Over the past decades, vast progress in sequencing and imaging has transformed our understanding of chromatin organization in the cell nucleus and its relevance in development and disease. Imaging methods have regained popularity in recent years, as they inherently facilitate single-cell resolution and visualization of subnuclear spatial distances and morphology. An exceptional strength of imaging is the relative ease of designing multimodal experiments, as DNA and RNA fluorescence in situ hybridization (FISH) can be combined with immunofluorescencestaining for joint visualization of genomic loci, transcripts, and proteins. In this mini-review, we highlight recent imaging developments that advance our views on 3D genome organization, focusing primarily on oligonucleotide-based DNA FISH and emerging applications (e.g.chromatin tracing, live-cell imaging, lineage reconstruction, and optical pooled screening). To help novices navigate this rapidly expanding field, we provide a comprehensive overview of the main oligo-based FISH methods.

Tuning the state: Matching condensate material properties with physiological demands and pathological dysfunction.

Now You See Me: Visualizing nuclear complexity by selective staining.

Understanding how fungi regulate mycotoxin production is critical for managing crop diseases and reducing contamination in food systems. Here, we elucidate the mechanism of the transcription factor FgSge1 in Fusarium graminearum, a significant fungal pathogen responsible for Fusarium head blight in cereal crops, in regulating mycotoxin biosynthesis and pathogenicity. FgSge1 specifically binds to the 8-bp cis-element TAARGTTT. Under mycotoxin-induced conditions, FgSge1 binds to this cis-element within its own promoter, activating its own transcription. Additionally, FgSge1 connects with this cis-element within the promoters of mycotoxin biosynthesis genes and interacts directly with the scaffold protein FgAda2 of the Spt-Ada-Gcn5-Acetyltransferase (SAGA) complex. This interaction recruits the histone acetyltransferase FgGcn5 to the promoters of DON biosynthetic genes, promoting histone acetylation and facilitating jet-like chromatin architecture, thereby activating transcription. In contrast, the FgSge1 mutant fails to recruit the SAGA complex, leading to reduced histone acetylation, disrupted chromatin structure, and impaired DON biosynthesis. Together, these findings establish FgSge1 as a critical self-activating regulatory factor that links histone acetylation to higher-order chromatin remodeling, orchestrating mycotoxin gene activation, and providing a framework for understanding epigenetic control of mycotoxin biosynthesis and virulence in fungi.

Post-replicative chromatin accessibility predicts cell fate change.

Eukaryotic DNA is wrapped around octamers of four core histones, forming nucleosomes. Histone post-translational modifications (PTMs) influence chromatin structure and the recruitment of regulatory factors, thereby affecting gene expression and DNA repair, including the response to DNA double-strand breaks (DSBs). Here, we describe a robust chromatin immunoprecipitation protocol combined with micrococcal nuclease digestion and DNA sequencing (MNase-ChIP-seq) to map histone modifications and their genome-wide distribution after the induction of a single DSB by the HO endonuclease in Saccharomyces cerevisiae. We validate the method by detecting changes in histone H3 methylation following HO transcriptional activation and DSB induction. This protocol enables reliable analysis of histone PTMs across mutant strains or stress conditions, supporting studies of chromatin dynamics in yeast.

Spatial controls of homology search.

Hi-C contact maps encode multiscale chromatin folding, yet extracting quantitative and physically interpretable descriptors directly from these matrices remains challenging due to sparsity, depth variation, and the coexistence of loop-, domain-, and compartment-scale interactions. We introduce VECTOR, a graph-spectral framework that quantifies chromatin organization through the von Neumann entropy of the normalized contact-map Laplacian. By constructing distance-banded egographs for each genomic locus, VECTOR provides scale-resolved measures of configurational disorder spanning ∼102-107 bp. Short-range entropy systematically decreases at topological associating domain (TAD) boundaries, whereas long-range entropy captures compartmental reorganization. Entropy scaling reveals shallow exponents (α ≈ 0.04-0.06) and a monotonic compaction-disorder relation linking P(s) scaling to entropy deficits. Polymer simulations with tunable loop strength and A/B contrast confirm predictable spectral and entropic responses to physically meaningful perturbations. VECTOR is reproducible across replicates, robust to resolution and sequencing depth, and remains informative for sparse single-nucleus Hi-C, offering a compact, physics-grounded framework for multiscale chromatin architecture.

The important economic traits of ruminants result from interactions between genetic background and environmental factors, but key traits such as reproductive performance, feed efficiency, disease resistance, and livestock product quality are often not fully explained by DNA sequence variations alone. Increasing evidence suggests that epigenetic regulation serves as a crucial molecular bridge connecting environmental stimuli with changes in gene expression, allowing organisms to exhibit stable and plastic phenotypic differences without altering the DNA sequence. This review provides a structured synthesis of recent research in the field of epigenetics in ruminants, elucidating how multiple layers of epigenetic mechanisms, including DNA methylation, histone modifications, non-coding RNAs, and higher-order chromatin structures, coordinate to regulate growth, development, reproductive performance, metabolic and immune homeostasis, and livestock product traits across different tissues and developmental stages. These epigenetic marks not only demonstrate high responsiveness to nutrition, management, and environmental stressors, but can exhibit context-dependent stability within the same tissue and physiological stage when environmental conditions are comparable, thereby contributing to the regulation of phenotypic plasticity and offering potential value as predictive biomarkers. Furthermore, epigenetic information can supplement our understanding of phenotypic variation in ways that traditional genomic selection methods are unable to capture, offering new data dimensions for the prediction and improvement of low heritability, environmentally sensitive traits. Overall, integrating epigenetic information with genomic selection strategies may improve the accuracy of ruminant trait prediction and enhance environmental adaptability. This integration also offers a conceptual basis and technical pathway for developing more precise and sustainable breeding systems.

Introduction. Chromatin remodelers regulate DNA–nucleosome interactions, shaping chromatin structure, accessibility, and gene expression. Their dysregulation is frequent in cancer and often represents a therapeutic vulnerability. HELLS, a key remodeler, preserves genome stability and controls gene expression in aggressive T-cell lymphomas, including ALK–anaplastic large cell lymphoma (ALCL). Previous studies showed that HELLS knockdown sensitizes ALCL cells to chemotherapy, highlighting its potential as a therapeutic target. This study aimed to define the transcriptional role of HELLS and evaluate its therapeutic relevance in ALCL. Methods. We analyzed formalin-fixed, paraffin-embedded biopsies from diagnostic and relapsed ALCL patients (n= 44) and performed integrated multi-omics analyses (RNA-seq, ChIP-seq, ATAC-seq, and Connectivity Map) alongside functional assays.Results. To explore the transcriptional function of HELLS, we performed HELLS Chromatin immunoprecipitation sequencing (ChIP‐seq) analysis and intersected these data with the list of HELLS-dependent genes we previously identified by performing RNA sequencing. Out of the 729 genes significantly affected by HELLSKD, 64% of the total were directly bound by HELLS based on ChIP-seq analysis. We termed these genes HELLS-direct genes otherwise named as HDGs. These genes belong to pathways known to drive the aggressiveness of ALK-ALCL like JAK/STAT signaling, Rho-GTPase and DNA damage. NCounter profiling in a cohort of 44 ALCL patients (15 ALK⁺, 29 ALK⁻) confirmed their clinical relevance and the association between HELLS and its HDGs in patients. By integrating the effects of the KD of HELLS on the binding profile of RNAPII by ChIP-seq, we observed RNAPII elongation defects in ~60% of HDGs and changes in RNAPII occupancy in ~40% of HDGs. Notably, this last set of genes associated with these promoters belongs to T-cell mediated immunity and PD-L1/PD1 signaling, indicating a previously unrecognized potential role of HELLS in immune-related signaling pathways.These data suggest that, depending on targets, HELLS may foster transcription via two different mechanisms: by promoting RNAPII loading and by facilitating RNAPII elongation along the gene body. To test this hypothesis, we evaluated changes in accessibility by performing ATAC-seq and H3K4me3 profiling, demonstrating that HELLS promotes chromatin accessibility and transcriptional activation at immune-related loci.To investigate the therapeutic potential of HELLS, we performed a drug repurposing analysis based on the HDG signature, which identified PI3K, JAK/STAT, and DNA-PK as synergistic targets. In vitro, HELLS depletion combined with Ruxolitinib (JAKi) or AZD7648 (DNA-PKi) induced synthetic lethality.CONCLUSION: This study shows that HELLS drives immune gene expression via chromatin remodeling in ALK-ALCL, and its inhibition reveals combinatorial vulnerabilities supporting dual targeting of JAK/STAT or DNA-PK pathways.

Genome replication is temporally regulated during S phase, with specific genomic regions replicating at defined times in a process that is known as replication timing (RT). Based on 3D cytology in replicating nuclei, we previously proposed a model in which maize euchromatin is subdivided into subcompartments distinguished by chromatin condensation and RT. However, whether this compartmentalization reflects a general nuclear architecture that persists throughout the cell cycle was unclear. To test this model, we conducted two orthogonal assays-Hi-C for genome-wide interaction data and 3D FISH for direct visualization of chromatin organization in maize (Zea mays L.). Hi-C analyses revealed distinct patterns of early-S regions exhibited negative insulation scores with long-range contacts, whereas middle-S regions showed the opposite. Early-S regions showed the strongest correlation with epigenomic signatures of open, transcriptionally active chromatin. 3D oligo FISH painting confirmed that early-S and middle-S replicating regions occupy adjacent but largely non-overlapping nucleoplasmic sub-territories throughout interphase stages, including G1. Together, our findings redefine the maize euchromatin "A" compartment as two spatially distinct subcompartments derived from high-frequency RT transitions between early and middle S along the linear genome. These findings have implications for chromatin-templated processes and underscore the importance of RT as a defining feature of genome organization.

Double-strand breaks (DSBs) threaten genomic stability and need immediate attention from DNA damage response (DDR) machinery involved in homologous recombination (HR) or nonhomologous end joining (NHEJ). DDR in heterochromatin is challenging owing to the distinct chromatin organization. Heterochromatin protein 1 (HP1) isoforms are central to heterochromatin structure and have been implicated in DDR. Mammalian HP1 has three isoforms, HP1α, HP1β, and HP1γ, which possess significant homology and yet have distinct functions. HP1α is the only isoform known to undergo liquid-liquid phase separation mediated by phosphorylation on the N-terminal extension (NTE). We show that the minute-scale dynamics of HP1α and HP1β differ dramatically and differentially influence the recruitment of HR vs. NHEJ factors at sites of laser-induced clustered DSBs. Perturbing HP1α phosphorylation impairs HR factor recruitment and reduces HR efficiency. Our study provides a potential link between phase separation and DDR-centric roles of HP1α and hints at spatial partitioning of repair pathways in response to damage in heterochromatin.

Chromatin organization, through the assembly of DNA with histones and the folding of nucleosome chains, regulates DNA accessibility for transcription, DNA replication and repair. Although models derived from in vitro studies have proposed distinct nucleosome chain geometries, the organization of chromatin within the crowded cell nucleus remains elusive. Using cryo-electron tomography of thin vitreous sections, we directly observed the path of nucleosomal and linker DNA in situ from a flash-frozen organism - Drosophila embryos. We quantified linker length and curvature, characterizing an irregular zig-zag chromatin-folding motif, with a low degree of linker bending. Nucleosome conformations could be identified on individual particles in favorable orientations without structure averaging. Additionally, we observed particles that accommodate a number of DNA gyres ranging from less than one to up to three, which resemble previously proposed non-octameric nucleosomal particles with variable DNA wrapping.

Lysyl oxidase (LOXs) are copper-dependent enzymes traditionally known for catalyzing the cross-linking of collagen and elastin, thereby ensuring extracellular matrix (ECM) stability. However, growing evidence reveals that their biological functions extend far beyond ECM remodeling. This review highlights the diverse roles of the LOX family, comprising LOX, LOXL1, LOXL2, LOXL3, and LOXL4, in tissue repair, vascular remodeling, inflammation, and cancer. Each isoform exhibits unique structural domains, regulatory pathways, and functional interactions with signaling cascades such as TGF-β, PDGF, VEGF, and HIF-1α. LOXs are essential for wound healing, coordinating ECM synthesis and cross-linking during different phases of tissue regeneration. Their expression is tightly modulated by inflammatory cytokines, and their dysregulation has been implicated in pathological fibrosis and impaired tissue repair. In cancer, LOXs contribute to epithelial-to-mesenchymal transition (EMT), cell invasion, and metastasis through both enzymatic and non-enzymatic mechanisms, including intracellular signaling, Snail1 stabilization, and cytoskeletal modulation. They also influence angiogenesis by regulating VEGF expression and promoting endothelial cell activation via PDGFRβ-AKT signaling. Intracellular and nuclear functions further expand their impact on gene regulation and chromatin structure. Given their involvement in matrix dynamics, mechanotransduction, and cell fate determination, LOXs emerge as key players in both physiological and pathological contexts. Understanding their multifactorial roles opens potential avenues for therapeutic targeting in cancer, fibrosis, and chronic inflammatory diseases.

Post-Translational Modifications Orchestrate Repair of Trapped Topoisomerase-Induced DNA Breaks via TDP1 and TDP2.

SMARCA4 deficiency in glioblastoma: Mitochondrial transfer from MSCs and the clinical dilemma in targeting the tumor microenvironment.

Epigenetic enzyme inhibitors targeting DNA, histone, and RNA methylation: Mechanisms and therapeutic applications in cancer.

The mouse model of induced sperm DNA damage caused by polystyrene microplastics exhibited distinct transcriptomic and proteomic features.

Single-cell Hi-C (scHi-C) provides unprecedented insight into 3D genome organization, but its sparse and noisy data pose challenges in accurately detecting A/B compartments, which are crucial for understanding chromatin structure and gene regulation. We presented scDIAGRAM, a data-driven method for annotating A/B compartments in single cells using direct statistical modeling and graph community detection. Unlike existing approaches, scDIAGRAM infers chromatin compartments directly from individual scHi-C matrix without imputation or external reference features, and subsequently assigns A/B labels using conventional genomic annotations. Accuracy and robustness of scDIAGRAM were illustrated through simulated scHi-C datasets and a human cell line. We applied scDIAGRAM to real scHi-C datasets from the mouse brain cortex, mouse embryonic development, and human acute myeloid leukemia, demonstrating its ability to capture compartmental shifts associated with transcriptional variation. This robust framework offers new insights into the functional roles of chromatin compartments at single-cell resolution across various biological contexts.

From bench to bedside: translating the histone deacetylase inhibitors into prostate cancer therapies.