Affect DNA Methylation Research Articles

Micronutrient supplementation affects DNA methylation in male gonads with potential intergenerational epigenetic inheritance involving the embryonic development through glutamate receptor-associated genes

BackgroundDNA methylation has an important role in intergenerational inheritance. An increasing number of studies have reported evidence of germline inheritance of DNA methylation induced by nutritional signals in mammals. Vitamins and minerals as micronutrients contribute to growth performance in vertebrates, including Atlantic salmon (Salmo salar), and also have a role in epigenetics as environmental factors that alter DNA methylation status. It is important to understand whether micronutrients in the paternal diet can influence the offspring through alterations of DNA methylation signatures in male germ cells.ResultsHere, we show the effect of micronutrient supplementation on DNA methylation profiles in the male gonad through a whole life cycle feeding trial of Atlantic salmon fed three graded levels of micronutrient components. Our results strongly indicate that micronutrient supplementation affects the DNA methylation status of genes associated with cell signalling, synaptic signalling, and embryonic development. In particular, it substantially affects DNA methylation status in the promoter region of a glutamate receptor gene, glutamate receptor ionotropic, NMDA 3A-like (grin3a-like), when the fish are fed both medium and high doses of micronutrients. Furthermore, two transcription factors, histone deacetylase 2 (hdac2) and a zinc finger protein, bind to the hyper-methylated site in the grin3a-like promoter. An estimated function of hdac2 together with a zinc finger indicates that grin3a-like has a potential role in intergenerational epigenetic inheritance and the regulation of embryonic development affected by paternal diet.ConclusionsThe present study demonstrates alterations of gene expression patterns and DNA methylation signatures in the male gonad when Atlantic salmon are fed different levels of micronutrients. Alterations of gene expression patterns are of great interest because the gonads are supposed to have limited metabolic activities compared to other organs, whereas alterations of DNA methylation signatures are of great importance in the field of nutritional epigenetics because the signatures affected by nutrition could be transferred to the next generation. We provide extensive data resources for future work in the context of potential intergenerational inheritance through the male germline.

Molecular mechanisms of antipsychotics - their influence on intracellular signaling pathways, and epigenetic and post-transcription processes.

The purpose of this paper is to describe some aspects of the intra-cellular mechanism of action of neuroleptics, drugs widely used in psychiatry in treatment of psychotic and affective disorders. The ability of neuroleptics to influence and modify the metabolic, energetic and structural processes of neurons, as well as their apoptosis, probably influence their therapeutic potential. The direct and indirect mechanisms of antipsychotics are discussed on the basis of epigenetic, intra-cellular and post-transcription processes. Antipsychotic drugs facilitate chromatin remodeling, decreasing or increasing histone acetylation, and affect DNA methylation differently. Antipsychotics modulate the intracellular signaling cascades like the cyclic adenosine monophosphate (cAMP), AKT/glycogen synthase kinase-3 (GSK-3) pathway and mitogen-activated protein kinase (MAPK) in a variety of ways that contribute to their different clinical and side-effect profiles. Among the cellular processes involved in the activity of antipsychotics are energy and metabolism, protein synthesis and processing, cytoskeleton functions like microtubule dynamics, dendritic branching, and spine dynamics, as well as cell adhesion and synaptic activity. Finally, antipsychotics have the ability to modulate the expression of a large number of miRNAs, which is related to oxidative stress and metabolism. Despite the efficacy of antipsychotics in treating schizophrenia and bipolar disorders over the last several decades, their molecular mechanisms of action turn out to be very complex and have not yet been fully elucidated. Recent thinking about a more personalized and endophenotype-specific diagnosing and treatment requires a more advanced genomic and proteomic approach and seems to be the next step in the treatment of mental disorders.

DNA methylation diversity analysis of Lycium barbarum samples from different cultivation areas based on MSAP

Obvious epigenetic differentiation occurred on Lycium barbarum in different cultivation areas in China. To investigate the difference and change rule of DNA methylation level and pattern of L. barbarum from different cultivation areas in China, the present study employed fluorescence-assisted methylation-sensitive amplified polymorphism(MSAP) to analyze the methylation level and polymorphism of 53 genomic DNA samples from Yinchuan Plain in Ningxia, Bayannur city in Inner Mongolia, Jingyuan county and Yumen city in Gansu, Delingha city in Qinghai, and Jinghe county in Xinjiang. The MSAP technical system suitable for the methylation analysis of L. barbarum genomic DNA was established and ten pairs of selective primers were selected. Among amplified 5'-CCGG-3' methylated sites, there were 35.85% full-methylated sites and 39.88% hemi-methylated sites, showing a high degree of epigenetic differentiation. Stoichiometric analysis showed that the ecological environment was the main factor affecting the epigenetic characteristics of L. barbarum, followed by cultivated varieties. Precipitation, air temperature, and soil pH were the main ecological factors affecting DNA methylation in different areas. This study provided a theoretical basis for the analysis of the epigenetic mechanism of L. barbarum to adapt to the diffe-rent ecological environments and research ideas for the introduction, cultivation, and germplasm traceability of L. barbarum.

Chromium induced neurotoxicity by altering metabolism in zebrafish larvae

BackgroundRecently, both trivalent chromium Cr (III) and hexavalent chromium Cr (VI) have been reported to produce neurotoxicity. However, the underlying mechanisms of the neurotoxicity caused by different chemical valence of chromium remain unclear. ObjectiveThe purpose of this study was to investigate the mechanism of neurotoxicity induced by exposure to chromium with different valence states based on metabolic disturbance in zebrafish larvae. MethodsZebrafish embryos were exposed to 1 mg/L Cr (III) and 1 mg/L Cr (VI) for 120 hpf respectively. The related indexes of neural development were observed by stereoscope and behavior analysis system. 8OH-dG were detected using enzyme-linked immunosorbent assay. The generation of reactive oxygen species was detected using an oxidant-sensing probe 2',7'-dichlorodihydrofluorescein diacetate. AChE activity was determined by a colorimetric assay based on hydrolysis of acetylcholine. The expression levels of neurodevelopmental genes and methyltransferase genes in juvenile zebrafish was analyzed by real-time PCR. The methylation status of neurogenin1 and neurod1 genes was detected by bisulfite sequencing PCR. The binding of H3K27me3 was detected by chromatin immunoprecipitation-qPCR. Metabolic profiles and one carbon metabolic analysis were performed by UPLC-MS. ResultsThere were no significant differences in survival rate, hatching rate and spontaneous movement of zebrafish in both Cr-exposed groups compared to the control. The malformation rate in Cr (VI) -exposed group was obviously increased compared to the control and Cr (III) -exposed group. At 48hpf and 72hpf of exposure, the embryonic heart rate in Cr (III)-exposed group was significantly higher than that of Cr (VI)-exposed group and the control. At 120hpf, zebrafish in both Cr-exposed groups exhibited decreasing changes in swimming distance and disturbance of sensitivity to light and dark. 8OH-dG in Cr (VI)-exposed group were significantly higher than that in the control. The generation of ROS in both Cr -exposed groups was significantly higher than that in the control. The activity of AchE was significantly decreased in both Cr-exposed groups compared to the control. Most of early neurogenesis related genes, such as α-tubulin, elavl3, gap43, sox19b, neurogenin1 and neurod1 in Cr-exposed groups were significantly up-regulated compared to those in the control. The expression of dnmt1 and dnmt3 genes was significantly down-regulated in both Cr-exposed groups. BSP-PCR results showed that genic sequences in the neurogenin1 and neurod1 genes have lower levels of DNA methylation in both Cr-exposed groups, especial in Cr (VI)-exposed group. ChIP analysis showed that there was a decrease in H3K27me3 binding within the corresponding region of neurogenin1 in both Cr-exposed groups and that of neurod1 in Cr (III)-exposed group. Untargeted metabolomic analysis showed that significant changes in metabolites induced by Cr exposure were associated with differences in primary bile acid biosynthesis, phospholipid biosynthesis (phosphatidylcholine biosynthesis and phosphatidylethanolamine biosynthesis), linoleic acid metabolism, arachidonic acid metabolism, amino acid metabolism, purine metabolism, betaine metabolism, spermidine and spermine biosynthesis, and folate metabolism, the last four of which are related to one carbon metabolism. Targeted analysis of one carbon metabolites (5-MT, Gly, Met, SAH and Hcy) related with folate cycle and methionine metabolism were significantly decreased upon Cr exposure. The elevated SAM to SAH ratio in both Cr- exposed group indicated the decreasing capacity for methylation reaction. ConclusionCr (III) and Cr (VI) can induce neurotoxicity by interfering with one carbon metabolism and affecting DNA methylation and histone methylation to regulate the expression of neuro-related genes. Cr exposure also influenced primary bile acid biosynthesis and phospholipid biosynthesis, which are associated with neuroprotective effects and need to be further validated.

Embryo culture media differentially alter DNA methylating enzymes and global DNA methylation in embryos and oocytes.

The effects of culture media on DNA methylation process, which is one of the epigenetic mechanisms, have not been clearly elucidated although it is known that in vitro culture conditions alter epigenetic mechanisms. This study was designed to address the question: does embryo culture media approach, sequential or single step, differentially affect DNA methylating enzymes and global DNA methylation. Mouse zygotes were cultured either in single step or sequential culture media until the blastocyst stage and in vivo developed blastocyst were utilized as control. Similarly, GV stage oocytes were in vitro matured either in single step or first step of sequential culture media. In vivo matured MII oocytes were used as control. The expression levels and cellular localization of Dnmt1 and 3a enzymes were analyzed by immunofluorescence and western blot analysis while global DNA methylation was evaluated by immunofluorescence. We found that signal intensities of Dnmt1 and Dnmt3a enzymes were significantly low in embryos or oocytes cultured in sequential media compared to single step media and control, which were comparable amongst themself. Similarly, global DNA methylation level in single step media and control groups was comparable but both was higher than the sequential media. This study demonstrated that composition of culture media may differentially affect DNA methylation levels in mouse embryos and oocytes. Since abnormal DNA methylation may cause aberrant oocyte or embryo development, we think that further studies are needed to test human embryos and oocyte, and to explain molecular mechanisms.

Association of exposure to deoxynivalenol with DNA methylation in white blood cells in children in China

Deoxynivalenol (DON) is a mycotoxin that commonly contaminates cereals worldwide. Dietary exposure to DON is a subject of great public health concern, but studies on the health effects of chronic exposure to DON are not available. In this study, we investigated the connection between DNA methylation levels and DON exposure in children. The DNA methylation status of white blood cells from 32 children aged 2~15 years old in Henan, China, was profiled. A total of 378 differentially methylated CpGs were identified between the high and low DON exposure groups, and 8 KEGG pathways were found to be significantly enriched among the differentially methylated genes. In addition, the quantitative methylation of EIF2AK4, EMID2 and GNASAS was analysed using the Sequenom MassARRAY platform. The results showed that the methylation level of EIF2AK4 was significantly different between the two groups, and the methylation levels were associated with exposure to DON. Conclusively, our study found that chronic exposure to DON during childhood could affect DNA methylation levels.

Structural Equation Modeling (SEM) Analysis of Sequence Variation and Green Plant Regeneration via Anther Culture in Barley.

The process of anther culture involves numerous abiotic stresses required for cellular reprogramming, microspore developmental switch, and plant regeneration. These stresses affect DNA methylation patterns, sequence variation, and the number of green plants regenerated. Recently, in barley (Hordeum vulgare L.), mediation analysis linked DNA methylation changes, copper (Cu2+) and silver (Ag+) ion concentrations, sequence variation, β-glucans, green plants, and duration of anther culture (Time). Although several models were used to explain particular aspects of the relationships between these factors, a generalized complex model employing all these types of data was not established. In this study, we combined the previously described partial models into a single complex model using the structural equation modeling approach. Based on the evaluated model, we demonstrated that stress conditions (such as starvation and darkness) influence β-glucans employed by cells for glycolysis and the tricarboxylic acid cycle. Additionally, Cu2+ and Ag+ ions affect DNA methylation and induce sequence variation. Moreover, these ions link DNA methylation with green plants. The structural equation model also showed the role of time in relationships between parameters included in the model and influencing plant regeneration via anther culture. Utilization of structural equation modeling may have both scientific and practical implications, as it demonstrates links between biological phenomena (e.g., culture-induced variation, green plant regeneration and biochemical pathways), and provides opportunities for regulating these phenomena for particular biotechnological purposes.

Misregulation of the expression and activity of DNA methyltransferases in cancer.

In mammals, DNA methyltransferases DNMT1 and DNMT3’s (A, B and L) deposit and maintain DNA methylation in dividing and nondividing cells. Although these enzymes have an unremarkable DNA sequence specificity (CpG), their regional specificity is regulated by interactions with various protein factors, chromatin modifiers, and post-translational modifications of histones. Changes in the DNMT expression or interacting partners affect DNA methylation patterns. Consequently, the acquired gene expression may increase the proliferative potential of cells, often concomitant with loss of cell identity as found in cancer. Aberrant DNA methylation, including hypermethylation and hypomethylation at various genomic regions, therefore, is a hallmark of most cancers. Additionally, somatic mutations in DNMTs that affect catalytic activity were mapped in Acute Myeloid Leukemia cancer cells. Despite being very effective in some cancers, the clinically approved DNMT inhibitors lack specificity, which could result in a wide range of deleterious effects. Elucidating distinct molecular mechanisms of DNMTs will facilitate the discovery of alternative cancer therapeutic targets. This review is focused on: (i) the structure and characteristics of DNMTs, (ii) the prevalence of mutations and abnormal expression of DNMTs in cancer, (iii) factors that mediate their abnormal expression and (iv) the effect of anomalous DNMT-complexes in cancer.

Interactions between the intestinal microbiota and epigenome in individuals with autism spectrum disorder.

Autism spectrum disorder (ASD) is a neurodevelopmental disorder characterized by variable impairment of cognitive function and interpersonal relationships. Furthermore, some individuals with ASD have gastrointestinal disorders that have been correlated with impairments in intestinal microbiota. Gut microbiota are important not only for intestinal health, but also for many other functions including food digestion, energy production, immune system regulation, and, according to current data, behavior. Disruption of the indigenous microbiota, microbial dysbiosis (imbalance between microorganisms present in the gut), overgrowth of potentially pathogenic microorganisms, a less diverse microbiome, or lower levels of beneficial bacteria in children with ASD can affect behavior. Metabolome analysis in children with ASD has identified perturbations in multiple metabolic pathways that might be associated with cognitive functions. Recent studies have shown that the intestinal microbiome provides environmental signals that can modify host response to stimuli by modifying the host epigenome, which affects DNA methylation, histone modification, and non-coding RNAs. The most studied microbiota-produced epigenetic modifiers are short-chain fatty acids, although other products of intestinal microbiota might also cause epigenetic modifications in the host's DNA. Here we review evidence suggesting that epigenetic alterations caused by modification of gene expression play an important role in understanding ASD.

Drought induces variation in the DNA methylation status of the barley HvDME promoter.

Cytosine methylation is an epigenetic modification with essential roles in diverse plant biological processes including vegetative and reproductive development and responsiveness to environmental stimuli. A dynamic process involving DNA methyltransferases and DNA demethylases establishes cytosine DNA methylation levels and distribution along the genome. A DNA demethylase gene from barley (Hordeum vulgare), DEMETER (HvDME), the homologue of the Arabidopsis thaliana DME (AtDME), has been characterized previously and found to respond to drought conditions. Here, the promoter of the HvDME gene was analysed further by in silico and DNA methylation analysis. The effect of drought conditions on the DNA methylation status of HvDME was investigated at single-cytosine resolution using bisulfite sequencing. It was demonstrated that the HvDME promoter can be divided into two discrete regions, in terms of DNA methylation level and density; a relatively unmethylated region proximal to the translational start site that is depleted of non-CG (CHG, CHH) methylation and another distal region, approximately 1500bp upstream of the translational start site, enriched in CG, as well as non-CG methylation. Drought stress provoked alterations in the methylation status of the HvDME promoter distal region, whereas the DNA methylation of the proximal region remained unaffected. Computational analysis of the HvDME promoter revealed the presence of several putative regulatory elements related to drought responsiveness, as well as transposable elements (TEs) that may affect DNA methylation. Overall, our results expand our investigations of the epigenetic regulation of the HvDME gene in response to drought stress in barley and may contribute to further understanding of the epigenetic mechanisms underlying abiotic stress responses in barley and other cereals.

Research progress of risk factors affecting DNA methylation changes and the development of chronic obstructive pulmonary disease

Research progress of risk factors affecting DNA methylation changes and the development of chronic obstructive pulmonary disease

The Effects of Exercise Duration and Intensity on Breast Cancer-Related DNA Methylation: A Randomized Controlled Trial.

Simple SummaryWhile physical activity has been associated with reduced cancer risk, it is not well understood why this is the case. One possible reason is that physical activity affects DNA methylation—a process that functions to turn certain genes “on” or “off”—which can affect cancer-related processes in the body. We tested this in an experimental study, where women aged 30–45 were randomly assigned to complete 16 weeks of exercise of varying intensity and duration. We hypothesized that higher levels of exercise would lead to changes in DNA methylation that would be associated with reduced cancer risk. Contrary to our hypotheses, we found that the total amount of exercise completed was not associated with changes in DNA methylation, though we did find that increases in VO2max, a marker of physical fitness, were associated with decreases in methylation of the BRCA1 gene, and higher levels of exercise during a follow-up period were associated with lower levels of methylation of the GALNT9 gene. This study provides preliminary evidence that increased exercise behavior or fitness may affect the methylation of some genes that are related to breast cancer.Emerging research suggests that one mechanism through which physical activity may decrease cancer risk is through its influence on the methylation of genes associated with cancer. The purpose of the current study was to prospectively test, using a rigorous experimental design, whether aerobic exercise affects DNA methylation in genes associated with breast cancer, as well as whether quantity of exercise completed affects change in DNA methylation in a dose–response manner. 276 women (M age = 37.25, SD = 4.64) were recruited from the Denver metro area for a randomized controlled trial in which participants were assigned to a supervised aerobic exercise program varying in a fully crossed design by intensity (55–65% versus 75–85% of VO2max) and duration (40 versus 20 min per session). DNA methylation was assessed via blood samples provided at baseline, after completing a 16-week supervised exercise intervention, and six months after the intervention. 137 participants completed the intervention, and 81 had viable pre-post methylation data. Contrary to our hypotheses, total exercise volume completed in kcal/kg/week was not associated with methylation from baseline to post-intervention for any of the genes of interest. An increase in VO2max over the course of the intervention, however, was associated with decreased post-intervention methylation of BRCA1, p = 0.01. Higher levels of self-reported exercise during the follow-up period were associated with lower levels of GALNT9 methylation at the six-month follow-up. This study provides hypothesis-generating evidence that increased exercise behavior and or increased fitness might affect methylation of some genes associated with breast cancer to reduce risk.

Association Between Periconceptional Weight of Maternal Grandmothers and Attention-Deficit/Hyperactivity Disorder in Grandchildren

Neurodevelopmental disorders have been proposed to involve alterations to epigenetic regulation, and epigenetic effects may extend to germline cells to affect later generations. Weight status may affect DNA methylation, and maternal weight before and during pregnancy has been associated with offspring DNA methylation as well as attention-deficit/hyperactivity disorder (ADHD). To assess whether a woman's weight before and during pregnancy is associated with ADHD in her grandchild. This cohort study analyzed data from 19 835 grandmother-mother dyads and 44 720 grandchildren in the Nurses' Health Study II (NHS-II) cohort (2001-2013), a population-based prospective cohort study. Cluster-weighted generalized estimating equations were modeled to estimate the association of grandmother's prepregnancy body mass index (BMI) and gestational weight gain with grandchild risk of ADHD. Data analyses were conducted from May 2018 to April 2021. Grandmothers reported their height and weight before, and weight gain during, their pregnancy with the NHS-II participants. Mothers self-reported height and weight prior to pregnancy. From those data, grandmother BMI and mother BMI were calculated as weight in kilograms divided by height in meters squared and categorized as underweight (<18.5), healthy/normal (18.5-24.9), overweight (25.0-29.9), or obese (≥30). Cases of ADHD identified by maternal report of having a child with a diagnosis of ADHD. In total, 19 835 grandmothers (97.6% White race/ethnicity; 2113 [10.7%] prepregnancy underweight and 1391 [7.0%] prepregnancy overweight or obese) were included in this cohort study. Of 44 720 grandchildren, 3593 (8%) received a diagnosis of ADHD. Higher odds of ADHD among grandchildren were found for those whose grandmother was underweight compared with healthy weight prior to pregnancy with the NHS-II participant (adjusted odds ratio, 1.25; 95% CI, 1.10-1.42). By contrast, grandmother gestational weight gain was not significantly associated with risk of grandchild ADHD (adjusted odds ratio for <20 lbs [9.1 kg], 1.06; 95% CI, 0.96-1.16; adjusted odds ratio for >29 lbs [13.2 kg], 1.01; 95% CI, 0.91-1.13). Mother prepregnancy BMI showed an association with ADHD among offspring, with a stronger association detected for obese status (adjusted odds ratio, 1.27; 95% CI, 1.07-1.49) than for overweight status (adjusted odds ratio, 1.13; 95% CI, 1.02-1.26) compared with normal weight as a reference group. The positive association between grandmother prepregnancy underweight and ADHD risk among the grandchildren remained unchanged after further adjustment for potential mediators, including maternal prepregnancy BMI. The results of this cohort study indicate that grandmother underweight prior to pregnancy is associated with an increased risk of ADHD among grandchildren, independent of grandmother gestational weight gain and independent of maternal prepregnancy weight status.

Polyphenol-Rich Aronia melanocarpa Juice Consumption Affects LINE-1 DNA Methylation in Peripheral Blood Leukocytes in Dyslipidemic Women.

Cardiovascular disease (CVD) is associated with alterations in DNA methylation and polyunsaturated fatty acid (PUFA) profile, both modulated by dietary polyphenols. The present parallel, placebo-controlled study (part of the original clinical study registered as NCT02800967 at www.clinicaltrials.gov) aimed to determine the impact of 4-week daily consumption of polyphenol-rich Aronia melanocarpa juice (AMJ) treatment on Long Interspersed Nucleotide Element-1 (LINE-1) methylation in peripheral blood leukocytes and on plasma PUFAs, in subjects (n = 54, age range of 40.2 ± 6.7 years) at moderate CVD risk, including an increased body mass index, central obesity, high normal blood pressure, and/or dyslipidemia. The goal was also to examine whether factors known to affect DNA methylation (folate intake levels, MTHFR C677T gene variant, anthropometric and metabolic parameters) modulated the LINE-1 methylation levels upon the consumption of polyphenol-rich aronia juice. Experimental analysis of LINE-1 methylation was done by MethyLight method. MTHFR C677T genotypes were determined by the polymerase chain reaction–restriction fragment length polymorphism method, and folate intake was assessed by processing the data from the food frequency questionnaire. PUFAs were measured by gas–liquid chromatography, and serum lipid profile was determined by using Roche Diagnostics kits. The statistical analyses were performed using Statistica software package. In the comparison after vs. before the treatment period, in dyslipidemic women (n = 22), we observed significant decreases in LINE-1 methylation levels (97.54 ± 1.50 vs. 98.39 ± 0.86%, respectively; P = 0.01) and arachidonic acid/eicosapentaenoic acid ratio [29.17 ± 15.21 vs. 38.42 (25.96–89.58), respectively; P = 0.02]. The change (after vs. before treatment) in LINE-1 methylation directly correlated with the presence of MTHFR 677T allele, average daily folate intake, and the change in serum low-density lipoprotein cholesterol but inversely correlated with the change in serum triacylglycerols (R = 0.72, R2 = 0.52, adjusted R2 = 0.36, P = 0.03). The current results imply potential cardioprotective effects of habitual polyphenol-rich aronia juice consumption achieved through the modifications of DNA methylation pattern and PUFAs in subjects at CVD risk, which should be further confirmed. Hence, the precision nutrition-driven modulations of both DNA methylation and PUFA profile may become targets for new approaches in the prevention of CVD.

Dietary Sodium Restriction Results in Tissue-Specific Changes in DNA Methylation in Humans.

[Figure: see text].

HeLa TI cell-based assay as a new approach to screen for chemicals able to reactivate the expression of epigenetically silenced genes.

Chemicals reactivating epigenetically silenced genes target diverse classes of enzymes, including DNMTs, HDACs, HMTs and BET protein family members. They can strongly influence the expression of genes and endogenous retroviral elements with concomitant dsRNA synthesis and massive transcription of LTRs. Chemicals reactivating gene expression may cause both beneficial effects in cancer cells and may be hazardous by promoting carcinogenesis. Among chemicals used in medicine and commerce, only a small fraction has been studied with respect to their influence on epigenetic silencing. Screening of chemicals reactivating silent genes requires adequate systems mimicking whole-genome processes. We used a HeLa TSA-inducible cell population (HeLa TI cells) obtained by retroviral infection of a GFP-containing vector followed by several rounds of cell sorting for screening purposes. Previously, the details of GFP epigenetic silencing in HeLa TI cells were thoroughly described. Herein, we show that the epigenetically repressed gene GFP is reactivated by 15 agents, including HDAC inhibitors–vorinostat, sodium butyrate, valproic acid, depsipeptide, pomiferin, and entinostat; DNMT inhibitors–decitabine, 5-azacytidine, RG108; HMT inhibitors–UNC0638, BIX01294, DZNep; a chromatin remodeler–curaxin CBL0137; and BET inhibitors–JQ-1 and JQ-35. We demonstrate that combinations of epigenetic modulators caused a significant increase in cell number with reactivated GFP compared to the individual effects of each agent. HeLa TI cells are competent to metabolize xenobiotics and possess constitutively expressed and inducible cytochrome P450 mono-oxygenases involved in xenobiotic biotransformation. Thus, HeLa TI cells may be used as an adequate test system for the extensive screening of chemicals, including those that must be metabolically activated. Studying the additional metabolic activation of xenobiotics, we surprisingly found that the rat liver S9 fraction, which has been widely used for xenobiotic activation in genotoxicity tests, reactivated epigenetically silenced genes. Applying the HeLa TI system, we show that N-nitrosodiphenylamine and N-nitrosodimethylamine reactivate epigenetically silenced genes, probably by affecting DNA methylation.

Does Arsenic Contamination Affect DNA Methylation Patterns in a Wild Bird Population? An Experimental Approach.

Pollutants, such as toxic metals, negatively influence organismal health and performance, even leading to population collapses. Studies in model organisms have shown that epigenetic marks, such as DNA methylation, can be modulated by various environmental factors, including pollutants, influencing gene expression, and various organismal traits. Yet experimental data on the effects of pollution on DNA methylation from wild animal populations are largely lacking. We here experimentally investigated for the first time the effects of early-life exposure to environmentally relevant levels of a key pollutant, arsenic (As), on genome-wide DNA methylation in a wild bird population. We experimentally exposed nestlings of great tits (Parus major) to arsenic during their postnatal developmental period (3 to 14 days post-hatching) and compared their erythrocyte DNA methylation levels to those of respective controls. In contrast to predictions, we found no overall hypomethylation in the arsenic group. We found evidence for loci to be differentially methylated between the treatment groups, but for five CpG sites only. Three of the sites were located in gene bodies of zinc finger and BTB domain containing 47 (ZBTB47), HIVEP zinc finger 3 (HIVEP3), and insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1). Further studies are needed to evaluate whether epigenetic dysregulation is a commonly observed phenomenon in polluted populations and what are the consequences for organism functioning and for population dynamics.

Heat stress and immune response phenotype affect DNA methylation in blood mononuclear cells from Holstein dairy cows

Heat stress negatively affects health and production in cows. Examining the cellular response to heat stress could reveal underlying protective molecular mechanisms associated with superior resilience and ultimately enable selection for more resilient cattle. This type of investigation is increasingly important as future predictions for the patterns of heat waves point to increases in frequency, severity, and duration. Cows identified as high immune responders based on High Immune Response technology (HIR) have lower disease occurrence compared to their average and low immune responder herd-mates. In this study, our goal was to identify epigenetic differences between high and low immune responder cows in response to heat stress. We examined genome-wide DNA methylation of blood mononuclear cells (BMCs) isolated from high and low cows, before and after in vitro heat stress. We identified differential methylation of promoter regions associated with a variety of biological processes including immune function, stress response, apoptosis, and cell signalling. The specific differentially methylated promoter regions differed between samples from high and low cows, and results revealed pathways associated with cellular protection during heat stress.

Salt stress and salt shock differently affect DNA methylation in salt-responsive genes in sugar beet and its wild, halophytic ancestor.

Here we determined the impact of salt shock and salt stress on the level of DNA methylation in selected CpG islands localized in promoters or first exons of sixteen salt-responsive genes in beets. Two subspecies differing in salt tolerance were subjected for analysis, a moderately salt-tolerant sugar beet Beta vulgaris ssp. vulgaris cv. Huzar and a halophytic beet, Beta vulgaris ssp. maritima. The CpG island methylation status was determined. All target sequences were hyper- or hypomethylated under salt shock and/or salt stress in one or both beet subspecies. It was revealed that the genomic regions analyzed were highly methylated in both, the salt treated plants and untreated controls. Methylation of the target sequences changed in a salt-dependent manner, being affected by either one or both treatments. Under both shock and stress, the hypomethylation was a predominant response in sugar beet. In Beta vulgaris ssp. maritima, the hypermethylation occurred with higher frequency than hypomethylation, especially under salt stress and in the promoter-located CpG sites. Conversely, the hypomethylation of the promoter-located CpG sites predominated in sugar beet plants subjected to salt stress. This findings suggest that DNA methylation may be involved in salt-tolerance and transcriptomic response to salinity in beets.

Combined healthy lifestyle score and risk of epigenetic aging: a discordant monozygotic twin study.

We investigated whether lifestyle influences epigenetic aging in 143 monozygotic twin pairs discordant for the combined healthy lifestyle score. Twins were scored for four lifestyle factors as unhealthy or healthy; non-smoker, moderate drinker, adequate fruit and vegetable intake, and sufficient physical activity. The combined healthy lifestyle score was calculated for each participant by summing the binary score for each factor. Individual and co-twin analyses were used to assess the relationship between single or combined lifestyle scores, along with DNA methylation age acceleration (AA) calculated using Horvath’s and Li’s epigenetic clocks, focusing on AA and intrinsic epigenetic age acceleration (IEAA) measures. Compared with the twins that scored no or one healthy lifestyle point, those who scored four healthy lifestyle points had lower Li_IEAA with similar results observed in the co-twin analysis. No significant relationships were found in analyses based on Horvath’s clock, although the direction of correlations was consistent with that determined using Li’s clock. Smoking and drinking did not significantly affect DNA methylation AA; however, physical activity and intake of vegetables and fruits did, although the influence varied depending on the epigenetic clock. Our findings suggest that a healthy lifestyle may be an important way to delay aging and prevent age-related diseases.