Background: Epigenomics has emerged as an essential field in modern molecular biology, providing a critical layer of gene regulation. DNA methylation, histone modifications and alterations to the chromatin accessibility of DNA have been widely associated with complex diseases including cancer. The most recent developments in high-throughput sequencing technology have made it possible to profile epigenetic landscapes genomically on a large scale. However, bulk averaging can obscure cellular heterogeneity essential for understanding complex disease states. The purpose of the review is to survey accessible tools and algorithms to conduct an Epigenomic study in the field of biomedical research, from bulk tissue analysis to the high-resolution frontier of single-cell epigenomics. Methods: We performed a comparative analysis of common methods used to analyze DNA methylation, chromatin immunoprecipitation, sequencing analysis and chromatin accessibility profiling. We described the standardized bioinformatics tools and pipelines required to transform raw sequencing data into mechanistic biological understanding, highlighting the role of quality control, peak calling, and differential analysis. Furthermore, we explore the integration of epigenomics with other “omics” layers through advanced computational frameworks, including machine learning and network-based modeling. Results: These advanced multi-omics techniques demonstrate promising clinical utility by enabling biomarker discovery, disease subtyping, and identification of novel therapeutic targets. Conclusions: Despite challenges with data complexity, the fusion of Artificial Intelligence (AI) and single-cell technologies will accelerate the transition toward precision medicine.
Background/Objectives: Parental environmental factors can shape developmental outcomes through epigenetic mechanisms that regulate gene expression. While maternal dietary effects on offspring epigenetics have been well characterized, the impact of paternal diet on embryonic DNA methylation remains poorly understood. Here, we investigated the effect of paternal methionine supplementation on DNA methylation patterns in preimplantation embryos in Polypay sheep. Methods: Four yearling rams (two control and two methionine-supplemented) were bred to twelve ewes following estrus synchronization and superovulation. Embryos were collected after natural mating and analyzed using whole-genome bisulfite sequencing (WGBS). Results: A total of 842 differentially methylated cytosines (DMCs) were identified in embryos derived from methionine-supplemented sires compared to controls, with 835 hypermethylated and 7 hypomethylated. The majority of DMCs were located in intergenic regions, with minimal representation in exonic regions. To assess overlap between parental dietary effects, DMCs identified in this study were compared with those previously reported in embryos derived from methionine-supplemented dams. Eight hypermethylated DMCs were shared between the two datasets, while no hypomethylated DMCs overlapped. To evaluate the functional relevance of differentially methylated genes, we performed siRNA-mediated knockdown of SSU72, a gene associated with multiple DMCs. Knockdown of SSU72 resulted in an average 18% decrease in blastocyst formation rate (p < 0.001). Conclusions: These results demonstrate that paternal methionine supplementation alters embryonic DNA methylation patterns and that affected genes may play critical roles in early embryonic development, contributing to fetal programming.
Background/Objectives: Doxorubicin (DOX) is an effective chemotherapeutic agent, but its clinical use is limited by dose-dependent cardiotoxicity. Emerging evidence suggests that epigenetic dysregulation, particularly altered DNA methylation, contributes to DOX-induced cardiac injury. Metformin has been reported to exert cardiometabolic and epigenetic regulatory effects. This study investigated genome-wide DNA methylation changes induced by chronic metformin exposure and their effects on doxorubicin sensitivity in H9c2 cardiomyoblast cells. Methods: Genome-wide DNA methylation changes induced by chronic metformin exposure were investigated in H9c2 cardiomyoblast cells using whole-genome bisulfite sequencing (WGBS). Cells were treated with metformin (0.7-2.8 mM) for four months prior to DOX exposure. Cellular sensitivity to DOX was evaluated using MTT-based dose-response analysis and IC50 estimation. Results: DOX reduced cell viability (IC50 = 0.164 µM). Chronic metformin pre-treatment produced a dose-dependent rightward shift in DOX dose-response curves, increasing IC50 values to 0.21, 0.289, and 0.51 µM at 0.7, 1.4, and 2.8 mM metformin, respectively. WGBS revealed distinct separation between treatment groups in principal component analysis. Significant methylation changes (adjusted p-value < 0.05) were identified in genes related to oxidative stress, mitochondrial function, apoptosis, and chromatin regulation. Conclusions: Chronic metformin exposure induces dose-dependent genome-wide DNA methylation remodeling in cardiac cells and is associated with altered cellular sensitivity to doxorubicin. These findings suggest that metabolic modulation by metformin may influence epigenetic regulation and cellular stress responses relevant to chemotherapy-induced cardiotoxicity.
Structural and functional disruptions of the epigenome are hallmarks of breast and ovarian carcinogenesis. This review dissects the reciprocal regulatory networks co-operated by DNA methyltransferases (DNMTs), ten-eleven translocation enzymes (TETs), and key non-coding RNAs (microRNAs and lncRNAs). We map the precise molecular mechanisms through which these epigenetic modulators alter chromatin accessibility, drive transcriptional reprogramming, and promote phenotypic plasticity in hormone-dependent malignancies. By systematically contrasting the distinct yet overlapping epigenetic profiles of breast and ovarian tumors, we elucidate how these aberrations dictate clinical outcomes. This comprehensive synthesis offers critical insights into the dual utility of these epigenetic elements as dual-purpose diagnostic biomarkers and druggable therapeutic targets, laying the groundwork for next-generation targeted epigenetical therapies.
Turner syndrome (TS), a disorder caused by the complete or partial absence of an X chromosome, exhibits significant clinical variability that cannot be fully explained by chromosomal anomalies alone. This narrative review highlights the crucial role of epigenetic mechanisms, particularly X-chromosome inactivation (XCI), in shaping the TS phenotype. The haploinsufficiency of genes that normally escape XCI is a primary driver of TS features. The specific epigenetic consequences depend on the chromosomal anomaly. In complete monosomy (45,X), the absence of escape-mediated dosage compensation genes from a second X chromosome amplifies haploinsufficiency across X-linked escape genes. Isochromosome Xq (i(Xq)) variants involve the loss of the short arm (Xp) and duplication of the long arm (Xq), creating a dual dosage imbalance with extreme XCI skewing. Carriers of i(Xq) also have a heightened risk for autoimmune disorders compared to those with 45,X TS. For ring-X chromosomes (r(X)), which are mitotically unstable, the functional status of the XIST gene is critical. If the ring is XIST-negative, it remains transcriptionally active, resulting in functional disomy and a more severe phenotype with pronounced neurodevelopmental and craniofacial features. Ultimately, the clinical heterogeneity in TS arises from a complex interplay of the specific chromosomal structure, tissue-specific mosaicism, XIST function, and variable escape from XCI, defining TS as a disorder of epigenetic and gene-regulatory imbalance. However, future research requires a better understanding of the complex mechanism of X-chromosome inactivation.
Background: It is observed that there are a limited number of scientific studies investigating the effect of cocaine use disorder on microRNA (miRNA) levels in human peripheral blood. This study aimed to identify candidate miRNAs that may play a role in the regulation of cocaine addiction by detecting changes in the expression of some miRNAs (miR-9-5p, miR-26b-5p, miR-132-3p, and miR-134-5p) in the peripheral whole blood of cocaine addicts. Methods: Peripheral blood samples were collected from 12 Turkish male individuals with cocaine abuse, 11 Turkish male individuals undergoing treatment for cocaine abuse, and 16 healthy Turkish male individuals without any substance abuse. The change in the expression of microRNAs was determined by quantitative real-time polymerase chain reaction (RT-qPCR). In statistical analyses, ΔCt values were analyzed for the expression of miRNAs. Receiver operating characteristic (ROC) analysis was used to assess the diagnostic adequacy of peripheral blood miRNAs. Results: miR-132-3p and miR-134-5p were downregulated in the addict group compared to the control group (p < 0.05). The areas under the curves (AUCs) of the ROC curve of miR-132-3p and miR-134-5p were significant at 0.778 and 0.744, respectively. Conclusions: This study suggests that miR-132-3p and miR-134-5p may have function as therapeutic markers in the treatment of cocaine use disorder.
Epigenetic regulation has played a fundamental role in the evolution of plant reproduction. Across more than a billion years, ancestral genome-defense mechanisms in early eukaryotes were progressively expanded, diversified, and repurposed throughout the green lineage. Streptophyte algae assembled the first plant-specific methylation and small RNA systems, providing pre-adaptations for terrestrial reproduction. In bryophytes and early vascular plants, these systems became integrated into gametophyte development, sporogenesis, and meiotic genome protection. Seed plants experienced substantial diversification and expansion of chromatin regulators and small RNA machinery, enabling increasingly sophisticated control of cone, ovule, and embryo development. Angiosperms underwent the most dramatic rewiring of epigenetic pathways, including gene-family diversification, subfunctionalization, and the emergence of genomic imprinting, endosperm-specific demethylation, and lineage-specific reproductive small RNAs such as phasiRNAs. Convergent solutions, including imprinting, meiotic transposable element (TE) silencing, and TE-derived regulatory elements, arose independently across lineages. Rather than reflecting the emergence of entirely new molecular machinery, these innovations illustrate repeated functional co-option and regulatory rewiring of deeply conserved epigenetic modules. Ecological and life-history pressures further shaped epigenetic diversification, linking environmental stress, mating systems, and domestication to reproductive epigenetic plasticity. Recent evidence further demonstrates that epigenetic plasticity underlies the recurrent evolution of alternative reproductive strategies such as apomixis and contributes to reproductive responses to environmental stress. Advances in comparative epigenomics, single-cell technologies, and epigenome editing are now providing unprecedented opportunities to reconstruct the evolutionary history of reproductive epigenetic pathways and to harness them for crop improvement. Together, these findings reveal epigenetic regulation as a dynamic, modular, and deeply evolvable framework that has repeatedly enabled reproductive innovation throughout plant evolution.
BACKGROUND:Glutathione peroxidases (GPXs) regulate peroxide detoxification and redox signaling, but their relationship with DNA methylation remains unclear in Oryza sativa. This study evaluated whether silencing mitochondrial GPX1 and GPX3 is associated with changes in growth, antioxidant activity, and DNA methylation-related profiles. METHODS:Non-transformed plants (NT) and five GPX-silenced lines were evaluated in a randomized complete block design. Morphophysiological traits, antioxidant enzyme activities, total 5-methylcytosine content, and methylation-sensitive restriction profiles were analyzed. RESULTS:GPX silencing impaired early establishment and significantly affected flowering time and leaf, root, seed, and total biomass. Total dry biomass decreased by 62.1% in the most affected GPX1 lines and by 29.2% in GPX3 lines relative to NT plants. Root biomass declined by up to 86.1%, and flowering was delayed by up to 30.7 days. Genotype significantly affected GPX-associated and glutathione reductase activities, whereas no significant genotype effects were detected for catalase, ascorbate peroxidase, or superoxide dismutase activities. GPX-associated and glutathione reductase activities were strongly correlated (r = 0.85, p < 0.001), consistent with selective alteration of GPX-associated and glutathione-linked redox metabolism. Total 5-methylcytosine content decreased by 41-42% in GPX-silenced groups. However, increased McrBC digestion and unchanged HpaII/MspI profiles indicated that methylation-related changes were nonuniform and depended on the genomic sites recognized by each enzymatic assay. CONCLUSIONS:These findings show that mitochondrial GPX knockdown is associated with impaired rice growth and reproductive development, as well as with altered total 5-methylcytosine content and restriction-sensitive methylation profiles, suggesting a potential relationship among redox homeostasis, developmental regulation, and epigenetic plasticity that requires further validation using locus-resolved and mechanistic approaches.
Background: Triple-negative breast cancer (TNBC) remains a clinical challenge due to its aggressive nature and the frequent emergence of therapeutic resistance. While the role of protein-coding genes in DNA repair is well-documented, the regulatory contributions of the non-coding genome, specifically long intergenic non-coding RNAs (lincRNAs), remain largely undefined. Objectives: In this study, we characterize the biological significance of LincRNA-BC7, a novel transcript identified within the breast cancer field effect. Methods: Through a combined in silico and in vitro approach, we investigated the transcriptional dynamics of the LincRNA-BC7/miR-663a/BRCA1 axis in response to the PARP inhibitor, Olaparib. Results: Our results demonstrate that Olaparib induces selective cytotoxicity in BRCA1-deficient MDA-MB-231 cells while sparing non-cancerous HEK293 cells, a response accompanied by a significant downregulation of LincRNA-BC7 and a reciprocal upregulation of BRCA1. Bioinformatics analysis through BLASTN, miRBase, and KEGG revealed that LincRNA-BC7 contains highly complementary binding sites for miR-663a, suggesting it functions as a competing endogenous RNA (ceRNA) or “molecular sponge.” Conclusions: By sequestering miR-663a, LincRNA-BC7 appears to modulate the expression of critical signaling nodes within the PI3K-AKT and TP53 pathways, thereby influencing cellular sensitivity to DNA-damaging agents. These findings suggest that LincRNA-BC7 is a key determinant of the aggressive TNBC phenotype and the response to PARP inhibition. Our study establishes the LincRNA-BC7/miR-663a axis as a novel biomarker for precision risk stratification and a promising therapeutic target to enhance treatment outcomes in BRCA1-associated breast cancers.
BACKGROUND:Depression is a heterogeneous and recurrent condition, whose underlying biological mechanisms remain poorly understood. MicroRNAs (miRNAs), small non-coding RNAs that regulate post-transcriptional gene expression, are increasingly implicated in cross sectional miRNA studies of depression and depressive symptoms; however, longitudinal studies capturing miRNA changes over time in relation to depression are scarce. METHODS:We conducted small RNA sequencing of leukocyte-derived miRNAs at two timepoints in a prospective community-based cohort (n = 185) to assess associations between within-person changes in depressive symptom severity (ΔPHQ-9) and longitudinal miRNA expression. Differential expression analyses were performed using a paired limma-voom framework, adjusting for covariates (baseline PHQ-9, sex, age, DNAm-derived immune cell covariates and ancestry components derived from matched blood samples) and within-subject correlation. RESULTS:Although no miRNAs survived multiple-testing correction, 68 mature unique miRNAs showed nominal associations (p < 0.05) with depressive symptom severity change (ΔPHQ-9). Several top candidates, including miR-493-3p, miR-409-3p, and miR-323a-3p, displayed expression patterns aligning with prior reports implicating these miRNAs in stress responsivity, synaptic plasticity, and neurodevelopmental regulation. Exploratory follow-up of predicted targets of the nominally symptom-associated miRNAs converged on genes in pathways central to depression biology, including neurotransmission, HPA axis/inflammatory signaling, neuroplasticity, and circadian regulation. Enrichment analyses highlighted receptor tyrosine kinase and intracellular signaling cascades, hypothalamic-pituitary-adrenal axis feedback, and inflammatory pathways. CONCLUSIONS:These findings provide preliminary evidence that peripheral miRNA expression changes may reflect depressive symptom trajectories, highlighting potential molecular pathways involved in depression. Further studies with larger samples and broader symptom severity are warranted to validate these dynamic miRNA signatures.
Epigenetics regulates gene activity without altering the underlying DNA sequences. Numerous studies have highlighted the importance of epigenetics in diverse physiological processes, including cell growth, differentiation, and tissue development. Increasingly, epigenetic modifications are recognized for their involvement in various diseases, notably corneal disorders. Corneal fibrosis, a common consequence of ocular injury or infection, significantly contributes to visual impairment and blindness worldwide. Recent evidence indicates that epigenetic changes regulate key processes in corneal pathogenesis, such as inflammation, wound healing, extracellular matrix remodeling, fibrosis, and neovascularization. These findings underscore the potential of developing novel therapeutic strategies that specifically target epigenetic mechanisms to treat or mitigate corneal pathology. Nevertheless, bringing epigenetic therapies into clinical practice remains challenging given the complexity of epigenetic regulation. Future research leveraging multi-omics technologies and specific gene manipulation will be essential to elucidate the mechanisms underlying epigenetic regulation in corneal diseases and to identify specific therapeutic targets. Such advancements will drive the development of effective, clinically relevant treatments for corneal fibrosis and related disorders.
DNA methylation, the covalent addition of methyl groups to cytosine (5mC) or adenine (6mA) in DNA, is a fundamental mechanism of epigenetic inheritance conserved from bacteria to humans. Fungi provide a uniquely informative window into the evolutionary logic of methylation systems. Spanning more than 1 billion years of diversification, the kingdom encompasses species that have lost cytosine methylation entirely, lineages that use 5mC to silence transposons and drive the irreversible genome-defense process known as repeat-induced point mutation (RIP), and early-diverging lineages, in which 6mA has emerged as a prominent chromatin mark. The methyltransferases underlying these strategies (DIM-2, RID, DNMT1-RFD, DNMT5, and the MT-A70 complex) and the recently characterized demethylases Dmt1 and CcTet are structurally and mechanistically distinct from their mammalian counterparts. Here we review the mechanisms, targets, and biological functions of fungal DNA methyltransferases and demethylases, incorporating cryo-EM structural insights into DIM-2 and DNMT5 catalysis, analyses of DNMT gene loss as a continuous evolutionary process, the antiviral role of DIM-2 in vegetative hyphae, and the emerging model of 6mA as a heritable regulatory mark in early-diverging lineages. By integrating these advances, this review offers the updated and comprehensive account of DNA methylation across fungi.
BACKGROUND/OBJECTIVES:As important post-transcriptional and epigenetic regulators of gene expression, miRNAs play a pivotal role in modulating host-virus interactions. While prior reviews have addressed either direct miRNA-HIV genome interactions or miRNA-mediated immune modulation in isolation, the integrated dual functionality of these molecules has not been systematically characterized. This review aimed to comprehensively explore how miRNAs that target the HIV-1 genome simultaneously modulate key innate and adaptive host immune signaling pathways. The conceptual novelty of this study is determined not by the identification of previously unknown miRNA-target gene pairs, but by the systemic integration of two regulatory levels (direct inhibition of the viral genome and modulation of the host cell immune signaling pathways) within a unified analytical framework. Such an integrated approach reveals a proviral regulatory network that remains non-obvious when each of these levels is examined separately. METHODS:A narrative review was conducted using PubMed, Scopus, Web of Science, and Google Scholar (all years through 2025). In Stage 1, publications reporting experimentally confirmed interactions between host miRNAs and the HIV-1 genome were identified, yielding a curated set of 15 miRNAs. In Stage 2, target genes for each miRNA were retrieved from miRTarBase, TarBase (experimentally validated) and TargetScan 8.0 (in silico predicted). In Stage 3, target genes were manually mapped to key immune signaling pathways (TLR, NF-κB, JAK-STAT). In Stage 4, targeted literature searches were performed for each miRNA-target gene pair to identify direct experimental evidence of interaction. All stages were performed by two independent researchers, with discrepancies resolved by a third. RESULTS:Fifteen host miRNAs with experimentally confirmed binding to the HIV-1 genome were identified, targeting viral genes including nef, pol, vpr, gag, env, vif, and the 3'-UTR. Thirteen of these miRNAs were found to regulate components of major immune pathways. miR-92a-3p, miR-29a/b-3p, miR-150-5p, and miR-125b-5p emerged as the most pleiotropic regulators, simultaneously suppressing TLR signaling (TLR3, TLR7, TLR8, MyD88, TRAF3/6, IRAK1/4), NF-κB components (REL, RELA, NFKB1), JAK-STAT effectors (STAT1-3, STAT5A/B, JAK2), and negative regulators of cytokine signaling (SOCS and PIAS family proteins). miR-133b and miR-196b-5p were found to selectively regulate SOCS/PIAS proteins without involvement in other analyzed pathways, suggesting potential for selective therapeutic targeting. CONCLUSIONS:The analyzed miRNAs exhibit functional dualism, acting as direct post-transcriptional suppressors of the HIV-1 genome while simultaneously functioning as epigenetic modulators of host immune signaling. These two modes of action are not independent but together form a conceptual framework of a self-reinforcing proviral regulatory network that, based on the synthesis of published evidence, is proposed to promote viral latency and immune evasion. The identified miRNAs represent promising, albeit complex, targets for novel therapeutic strategies aimed at eliminating latent HIV reservoirs.
Background: The transfer of a nucleus from one oocyte to another offers patients harbouring high levels of mitochondrial DNA mutation and sufferers of frequent fertilisation failure or early embryonic arrest the potential to have healthy children. However, a small amount of mtDNA is carried over with the nucleus as the transfer takes place. Consequently, we still need to distinguish between the effects of the carryover and the transfer of a nucleus itself from a mature oocyte. Methods: To overcome this, we analysed a series of hatching stage blastocysts generated using metaphase II spindle transfer and mitochondrial supplementation. The latter approach also introduces a small amount of mtDNA into the oocyte as fertilisation takes place. For both manipulations, an autologous approach was used to overcome the effects of third-party transfer. Results: We then compared the changes in global gene expression between the two groups. We found that the nuclear transfer process affected a number of gene networks and pathways. These included metabolic, cell cycle, inflammatory and immune, and epigenetic responses. A comparison with earlier stage blastocysts did not suggest that the cause was due to developmental delay. Conclusions: Critically, these changes could affect offspring health and well-being as is the case following somatic cell nuclear transfer.
Aim: The aim of this study was to correlate single-nucleotide polymorphisms (SNPs) in the FTO and MC4R genes with body composition (BC) in populations with various levels of physical activity, and to investigate associations of SREBF1 methylation with the level of physical activity (PA) and BC. Methods: Fifty-six participants aged 18-65 years old with no underlying medical conditions were included in the study and were classified into sedentary/light PA (SLPA), moderate PA (MPA) and vigorous PA (VPA) groups using the International PA questionnaire (IPAQ). Anthropometric measures such as age, gender, body mass index (BMI) and body fat percentage (BFP) were recorded at the time of recruitment. Venous blood samples were collected during participant recruitment and DNA was extracted. Genotyping assays were performed for SNPs in FTO (rs9939609) and MC4R (rs17782313) using Taqman® RT qPCR and TaqMan Genotyper software 1.7.1. Methylation analysis assay for CpG sites in the SREBF1 gene was performed on 56 samples using PyroMark® Q48 Autoprep (Qiagen, Venlo, The Netherlands). The results were statistically analysed to identify any associations between FTO/MC4R genotypes and the level of PA, and between SREBF1 methylation status and the level of PA. This is the first study to investigate links between PA and quantitative methylation of SREBF1. Results: According to IPAQ guidance, the 56 participants were classified into SLPA n = 14, MPA n = 11 and VPA n = 31. The correlation analysis revealed that the FTO rs9939609 'A' risk allele had a significant negative association with BFP in the VPA group (p = 0.0387); the MC4R rs17782313 'C' risk allele had a significant positive association with BMI in the VPA group (p = 0.0256). In the SREBF1 pyrosequencing analysis, higher levels of methylation were observed in the VPA group (p = 0.07). Conclusions: We concluded that SNPs associated with obesity identified in FTO rs9939609 and MC4R rs17782313 could help to predict the molecular effects of PA. A high frequency of FTO risk variants in the cohort was observed and the VPA group could help maintain a healthy BFP.
Background/Objectives: West Nile virus (WNV) remains a significant threat to human health, with no approved antiviral treatments or vaccine available. A better understanding of the molecular mechanisms governing flavivirus-host interactions is needed to identify host regulatory pathways involved in infection. This study aimed to investigate how WNV infection remodels the host miRNA-mRNA regulatory landscape. Methods: WNV-induced changes in host miRNA expression in HEK-293 cells were profiled using miRNA-Seq. Transcriptome-wide host gene expression changes in WNV-infected cells were analysed using RNA-Seq. Gene Ontology and pathway enrichment analyses were conducted using DAVID. Integrated miRNA-mRNA network reconstruction was performed using Cytoscape based on the experimentally validated miRNA-mRNA interactions in miRNet database. Results: WNV infection induced global changes in host miRNA expression, with pathogenic NY99 and non-pathogenic Kunjin strains of the virus producing overlapping and strain-specific alterations in the miRNA landscape. Transcriptome analysis showed strong induction of interferon-related responses and activation of NF-κB and MAPK signalling pathways in the infected cells. In contrast, pathways associated with RNA processing, splicing, and proteasomal degradation were downregulated. Integrated miRNA-mRNA network analysis identified miR-197-3p, miR-301b-3p, miR-129-3p, miR-3662, and miR-128-5p as candidate regulatory hubs involved in WNV-induced transcriptome remodelling. These networks suggested that miRNA-mediated regulation may influence antiviral signalling, apoptosis, and RNA metabolism during infection. Conclusions: These findings suggest that WNV infection broadly remodels host miRNA-mRNA regulatory networks and identifies candidate miRNAs that may contribute to the regulation of antiviral and cellular stress responses. These predicted regulatory interactions provide a foundation for future experimental validation.
Background: Long-read, single-CpG-resolution sequencing is redefining the information-to-depth ratio in epigenomics. While conventional methylome analysis often requires high coverage, we propose a scalable pipeline designed to extract high-density regulatory logic from shallow sequencing data. Methods: By utilizing the progenitor-like HepaRG cell line as a model for liver plasticity, we validated this framework across two divergent developmental trajectories: hepatic maturation and sphere-induced retrodifferentiation. Our technical approach combines CpG-centric enrichment and regional methylation aggregation to reconstruct regulatory landscapes from sparse data. Using long-read Nanopore sequencing, we mapped the dynamics of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC). Results: Our pipeline revealed that these trajectories are not inverse processes but engage distinct epigenetic strategies. Hepatic maturation is characterized by the accumulation of 5hmC that partially targets repressive heterochromatin (H3K9me3, H4K20me3) and pioneer factors such as FOXA2. In contrast, retrodifferentiation increases 5mC, potentially silencing adult regulators such as HNF1A via Polycomb-associated networks. In addition, aggregation-based analysis can distinguish widespread focal perturbations from a restricted subset of transcription factors that translate epigenetic changes into regional accessibility. Conclusions: This study provides a scalable computational framework for investigating cellular fate transitions, proving that high-value epigenetic insights are attainable even at reduced sequencing depths.
BACKGROUND/OBJECTIVES:Opioid use disorder (OUD) is caused by a complex interplay between genetic and non-genetic factors. DNA methylation is an epigenetic mechanism that modulates gene expression. Data on DNA methylation and opioid addiction and treatment are limited. This association study was designed to assess the difference in genome-wide methylation patterns between individuals with OUD in methadone maintenance treatment (MMT) (n = 114) and those with OUD who achieved long-term abstinence (>10 years) without mu opioid receptor agonist treatment (n = 136). METHODS:Differential DNA methylation analysis was performed in whole blood using the Illumina EPIC array. RESULTS:A total of 135 differentially methylated probes (DMPs) reached epigenome-wide significance (p < 1 × 10-7), controlling for sex, age, estimates of blood cell proportions, and the first two principal components based on genome-wide SNP genotypes. The methylation sites were annotated to 157 genes, including 32% long non-coding RNAs. These genes are related to several systems, including cell adhesion (e.g., SAXO4), immune system and inflammation (e.g., UBTF, USP39, C10orf90, PRKCA), stress response (e.g., CRHR1, GPR19), and spermatogenesis (e.g., SPATA16, COX7B2). DMP cg11641410 is located in lncRNA ENSG00000254687, an antisense to OPRK1. Six of the DMPs were also identified in a related longitudinal study of MMT. CONCLUSIONS:At this point, it is not possible to determine whether the minor methylation differences observed in this study cause clinically relevant changes in gene expression. However, these findings have the potential to identify biomarkers and to provide new targets for treatment optimization.
The ubiquitously transcribed tetratricopeptide repeat Y-linked gene (UTY/KDM6C), a catalytically impaired histone demethylase encoded on the Y chromosome, has garnered increasing attention for its emerging roles in tumorigenesis and cancer progression. Despite high sequence homology with its X-linked paralog UTX/KDM6A, UTY exhibits markedly reduced or absent H3K27me3 demethylase activity due to critical amino acid substitutions in its Jumonji C domain. Consequently, UTY primarily functions through non-enzymatic mechanisms, acting as a scaffold in chromatin-remodelling complexes like COMPASS and SWI/SNF, or mediating protein-protein interactions that regulate transcriptional programs independent of demethylation. This aligns with epigenetic dysregulation in cancers, where imbalances in repressive H3K27me3 and active H3K4me either drive tumour suppressor silencing or oncogene activation. Unlike frequently mutated UTX in cancers such as breast, renal cell carcinoma, and acute myeloid leukaemia, UTY's contributions in cancer are less defined, constrained by male-specific expression. Emerging evidence suggests UTY as a context-dependent tumour suppressor in AML and squamous-like pancreatic ductal adenocarcinoma. While direct functional validation remains limited in several cancer types, UTY is increasingly implicated as a potential tumour suppressor in haematological malignancies and prostate cancer. Therapeutically targeting UTY's scaffold functions shows promise for male-specific cancers and merits future investigation.
Background/Objectives: Parity, the number of times a woman carries a pregnancy to viability, has been linked to long-term maternal health outcomes. The mechanisms linking parity to health outcomes are poorly understood but may reflect influences of pregnancy on the maternal epigenome. Methods: This study examines the relationship between parity and DNA methylation (DNAm) during pregnancy using data from three cohorts: the Norwegian Mother, Father and Child Cohort Study (MoBa), the Atlanta African American Maternal-Child (AAAMC) cohort, and the Isle of Wight (IOW) Birth Cohort. Results: An epigenome-wide association study (EWAS) in MoBa identified 5374 cytosine-phosphate-guanine sites (CpGs) that were statistically significantly associated with parity, of which 69% were positively and 31% negatively correlated. Replication analyses confirmed 3491 CpGs in at least one cohort, and 93 CpGs in both AAAMC and IOW. Gene enrichment analysis revealed significant involvement of developmental and signaling pathways, including calcium signaling and neuroactive ligand-receptor interaction. Additionally, 584 differentially methylated regions (DMRs) were detected, with 90% overlapping individual parity-related CpGs. Conclusions: These findings suggest that parity influences epigenetic patterns, potentially affecting biological processes and molecular functions relevant to maternal health later in life.
