DNA, Volume 5, Issue 4 (December 2025) – 15 articles

In animals and humans, nutrients influence signaling cascades, transcriptional programs, chromatin dynamics, and mitochondrial function, collectively shaping traits related to growth, immunity, reproduction, and stress resilience. This review synthesizes evidence supporting nutrient-mediated regulation of DNA methylation, histone modifications, non-coding RNAs, and mitochondrial biogenesis, and emphasizes their integration within metabolic and developmental pathways. Recent advances in epigenome-wide association studies (EWAS), single-cell multi-omics, and systems biology approaches have revealed how diet composition and timing can reprogram gene networks, sometimes across generations. Particular attention is given to central metabolic regulators (e.g., PPARs, mTOR) and to interactions among methyl donors, fatty acids, vitamins, and trace elements that maintain genomic stability and metabolic homeostasis. Nutrigenetic evidence further shows how genetic polymorphisms (SNPs) in loci such as IGF-1, MSTN, PPARs, and FASN alter nutrient responsiveness and influence traits like feed efficiency, body composition, and egg quality, information that can be exploited via marker-assisted or genomic selection. Mitochondrial DNA integrity and oxidative capacity are key determinants of feed conversion and energy efficiency, while dietary antioxidants and mitochondria-targeted nutrients help preserve bioenergetic function. The gut microbiome acts as a co-regulator of host gene expression through metabolite-mediated epigenetic effects, linking diet, microbial metabolites (e.g., SCFAs), and host genomic responses via the gut–liver axis. Emerging tools such as whole-genome and transcriptome sequencing, EWAS, integrated multi-omics, and CRISPR-based functional studies are transforming the field and enabling DNA-informed precision nutrition. Integrating genetic, epigenetic, and molecular data will enable genotype-specific feeding strategies, maternal and early-life programming, and predictive models that enhance productivity, health, and sustainability in poultry production. Translating these molecular insights into practice offers pathways to enhance animal welfare, reduce environmental impact, and shift nutrition from empirical feeding toward mechanistically informed precision approaches. Full article

The cannabinoid receptor type 2 (CB₂) is increasingly recognized as a crucial regulator of neuroimmune balance in the brain. In addition to its well-established role in immunity, the CB₂ receptor has been identified in specific populations of neurons and glial cells throughout various brain regions, and its expression is dynamically increased during inflammatory and neuropathological conditions, positioning it as a potential non-psychoactive target for modifying neurological diseases. The expression of the CB₂ gene (CNR2) is finely tuned by epigenetic processes, including promoter CpG methylation, histone modifications, and non-coding RNAs, which regulate receptor availability and signaling preferences in response to stress, inflammation, and environmental factors. CB₂ signaling interacts with TRP channels (such as TRPV1), nuclear receptors (PPARγ), and orphan G Protein-Coupled Receptors (GPCRs, including GPR55 and GPR18) within the endocannabinoidome (eCBome), influencing microglial characteristics, cytokine production, and synaptic activity. We review how these interconnected mechanisms affect neurodegenerative and neuropsychiatric disorders, underscore the species- and cell-type-specificities that pose challenges for translation, and explore emerging strategies, including selective agonists, positive allosteric modulators, and biased ligands, that leverage the signaling adaptability of the CB₂ receptor while reducing central effects mediated by the CB₁ receptor. This focus on the neuro-centric perspective repositions the CB₂ receptor as an epigenetically informed, context-dependent hub within the eCBome, making it a promising candidate for precision therapies in conditions featuring neuroinflammation. Full article

Background/Objectives: Goat and sheep farming is an important agro-economic resource in Benin. However, their milk is both underutilized and insufficiently characterized, which limits the development of innovative dairy products and raises concerns about its safety. Against this backdrop, our pioneering study set out to investigate, for the first time in Benin and using an advanced metagenomic approach, the microbial diversity present in goat and sheep raw milk. The aim was to lay the groundwork for safer and more efficient dairy valorization. Methods: To achieve this, metagenomic DNA was extracted from 20 pooled milk samples representing both animal species, followed by shotgun sequencing. Results: Analyses revealed seven dominant phyla: Bacillota (17.44–27.23%), Pseudomonadota (12.39–15.55%), Campylobacterota (3.65–5.29%), Actinomycetota (1.47–6.03%), Spirochaetota (1.14–2.02%), Apicomplexa (0.28–0.50%), and Bacteroidota (0.17–0.22%) in the raw milk of both species. However, their proportions differ. Bacillota, which was the most dominant in both types of milk, was significantly more abundant in goat (27.23 ± 5.33) than in sheep milk (17.44 ± 8.44). In sheep milk, Enterobacteriaceae (11.36 ± 5.79) were the most predominant family, followed by Streptococcaceae (5.57 ± 2.29) and Staphylococcaceae (4.51 ± 3.63). Goat milk, on the other hand, presents a different hierarchy. Streptococcaceae (6.65 ± 2.19) and Staphylococcaceae (6.43 ± 2.33) were the most abundant families, surpassing Enterobacteriaceae (5.33 ± 1.66). The genus Escherichia was the most abundant in sheep milk (6.18 ± 5.33). The genera Staphylococcus (4.50 ± 3.63) and Streptococcus (5.05 ± 1.98) were also present. In contrast, in goat milk, the genera Streptococcus (6.54 ± 2.35) and Staphylococcus (6.42 ± 2.32) were the most dominant, while the average abundance of Escherichia was much lower (1.98 ± 1.28). In terms of species, Sheep milk was dominated by Escherichia coli (6.14 ± 5.28) and Staphylococcus aureus (5.17 ± 2.28) while Klebsiella pneumoniae (2.82 ± 1.72), Streptococcus pneumoniae (1.92 ± 1.36), and Campylobacter coli (1.52 ± 1.27) were also found. In addition to a relatively high abundance of Staphylococcus aureus (6.40 ± 2.45), goat milk was characterized by the presence of Corynebacterium praerotentium (5.32 ± 2.28) and Clostridium perfringens (3.39 ± 2.09). Additional pathogens identified included Clostridioides difficile (1.17–2.00%), Clostridium botulinum (0.27–0.43%), Listeria monocytogenes, Mycobacterium tuberculosis, Helicobacter pylori (0.36–0.62%), Salmonella enterica (0.22–0.26%). As for fungi, Ascomycota were predominant, with the presence of Aspergillus fumigatus, Saccharomyces cerevisiae, Trichophyton mentagrophytes, and Candida auris. Moreover, lactic acid bacteria with technological interest such as Oenococcus oeni (0.60–0.97%), Levilactobacillus namurensis (0.25–0.44%), Lactobacillus agrestimuris, and Lacticaseibacillus rhamnosus were also detected. Conclusions: These findings provide essential insights into the technological potential and health risks associated with these milks, which are key to developing safer and more efficient local dairy value chains. Full article

CRISPR-based transcriptional regulation technologies, including CRISPR activation (CRISPRa) and CRISPR interference (CRISPRi), offer precise and programmable control over gene expression, representing a major advance in gene and epigenetic therapy. CRISPRa uses nuclease-inactive Cas proteins fused to transcriptional activators to upregulate target genes, while CRISPRi employs repressor domains for gene silencing. Preclinical studies have demonstrated the efficacy of CRISPRa/i in models of metabolic, neurological, muscular, and oncological diseases. Notably, CRISPRi-based therapies have entered clinical trials for conditions like hepatitis B and muscular dystrophy, showing encouraging safety and efficacy profiles. Despite ongoing challenges related to delivery efficiency, immunogenicity, and off-target activity, innovations in protein engineering and guide RNA design are rapidly enhancing the precision and safety of these technologies. Overall, CRISPRa and CRISPRi are poised to transform the treatment of genetic and epigenetic disorders, with continued optimization expected to accelerate their clinical adoption and broaden their therapeutic impact. Full article

Resistance exercise (RE) is a well-known modality to increase skeletal muscle strength and hypertrophy. While both high-load (HL) and low-load (LL) RE stimulate skeletal muscle growth, the effects of RE load on androgen-regulated genes remain unclear. Further, the relationship between circulating and intramuscular androgen-associated targets and muscular strength and mass has not been well defined. Purpose: This investigation therein aimed to examine acute gene and hormone responses to volume- and intensity-equated RE at different loads, examining their relationships with lean body mass (LBM), strength, and circulating and intramuscular androgen-related biomarkers. Methods: Ten resistance-trained males completed one-repetition maximum (1RM) testing, as well as body composition testing, before two volume- and intensity-equated RE sessions, separated by a 7–10 day crossover period. Serum and skeletal muscle samples were collected at baseline, 3 h, and 24 h post-exercise to assess testosterone (TST), dihydrotestosterone (DHT), AR protein, AR mRNA, and AR–DNA binding. Pearson correlations evaluated any potential associations between LBM, strength, and androgen/AR biomarkers. Results: Training load did not significantly impact gene expression, but time effects were observed, whereby MyoD peaked 3 h post-exercise (2.03 ± 1.64 fold; p = 0.005), while AR mRNA decreased at 24 h (0.54 ± 0.42 fold; p = 0.021) versus baseline. LBM also correlated with bench press (r = 0.607, p = 0.048) and leg press (r = 0.705, p = 0.015) 1RM. Serum total TST correlated with leg press 1RM (r = 0.909, p = 0.012), while serum-free TST correlated with AR mRNA fold-change (r = 0.392, p = 0.001) and AR–DNA binding (r = 0.287, p = 0.021). Intramuscular DHT correlated with intramuscular TST (r = 0.415, p < 0.001) and AR protein (r = 0.421, p < 0.001). Lastly, fold changes in AR mRNA were correlated with MyoD mRNA fold changes (r = 0.785, p = 0.007) along with IGF1-Ea mRNA fold changes being significantly correlated with both myogenin mRNA fold changes (r = 0.865, p = 0.001) and AR-DNA binding (r = −0.727, p = 0.017). Conclusions: Despite no observable load-specific effects, RE elicited time-dependent increases in MyoD and AR mRNA expression. This reinforces prior LBM and maximal muscular strength relationship evidence whilst also lending new insights into circulating and intramuscular androgen interactions with AR. Full article

Best known as an organizer of the mitotic spindle, the protein product of the human assembly factor for spindle microtubules (ASPM) gene has recently been shown to function in the interphase nucleus during multiple DNA-associated processes, including BRCA1-mediated DNA DSB repair, ATR-CHK1 activation during replication stress, and transcription regulation alongside the transcription factor FOXM1. In this review, we provide an overview of these DNA-related roles of ASPM. Additionally, we suggest the facilitation of liquid–liquid phase separation (LLPS) as a potential unifying mechanism underlying ASPM function. We also consider the implications of LLPS and ASPM dysfunction in disease, and highlight the impact of cellular context including cell cycle phase-dependent post-translational protein modifications and ion concentrations. An increased understanding of LLPS in ASPM function relevant to genome stability may enable future drug discovery for diseases such as cancer. Full article

Human populations are classified as secretors or non-secretors by, respectively, the ability to produce or not produce FUT2 enzyme (alpha-1,2-fucosyltransferase; FUT2 gene). Non-secretors have some protection against diseases and viral infections. Two single-nucleotide polymorphisms (SNPs), rs601338 (non-secretor; sese), and rs1047781 (weak secretor; Se^w), are known population markers. In this study, 68 saliva samples collected from children living in the Brazilian Amazon region were evaluated for zygosity—genotyping (homozygous or heterozygous) of the rs601338 and rs1047781 SNPs by pyrosequencing. Nine children were heterozygous (Sese) for the rs601338 SNP (13.2%; 9/68) and one homozygous (sese) (1.5%; 1/68). One child that was heterozygous for the rs601338 SNP was also heterozygous for the rs1047781 SNP (Se^w) (1.5%; 1/68). By using Sanger nucleotide sequencing of the FUT2 coding region, strongly linked SNPs (171A>G, 216C>T, 357T>C, 428G>A, 739G>A, 960A>G), including the FUT2*01N.02 allele (428G>A; 739A>G), were detected and have been associated with non-secretor children. A novel SNP (315C>T) and others (40A>G; 480C>T; 863C>T) detected in worldwide populations were also detected. The sensitivity of the pyrosequencing method provided an unprecedented discovery of the zygosity of the SNP rs1047781 only previously detected in East and Southeast Asians. The identification of novel SNPs in this population expands our knowledge of genetic susceptibility to viral infections. Full article

Myalgic Encephalomyelitis (ME), also known as chronic fatigue syndrome (CFS), is a debilitating and heterogeneous disorder marked by persistent fatigue, post-exertional malaise, cognitive impairment, and multisystem dysfunction. Despite its prevalence and impact, the molecular mechanisms underlying ME remain poorly understood. This review synthesizes current evidence on the role of DNA, both nuclear and mitochondrial, in the susceptibility and pathophysiology of ME. We examined genetic predispositions, including familial clustering and candidate gene associations, and highlighted emerging insights from genome-wide and multi-omics studies. Mitochondrial DNA variants and oxidative stress-related damage are discussed in relation to impaired bioenergetics and symptom severity. Epigenetic modifications, particularly DNA methylation dynamics and transposable element activation, are explored as mediators of gene–environment interactions and immune dysregulation. Finally, we explored the translational potential of DNA-based biomarkers and therapeutic targets, emphasizing the need for integrative molecular approaches to advance diagnosis and treatment. Understanding the DNA-associated mechanisms in ME offers a promising path toward precision medicine in post-viral chronic diseases. Full article

Background/Objectives: DNA interstrand cross-links (ICLs) mark one of the most deleterious lesions that can preclude strand separation required for essential cellular processes. Efforts to discover ICL-inducing agents and endogenous substrates for ICL repair pathways have led to the identification of structurally diverse ICLs produced by reactive aldehydes and abasic sites, among others. While several studies point to UV rays as ICL-inducing agents, UV ray-induced ICL formation from biologically relevant DNA lesions has been rarely reported. We conjectured that solar radiation-induced reactive oxygen species may give rise to ICLs via further oxidation of DNA lesions with lower redox potential (e.g., 8-oxoadenine (oxoA)). Here, we present the discovery of ICL production via light-induced modification of the major oxidative adenine lesion oxoA. Methods/Results: In the absence of a photosensitizer, both UVC and UVB rays, but not UVA and visible rays, trigger the formation of oxoA-G ICLs, albeit in low yields. By contrast, the inclusion of the naturally occurring photosensitizer riboflavin in the cross-linking reaction makes UVA and visible rays readily generate oxoA-G ICLs, suggesting solar radiation facilitates the formation of oxoA ICLs in vivo. Conclusions: The plausible oxoA-G ICL formation mechanism concerns the further oxidation of oxoA into an iminoquinone, followed by the nucleophilic attack of the opposite guanine on the iminoquinone. OxoA-G ICLs represent rare examples of ICLs produced by photosensitization. These results will contribute to the discovery of a novel form of ICLs induced by solar radiation. Full article

Glucosinolates (GSLs) are nitrogen- and sulfur-containing secondary metabolites central to the defense, development, and environmental responsiveness of Brassicaceae species. While the enzymatic steps and transcriptional networks underlying GSL biosynthesis have been extensively characterized, mounting evidence reveals that chromatin-based processes add a critical, yet underexplored, layer of regulatory complexity. Recent studies highlight the roles of DNA methylation, histone modifications, and non-coding RNAs in modulating the spatial and temporal expression of GSL biosynthetic genes and their transcriptional regulators in response to developmental cues and environmental signals. This review provides a comprehensive overview of GSL classification, biosynthetic pathway architecture, transcriptional regulation, and metabolite transport, with a focus on emerging epigenetic mechanisms that shape pathway plasticity. We also discuss how these insights may be leveraged in precision breeding and epigenome engineering, including the use of CRISPR/dCas9-based chromatin editing and epigenomic selection, to optimize GSL content, composition, and stress resilience in cruciferous crops. Integrating transcriptional and epigenetic regulation thus offers a novel framework for the dynamic control of specialized metabolism in plants. Full article

Background/Objectives: Environmental DNA (eDNA) is increasingly recognised as a powerful molecular tool for biodiversity monitoring, enabling the detection of species through trace genetic material found in environmental samples. This study investigates the utility of eDNA analysis for identifying vertebrate marine species in the central Mediterranean, with a focus on taxa that serve as ecological indicators to local ecosystems. Methods: Seawater samples were collected from nine sites around the Maltese Islands between May and August 2021, at depths ranging from 2 to 5 m. Samples were filtered and DNA was extracted, amplified and sequenced. The resulting sequences were processed through a bioinformatics pipeline, clustered into molecular operational taxonomic units (MOTUs) and assigned taxonomic identities using reference databases. Results: This study led to the detection of 70 MOTUs, including ecologically important species such as the loggerhead turtle (Caretta caretta), the striped dolphin (Stenella coeruleoalba) and the bottlenose dolphin (Tursiops truncatus), underscoring the method’s effectiveness in the detection of taxa of conservation value. Additionally, we detected a number of overlooked Blenniidae and Gobiidae taxa and deep-water or rarely encountered species such as the ocean sunfish (Mola mola), Cornish blackfish (Schedophilus medusophagus), Haifa grouper (Hyporthodus haifensis) and Madeira lantern fish (Ceratoscopelus maderensis). eDNA of the invasive dusky spinefoot (Siganus luridus) and that of the lumpfish (Cyclopterus lumpus), a species not previously recorded in Maltese waters, was also detected during this study. The latter’s detection highlights the potential of this methodology as an early detection tool for biological invasions. Conclusions: These findings support the integration of eDNA surveillance into marine biodiversity monitoring frameworks, particularly within marine protected areas to monitor native indicator taxa and assess the effectiveness of conservation measures, but also in ports and bunkering zones, where the risk of alien species introduction is elevated, with potential subsequent invasive species expansion that impacts native species and habitats. Full article

Background/Objectives: Mutations in NACHT, LRR and PYD domain-containing protein 2 (NLRP2) and NLRP7 genes, members of the NOD-like receptor (NLR) family of innate immune sensors, result in recurrent miscarriages and reproductive wastage in women. These genes have been identified to be maternal effect genes in humans and mice regulating early embryo development. Previous research in vitro suggests that NLRP2 and NLRP7 regulate DNA methylation and/or immune signaling through inflammasome formation. However, the exact mechanisms underlying NLRP2 and NLRP7 function are not well defined. Methods: To determine the interacting proteins required for NLRP2/NLRP7-mediated regulation of DNA methylation, yeast 2-hybrid screens, coimmunoprecipitation, and FRET studies were performed and verified the ability of novel protein interactions to affect global DNA methylation by 5-methylcytosine-specific ELISA. Results: Various methodologies employed in this research demonstrate a novel protein interaction between human ErbB3-binding protein 1 (EBP1, also known as proliferation-associated protein 2G4 (PA2G4) and NLRP2 or NLRP7. In addition, NLRP2 and NLRP7 regulate EBP1 gene expression. Functionally, global DNA methylation levels appeared to decrease further when NLRP2 and NLRP7 were co-expressed with EBP1, although additional studies may need to confirm the significance of this effect. Conclusions: Since EBP1 is implicated in apoptosis, cell proliferation, DNA methylation, and differentiation, our discovery significantly advances our understanding of how mutations in NLRP2 or NLRP7 may contribute to reproductive wastage in women through EBP1. Full article

Multiple sclerosis (MS) is a chronic autoimmune neurodegenerative disorder characterized by progressive demyelination and axonal degeneration within the central nervous system, driven by complex genomic and epigenomic dysregulation. Its pathogenesis involves aberrant DNA methylation patterns at CpG islands of numbers of genes like OLIG1 and OLIG2 disrupting protein expression at myelin with compromised oligodendrocyte differentiation. Furthermore, histone modifications, particularly H3K4me3 and H3K27ac, alter the promoter regions of genes responsible for myelination, affecting myelin synthesis. MS exhibits chromosomal instability and copy number variations in immune-regulatory gene loci, contributing to the elevated expression of genes for pro-inflammatory cytokines (TNF-α, IL-6) and reductions in anti-inflammatory molecules (IL-10, TGF-β1). Vitamin D deficiency correlates with compromised immune regulation through hypermethylation and reduced chromatin accessibility of vitamin D receptor (VDR) dysfunction and is reported to be associated with dopaminergic neuronal loss. Vitamin D supplementation demonstrates therapeutic potential through binding with VDR, which facilitates nuclear translocation and subsequent transcriptional activation of target genes via vitamin D response elements (VDREs), resulting in suppression of NF-κB signalling, enhancement of regulatory T-cell (T_reg) responses due to upregulation of specific genes like FOXP3, downregulation of pro-inflammatory pathways, and potential restoration of the chromatin accessibility of oligodendrocyte-specific gene promoters, which normalizes oligodendrocyte activity. Identification of differentially methylated regions (DMRs) and differentially expressed genes (DEGs) that are in proximity to VDR-mediated gene regulation supports vitamin D supplementation as a promising, economically viable, and sustainable therapeutic strategy for MS. This systematic review integrates clinical evidence and eventual bioinformatical meta-analyses that reference transcriptome and methylome profiling and identify prospective molecular targets that represent potential genetic and epigenetic biomarkers for personalized therapeutic intervention. Full article

Plants have evolved a complex, multilayered immune system that integrates molecular recognition, signaling pathways, epigenetic regulation, and small RNA-mediated control. Recent studies have shown that DNA-level regulatory mechanisms, such as RNA-directed DNA methylation (RdDM), histone modifications, and chromatin remodeling, are critical for modulating immune gene expression, allowing for rapid and accurate pathogen-defense responses. The epigenetic landscape not only maintains immunological homeostasis but also promotes stress-responsive transcription via stable chromatin modifications. These changes contribute to immunological priming, a process in which earlier exposure to pathogens or abiotic stress causes a heightened state of preparedness for future encounters. Small RNAs, including siRNAs, miRNAs, and phasiRNAs, are essential for gene silencing before and after transcription, fine-tuning immune responses, and inhibiting negative regulators. These RNA molecules interact closely with chromatin features, influencing histone acetylation/methylation (e.g., H3K4me3, H3K27me3) and guiding DNA methylation patterns. Epigenetically encoded immune memory can be stable across multiple generations, resulting in the transgenerational inheritance of stress resilience. Such memory effects have been observed in rice, tomato, maize, and Arabidopsis. This review summarizes new findings on short RNA biology, chromatin-level immunological control, and epigenetic memory in plant defense. Emerging technologies, such as ATAC-seq (Assay for Transposase-Accessible Chromatin using Sequencing), ChIP-seq (Chromatin Immunoprecipitation followed by Sequencing), bisulfite sequencing, and CRISPR/dCas9-based epigenome editing, are helping researchers comprehend these pathways. These developments hold an opportunity for establishing epigenetic breeding strategies that target the production of non-GMO, stress-resistant crops for sustainable agriculture. Full article

Background/Objectives: Intrinsically disordered protein regions (IDRs) are difficult to study due to their flexible nature and transient interactions. Computational folding using AlphaFold may offer one way to explore potential folding of these regions under various conditions. Human DNA topoisomerase IIα (TOP2A) is an essential enzyme involved in regulating DNA topology during replication and cell division. TOP2A has an IDR at the C-terminal domain (CTD) that has been shown to be important for regulating TOP2A function, but little is known about potential conformations that it may undertake. Methods: Utilizing the AlphaFold 3 (AF3) model by way of AlphaFold Server, TOP2A was folded as a dimer first without and then with 29 literature-supported post-translational modifications (PTMs) and DNA to observe whether there is predicted folding. Results: TOP2A CTD does not fold in the absence of PTMs. With the addition of PTMs, however, the CTD is predicted to fold into a globular bundle of loops and α-helices. While DNA alone did not induce folding, in the presence of PTMs, DNA ligands increased helicity of the folded CTD and caused it to interact at different core domain interfaces. In addition, DNA is predicted to enable folding of the TOP2A CTD in the presence of fewer PTMs when compared to the absence of DNA. Conclusions: AF3 predicts the folding of TOP2A CTD in the presence of specific PTMs, and this folding appears to shift to allow binding to DNA in functionally relevant regions. These studies provide predicted folding patterns that can be tested by biochemical approaches. AF3 may support the development of testable hypotheses regarding IDRs and enables researchers to model protein-DNA interactions. Full article