere were no significant correlations between methylation and OS indices. The presence and severity of COPD is associated with progressively lower DNA methylation in blood. However, this epigenetic alteration seems independent of oxidative stress.

Genome-wide mapping of transcriptional enhancer candidates using DNA and chromatin features in maize

BackgroundWhile most cells in multicellular organisms carry the same genetic information, in each cell type only a subset of genes is being transcribed. Such differentiation in gene expression depends, for a large part, on the activation and repression of regulatory sequences, including transcriptional enhancers. Transcriptional enhancers can be located tens of kilobases from their target genes, but display characteristic chromatin and DNA features, allowing their identification by genome-wide profiling. Here we show that integration of chromatin characteristics can be applied to predict distal enhancer candidates in Zea mays, thereby providing a basis for a better understanding of gene regulation in this important crop plant.ResultTo predict transcriptional enhancers in the crop plant maize (Zea mays L. ssp. mays), we integrated available genome-wide DNA methylation data with newly generated maps for chromatin accessibility and histone 3 lysine 9 acetylation (H3K9ac) enrichment in young seedling and husk tissue. Approximately 1500 intergenic regions, displaying low DNA methylation, high chromatin accessibility and H3K9ac enrichment, were classified as enhancer candidates. Based on their chromatin profiles, candidate sequences can be classified into four subcategories. Tissue-specificity of enhancer candidates is defined based on the tissues in which they are identified and putative target genes are assigned based on tissue-specific expression patterns of flanking genes.ConclusionsOur method identifies three previously identified distal enhancers in maize, validating the new set of enhancer candidates and enlarging the toolbox for the functional characterization of gene regulation in the highly repetitive maize genome.