bims-crepig 2025-07-06 papers

bims-crepig

Biomed News

on Chromatin regulation and epigenetics in cell fate and cancer

Issue of 2025–07–06
39 papers selected by
Connor Rogerson, University of Cambridge

Extrusion fountains are restricted by WAPL-dependent cohesin release and CTCF barriers.
Nucleosome organization of mouse embryos during pre-implantation development.
CTCF binding landscape is shaped by the epigenetic state of the N-terminal nucleosome in relation to CTCF motif orientation.
Klf5-adjacent super-enhancer functions as a 3D genome structure-dependent transcriptional driver to safeguard ESC identity.
Cold-induced nucleosome dynamics linked to silencing of Arabidopsis FLC.
Echs1-mediated histone crotonylation facilitates zygotic genome activation and expression of repetitive elements in early mammalian embryos.
Rhabdomyosarcoma fusion oncoprotein initially pioneers a neural signature in vivo.
Characterizing DNA recognition preferences of transcription factors using global couplings and high-throughput sequencing.
Single-cell long-read Hi-C, scNanoHi-C2, details 3D genome reorganization in embryonic-stage germ cells.
Structure of a transcribing Pol II-DSIF-SPT6-U1 snRNP complex.
Deciphering the role of cis-regulatory elements and TFAP2C in the activation of zygotic Sox2 expression in mouse preimplantation embryos.
Single-cell Micro-C profiles 3D genome structures at high resolution and characterizes multi-enhancer hubs.
Regulation of somatic hypermutation by higher-order chromatin structure.
Synergistic DNA and RNA binding of the Hox transcription factor Ultrabithorax coordinates splicing and shapes in vivo homeotic functions.
H3K79 methylation and H3K36 trimethylation synergistically regulate gene expression in pluripotent stem cells.
EBNA leader protein orchestrates chromatin architecture remodeling during Epstein-Barr virus-induced B cell transformation.
Range extender mediates long-distance enhancer activity.
Self-amplifying NRF2-EZH2 epigenetic loop converts KRAS-initiated progenitors to invasive pancreatic cancer.
Differential oligomerization regulates PHF13 chromatin affinity and function.
Zincore, an atypical coregulator, binds zinc finger transcription factors to control gene expression.
Adversarial attack of sequence-free enhancer prediction identifies chromatin architecture.
TEAD-targeting small molecules induce a cofactor switch to regulate the Hippo pathway.
A dual enhancer-attenuator element ensures transient Cdx2 expression during mouse posterior body formation.
Epigenetic profiling identifies markers of endocrine resistance and therapeutic options for metastatic castration-resistant prostate cancer.
A new IDH-independent hypermethylation phenotype is associated with astrocyte-like cell state in glioblastoma.
The length and strength of compartmental interactions are modulated by condensin II activity.
Chromosomal tethering and mitotic transcription promote ecDNA nuclear inheritance.
Deciphering gene regulatory programs in mouse embryonic skin through single-cell multiomics analysis.
Reconciling competing models on the roles of condensates and soluble complexes in transcription factor function.
Mapping the nuclear landscape with multiplexed super-resolution fluorescence microscopy.
Spatial profiling of chromatin accessibility in formalin-fixed paraffin-embedded tissues.
O-GlcNAcylation of FOXK1 co-opts BAP1 to orchestrate the E2F pathway and promotes oncogenesis.
Histone lysine methyltransferases MLL3 and MLL4 direct gene expression to produce platelets efficiently.
Deep learning deciphers the related role of master regulators and G-quadruplexes in tissue specification.
BMAL1 and ARNT enable circadian HIF2α responses in clear cell renal cell carcinoma.
Disruption of TAD hierarchy promotes LTR co-option in cancer.
Precision-edited histone tails disrupt polycistronic gene expression controls in trypanosomes.
De novo design of insulated cis-regulatory elements based on deep learning-predicted fitness landscape.
MORC2 is a phosphorylation-dependent DNA compaction machine.

Nucleic Acids Res. 2025 Jun 20. pii: gkaf549. [Epub ahead of print]53(12):

Extrusion fountains are restricted by WAPL-dependent cohesin release and CTCF barriers.

Ning Qing Liu, Mikhail Magnitov, Marijne M G A Schijns, Tom van Schaik, Hans Teunissen, Bas van Steensel, Elzo de Wit.

Interphase chromosomes are mainly shaped by loop extrusion and compartmentalisation mechanisms. However, their temporal component and cause-effect relationships remain largely unknown. In this study, we use acute degradation of WAPL, CTCF and cohesin in mouse embryonic stem cells to investigate the dynamics of loop extrusion and its relationship to compartmentalisation. Stabilisation of cohesin on chromatin by depletion of WAPL results in the formation of extended loops and promotes looping between non-convergent CTCF sites. Loss of WAPL also results in a rapid decrease in compartmentalisation, which is reversed by subsequent removal of cohesin, directly demonstrating the opposite role of extrusion on compartmentalisation. Using combined depletion of WAPL and CTCF, we identify fountains, a feature of chromosome organisation that emanates from enhancer regions and exhibits strong cohesin binding. Fountains form rapidly after mitosis and early in mammalian development. Cohesin depletion confirms that fountains are cohesin dependent, and their disruption leads to the downregulation of fountain-proximal genes, suggesting a role in gene regulation. Taken together, by exploiting the temporal precision of acute protein depletion, our study reveals fountains as an extrusion-mediated, fast-forming feature of 3D genome organisation.

DOI: https://doi.org/10.1093/nar/gkaf549
Sci Rep. 2025 Jul 01. 15(1): 21894

Nucleosome organization of mouse embryos during pre-implantation development.

Jitao Zeng, Xiaojuan Tu, Siming Qu, Xichan Chen, Ying Wang, Xiaona Liu, Enchuan Kang, Yi Tian, Qianming Xiang, Binbin Lai, Weiwu Gao, Bing Ni, Wei He.

  Sexual reproduction begins with sperm-oocyte fusion to form a zygote, where chromatin undergoes dramatic reorganization to establish totipotency. Although nucleosomes- the basic units of eukaryotic chromatin and key epigenetic regulators- are extensively remodeled during early embryogenesis, their dynamic repositioning mechanisms and biological implications remain unclear. Here, we employed single-cell MNase sequencing (scMNase-seq) to map genome-wide nucleosome positioning and chromatin accessibility in individual mammalian embryos. We found that nucleosome positioning mirrored somatic cell patterns until the 4-cell stage, with nucleosome depletion and phasing at CTCF sites not fully established until morula formation. By integrating H3K4me3 localization and transcriptomic data, we revealed that nucleosome sparsity at transcription start sites (TSS) and flanking regions correlated with expression levels of genes critical for preimplantation development. Notably, these nucleosome-depleted regions likely serve as regulatory hubs influencing histone modification dynamics. Our study systematically delineates nucleosome reorganization principles during mammalian embryogenesis and provides a high-resolution resource for understanding chromatin remodeling in early development.

Keywords:  Chromatin organization; Embryos; H3K4me3; Nucleosome position; scMNase-seq

DOI:  https://doi.org/10.1038/s41598-025-05642-5
Nucleic Acids Res. 2025 Jun 20. pii: gkaf587. [Epub ahead of print]53(12):

CTCF binding landscape is shaped by the epigenetic state of the N-terminal nucleosome in relation to CTCF motif orientation.

Md Tajmul, Dharmendra Nath Bhatt, Luminita Ruje, Emma Price, Yon Ji, Dmitri Loukinov, Vladimir B Teif, Victor V Lobanenkov, Elena M Pugacheva.

CTCF binding sites serve as anchors for the 3D chromatin architecture in vertebrates. The functionality of these anchors is influenced by the residence time of CTCF on chromatin, which is determined by its binding affinity and its interactions with nucleosomes and other chromatin-associated factors. In this study, we demonstrate that CTCF occupancy is driven by CTCF motifs, strategically positioned at the entry sides of a well-positioned nucleosome, such that, upon binding, the N-terminus of CTCF is oriented towards the nucleosome. We refer to this nucleosome as the CTCF priming nucleosome (CPN). Our analyses suggest that CTCF can more easily displace the CPN if the nucleosome is not marked by CpG methylation or repressive histone modifications. Under these permissive conditions, the N-terminus of CTCF recruits SMARCA5 to reposition the CPN downstream, thereby creating nucleosome-free regions that enhance CTCF occupancy and cohesin stalling. In contrast, when CPNs carry repressive epigenetic marks, CTCF binding is weaker, with no nucleosome displacement or chromatin opening, and cohesin is retained less effectively at CTCF binding sites. We propose that the epigenetic status of CPNs shapes cell-specific CTCF binding patterns, ensuring the maintenance of chromatin architecture throughout the cell cycle.

DOI: https://doi.org/10.1093/nar/gkaf587
Nat Commun. 2025 Jul 01. 16(1): 5540

Klf5-adjacent super-enhancer functions as a 3D genome structure-dependent transcriptional driver to safeguard ESC identity.

Guangsong Su, Bohan Chen, Yingjie Song, Qingqing Yin, Wenbin Wang, Xueyuan Zhao, Sibo Fan, Jie Lian, Dongqing Li, Jinfang Bi, Peng Li, Zhongfang Zhao, Lei Zhang, Jiandang Shi, Wange Lu.

Cell-specific super-enhancers (SEs) and master transcription factors (TFs) dynamically remodel embryonic stem cell (ESC) fate, yet their regulatory interplay remains unclear. By integrating multi-omics data (H3K27ac, Hi-C, scRNA-seq) across ESC states, we identified SEs interacting with master TFs, exemplified by the Klf5-adjacent SE (K5aSE). K5aSE deletion impaired proliferation, differentiation, and Klf5 expression, partially rescued by KLF5 reintroduction. Despite phenotypic similarities between Klf5-KO and K5aSE-KO ESCs, scRNA-seq of embryoid bodies revealed distinct differentiation trajectories, suggesting K5aSE targets beyond Klf5. High-throughput 3D genome screening demonstrated K5aSE activates four distal genes via chromatin looping. CRISPRa-mediated activation of these targets rescued K5aSE-KO phenotypes and uncovered their regulatory roles. Furthermore, CTCF depletion disrupted topologically associated domains (TADs) near K5aSE, suppressing Klf5 and target gene expression, indicating CTCF-mediated TADs sustain K5aSE activity. Our study unveils a 3D genome-dependent mechanism by which SEs govern ESC identity through coordinated TF interaction and multi-gene regulation.

DOI: https://doi.org/10.1038/s41467-025-60389-x
Nat Commun. 2025 Jul 01. 16(1): 5550

Cold-induced nucleosome dynamics linked to silencing of Arabidopsis FLC.

Miguel Montez, Danling Zhu, Jan Huertas, M Julia Maristany, Bas Rutjens, Mathias Nielsen, Rosana Collepardo-Guevara, Caroline Dean.

Temperature influences nucleosome dynamics, and thus chromatin, to regulate gene expression. Such mechanisms underlie the epigenetic silencing of Arabidopsis FLOWERING LOCUS C (FLC) by prolonged cold. Here, we show a temperature-dependent transition in local chromatin structure at the H3K27me3 nucleation region, from a modality active for transcription to a state that can be Polycomb silenced. In vivo chromatin measurements and coarse-grained simulations at near-atomistic resolution show that the active transcription state is characterised by a highly dynamic nucleosome arrangement that exposes the FLC transcription start site (TSS). Cold exposure then changes the chromatin by reducing nucleosome dynamics and re-positioning the + 1 nucleosome, leading to transcriptional repression. This local chromatin transition partially depends on VERNALIZATION1 (VRN1), a non-sequence-specific DNA-binding protein. Loss of VRN1 results in hyperaccumulation of H2A.Z, more dynamic nucleosomes and an inability to accumulate H2Aub and H3K27me3. Our work highlights how local nucleosome dynamics link to chromatin structure transitions to integrate temperature inputs into epigenetic switching mechanisms in plants.

DOI: https://doi.org/10.1038/s41467-025-60735-z
Nat Commun. 2025 Jul 01. 16(1): 5630

Echs1-mediated histone crotonylation facilitates zygotic genome activation and expression of repetitive elements in early mammalian embryos.

Yong-Feng Wang, Yu-Ting Wan, Qian-Rong Qi, Qing Tian, Xin-Mei Liu, Qing-Zhen Xie, Ying Yin, Li-Quan Zhou.

Histone crotonylation, a conserved post-translational histone modification, plays a crucial role in transcriptional regulation. However, its function in early embryonic development remains largely unexplored. Here, we perform genome-wide mapping of histone crotonylation in mouse and human early embryos. Our analysis reveals that histone crotonylation is highly enriched at promoter regions and exhibits distinct dynamic patterns throughout embryogenesis. Notably, strong histone crotonylation signals are observed at the mouse 2-cell and human 4-to-8-cell stages, coinciding with zygotic genome activation. In mice, Echs1 knockdown in oocytes, which suppresses histone crotonylation, results in developmental arrest at the 2-cell stage. Further investigation demonstrates that reduced histone crotonylation impairs transcriptional activity at zygotic genome activation genes, retrotransposon elements, and ribosomal DNA loci. Moreover, early embryos from aged female mice exhibit significantly diminished histone crotonylation, while supplementation with exogenous sodium crotonate enhances blastocyst formation. Collectively, our findings establish histone crotonylation as a key regulatory mechanism in early mammalian embryogenesis by facilitating transcriptional activation of zygotic genome activation genes and repetitive elements.

DOI: https://doi.org/10.1038/s41467-025-60565-z
Cell Rep. 2025 Jul 02. pii: S2211-1247(25)00694-1. [Epub ahead of print]44(7): 115923

Rhabdomyosarcoma fusion oncoprotein initially pioneers a neural signature in vivo.

Jack Kucinski, Alexi Tallan, Cenny Taslim, Andrew M Vontell, Katherine M Silvius, Meng Wang, Matthew V Cannon, Benjamin Z Stanton, Genevieve C Kendall.

  Fusion-positive rhabdomyosarcoma is an aggressive pediatric cancer molecularly characterized by arrested myogenesis. The defining genetic driver, PAX3::FOXO1, encodes a chimeric gain-of-function transcription factor. An incomplete understanding of the in vivo chromatin regulatory mechanisms of PAX3::FOXO1 has hindered therapeutic development. Here, we establish a PAX3::FOXO1 zebrafish injection model and a semi-automated ChIP-seq normalization strategy to evaluate how PAX3::FOXO1 initially interfaces with and modulates chromatin in a developmental context. We find that PAX3::FOXO1 interacts with inaccessible chromatin through partial/homeobox motif recognition consistent with pioneering activity. However, PAX3::FOXO1-genome binding through a composite paired box/homeobox motif alters chromatin accessibility and redistributes H3K27ac to activate neural transcriptional programs. We uncover neural signatures that are highly representative of clinical rhabdomyosarcoma gene expression programs that are enriched following chemotherapy. Overall, we identify partial/homeobox motif recognition as a key mode for PAX3::FOXO1 pioneer function and identify neural signatures as a potentially critical PAX3::FOXO1 tumor initiation event.

Keywords:  CP: Cancer; chromatin biology; functional epigenomics; fusion oncogenes; pediatric sarcoma; pioneer factors; rhabdomyosarcoma; zebrafish cancer models

DOI:  https://doi.org/10.1016/j.celrep.2025.115923
Nucleic Acids Res. 2025 Jun 20. pii: gkaf592. [Epub ahead of print]53(12):

Characterizing DNA recognition preferences of transcription factors using global couplings and high-throughput sequencing.

Qin Zhou, Jose Alberto de la Paz, Alexander D Stanowick, Xingcheng Lin, Faruck Morcos.

DNA-transcription factor (TF) interactions are essential for gene regulation. Fully characterizing TF recognition specificities and identifying their genomic binding targets are important to understand TF function and regulatory networks. Recently, high-throughput sequencing technology HT-SELEX (high-throughput systematic evolution of ligands by exponential enrichment) has been used to measure hundreds of TFs, providing massive datasets that comprise TF binding preferences. However, there is a need to develop comprehensive computational modeling to fully extract and characterize critical TF binding preferences and fail to distinguish genome-wide binding targets. In this study, we developed a global pairwise model called DCA-Scapes trained with experimental HT-SELEX data. Our approach uncovered high-resolution TF recognition specificity landscapes, enabled the prediction of in vivo binding sequences, and was validated with ChIP-seq (ChIP sequencing) data. In addition, the DCA-Scapes model was utilized to refine the locations of binding regions and accurately identify the binding sites within the ChIP-seq enriched peaks. Moreover, we extended our model to cover the entire human genome, uncovering potential TF target sites that exhibit tissue-specific TF recognition across various cellular environments.

DOI: https://doi.org/10.1093/nar/gkaf592
Nat Struct Mol Biol. 2025 Jul 04.

Single-cell long-read Hi-C, scNanoHi-C2, details 3D genome reorganization in embryonic-stage germ cells.

Jiansen Lu, Wen Li, Fuchou Tang.

During mouse development, embryonic-stage germ cells (EGCs) make crucial fate decisions, with female EGCs embarking on meiosis whereas male EGCs enter mitotic arrest until birth. Despite increasing understanding of the reprogramming of epigenetic modifications, the dynamics of three-dimensional (3D) genome structures within individual EGCs remains elusive. Here we present a single-cell input, long-read Hi-C method, termed scNanoHi-C2. We use scNanoHi-C2 to systematically dissect the dynamics of EGC chromatin structures. We find that, despite changes in autosomes similar to spermatogenesis, the X chromosomes of female EGCs show enhanced specific interactions between B compartments. By reconstructing 3D genome models, we observe dynamic chromosome positioning during meiosis, showing that the neighborhood between nonhomologous chromosomes of EGCs is relatively random. Simultaneously, transposable elements undergo dramatic chromatin reorganization and display an asymmetric distribution of Alu/B2 elements around meiotic topologically associated domain boundaries. Moreover, we find that high-order interactions in EGCs at the mitosis stage are mainly enriched in the B compartment, whereas, after the mitosis-to-meiosis transition, enriched high-order interactions shift to refined A compartments, to potentially promote meiotic-specific transcription programs during global genomic condensation. We also reveal an unexpected chromatin structure in mitotic-arrested male EGCs distinct from the previously assumed G0 status, which may prime the unique genome structure for subsequent spermatogenesis. Altogether, our study highlights the potential of scNanoHi-C2 and reveals key features of the chromatin structure reprogramming in EGCs.

DOI: https://doi.org/10.1038/s41594-025-01604-7
Nat Commun. 2025 Jul 01. 16(1): 5823

Structure of a transcribing Pol II-DSIF-SPT6-U1 snRNP complex.

Luojia Zhang, Christopher Batters, Shintaro Aibara, Yuliya Gordiyenko, Kristina Žumer, Jana Schmitzová, Kerstin Maier, Patrick Cramer, Suyang Zhang.

In eukaryotic cells, splicing occurs predominantly co-transcriptionally, enhancing splicing efficiency and fidelity while introducing an additional layer of regulation over gene expression. RNA polymerase II (Pol II) facilitates co-transcriptional splicing by recruiting the U1 small nuclear ribonucleoprotein particle (U1 snRNP) to the nascent transcripts. Here, we report the cryo-electron microscopy structure of a transcribing Pol II-U1 snRNP complex with elongation factors DSIF and SPT6. In addition, our biochemical analysis reveals that the phosphorylated Pol II carboxyl-terminal domain and SPT6 interact directly with U1 snRNP proteins, facilitating its recruitment to the elongation complex. This multivalent interaction between U1 snRNP and the transcription elongation complex may both allow efficient spliceosome assembly and ensure transcription processivity.

DOI: https://doi.org/10.1038/s41467-025-60979-9
Development. 2025 Jul 02. pii: dev.204626. [Epub ahead of print]

Deciphering the role of cis-regulatory elements and TFAP2C in the activation of zygotic Sox2 expression in mouse preimplantation embryos.

Jaehwan Kim, Chad S Driscoll, Lijia Li, Catherine A Wilson, Wei Xie, Jason G Knott.

  Cell-fate decisions in preimplantation embryos require the coordinated expression of pluripotency and lineage-specific transcription factors. SOX2 represents the first pluripotency regulator whose expression is restricted to the inside cells of mouse preimplantation embryos. However, the genetic mechanisms that activate the expression of zygotic Sox2 are poorly understood. Here we report that Sox2 expression in mouse embryos is controlled by the actions of key cis-regulatory elements, including a proximal promoter and super enhancer. We show that TFAP2C, a key trophoblast lineage regulator, binds to the Sox2 proximal promoter to activate its expression. Lastly, we provide evidence that TFAP2C and the HIPPO signaling pathway cooperatively regulate Sox2 expression. In summary, this work has important implications in understanding how conventional trophoblast transcription factors, such as TFAP2C, contribute to the activation of early pluripotency genes to facilitate divergent cellular states that support lineage formation.

Keywords:   Sox2 regulatory regions; HIPPO signaling; Pluripotency; Preimplantation embryo; TFAP2C

DOI:  https://doi.org/10.1242/dev.204626
Nat Genet. 2025 Jul 02.

Single-cell Micro-C profiles 3D genome structures at high resolution and characterizes multi-enhancer hubs.

Honggui Wu, Jiankun Zhang, Longzhi Tan, Xiaoliang Sunney Xie.

In animal genomes, regulatory DNA elements called enhancers govern precise spatiotemporal gene expression patterns in specific cell types. However, the spatial organization of enhancers within the nucleus to regulate target genes remains poorly understood. Here we report single-cell Micro-C (scMicro-C), a micrococcal nuclease-based three-dimensional (3D) genome mapping technique with an improved spatial resolution of 5 kb, and identified a specialized 3D enhancer structure termed 'promoter-enhancer stripes (PESs)', connecting a gene's promoter to multiple enhancers. PES are formed by cohesin-mediated loop extrusion, which potentially brings multiple enhancers to the promoter. Further, we observed the prevalence of multi-enhancer hubs on genes with PES within single-cell 3D genome structures, wherein multiple enhancers form a spatial cluster in association with the gene promoter. Through its improved resolution, scMicro-C elucidates how enhancers are spatially coordinated to control genes.

DOI: https://doi.org/10.1038/s41588-025-02247-6
Mol Cell. 2025 Jun 20. pii: S1097-2765(25)00503-9. [Epub ahead of print]

Regulation of somatic hypermutation by higher-order chromatin structure.

Ursula E Schoeberl, Johanna Fitz, Maximilian von der Linde, Renan Valieris, Bernd Bauer, Daniel Malzl, Kimon Froussios, Julia Costea, Franziska Schmidt, Jeremy Pflaum, Bianca Bartl, Johanna Albrecht, Eva-Maria Wiedemann, Adriana Cantoran Garcia, Mihaela Peycheva, Marta Rizzi, Anton Goloborodko, Israel Tojal da Silva, Rushad Pavri.

  The generation of protective antibodies by somatic hypermutation (SHM) is essential for antibody maturation and adaptive immunity. SHM involves co-transcriptional mutagenesis of immunoglobulin variable (V) regions regulated by enhancers located hundreds of kilobases away. How 3D chromatin topology affects SHM is poorly understood. Here, we measure higher-order interactions on single alleles of the human immunoglobulin heavy-chain locus (IGH) using Tri-C. We find that SHM is underpinned by a multiway hub wherein the V region is proximal to all enhancers. Cohesin-mediated loop extrusion is dispensable for IGH transcription and hub architecture. Transcription and mutagenesis of IGH switch regions, which are necessary for antibody class-switch recombination, create new chromatin loops that can form without cohesin. However, these additional loops do not compromise hub integrity, V region transcription, or SHM. Thus, antibody maturation occurs within a multiway hub accommodating several gene-enhancer loops in which transcription and mutagenesis of different segments occur non-competitively.

Keywords:  PRO-seq; Tri-C; antibody maturation; chromatin architecture; chromatin loop extrusion; class switch recombination; cohesin; human immunoglobulin heavy-chain locus; somatic hypermutation; transcription hub

DOI:  https://doi.org/10.1016/j.molcel.2025.06.003
Nucleic Acids Res. 2025 Jun 20. pii: gkaf602. [Epub ahead of print]53(12):

Synergistic DNA and RNA binding of the Hox transcription factor Ultrabithorax coordinates splicing and shapes in vivo homeotic functions.

Constanza Blanco, Wan Xiang, Panagiotis Boumpas, Elzéar Buvry, Maily Scorcelletti, Ashley Suraj Hermon, Jiemin Wong, Samir Merabet, Julie Carnesecchi.

The dual interaction of many transcription factors (TFs) with both DNA and RNA is an underexplored issue that could fundamentally reshape our understanding of gene regulation. We address this central issue by investigating the RNA binding activity of the Drosophila Hox TF Ultrabithorax (Ubx) in alternative splicing and morphogenesis. Relying on molecular and genetic interactions, we uncover a homodimerisation-dependent mechanism by which Ubx regulates splicing. Notably, this mechanism enables the decoupling of Ubx-DNA and -RNA binding activity in splicing. We identify a critical residue for Ubx-RNA binding and demonstrate the essential role of Ubx-RNA binding ability for its homeotic functions. Overall, we uncover a unique mechanism for Ubx-mediated splicing and underscore the critical contribution of synergistic DNA/RNA binding for its morphogenetic functions. These findings advance our understanding of co-transcriptional regulation and highlight the significance of TF-DNA/RNA synergistic function in shaping gene regulatory networks in living organisms.

DOI: https://doi.org/10.1093/nar/gkaf602
Sci Adv. 2025 Jul 04. 11(27): eadt8765

H3K79 methylation and H3K36 trimethylation synergistically regulate gene expression in pluripotent stem cells.

Emmalee W Cooke, Cheng Zeng, Suza Mohammad Nur, Yunbo Jia, Aileen Huang, Jiwei Chen, Peidong Gao, Fei Xavier Chen, Fulai Jin, Kaixiang Cao.

Metazoan nucleosomes harboring H3K79 methylation (H3K79me) deposited by the methyltransferase DOT1L (disruptor of telomeric silencing 1-like) decorate actively transcribed genes. While DOT1L regulates transcription and the pathogenesis of leukemia and neurological disorders, the role of H3K79me remains elusive. Here, we reveal a functional synergism between H3K79me and H3K36 trimethylation (H3K36me3) in regulating gene expression and cellular differentiation. Simultaneous catalytic inactivation of DOT1L and the H3K36 methyltransferase SETD2 (SET domain containing 2) leads to hyperactive transcription and failures in neural differentiation. H3K79me/H3K36me3 loss causes increased transcription elongation, gained chromatin accessibility at a group of enhancers, and increased recruitment of TEAD4 (TEA domain transcription factor 4) and its coactivator YAP1 (Yes-associated protein 1) to these enhancers. Furthermore, YAP-TEAD inhibition restores the expression levels of genes hyperactivated by H3K79me/H3K36me3 loss. Together, we demonstrate a synergism of H3K79me and H3K36me3 in regulating transcription and cell fate transition, unveil the underlying mechanisms, and provide insight into targeting diseases driven by misregulation/mutations of DOT1L and/or SETD2.

DOI: https://doi.org/10.1126/sciadv.adt8765
Nucleic Acids Res. 2025 Jun 20. pii: gkaf629. [Epub ahead of print]53(12):

EBNA leader protein orchestrates chromatin architecture remodeling during Epstein-Barr virus-induced B cell transformation.

Davide Maestri, Lisa B Caruso, Jana M Cable, Rachel Sklutuis, Sarah Preston-Alp, Robert E White, Micah A Luftig, Italo Tempera.

Epstein-Barr virus Nuclear Antigen Leader Protein (EBNA-LP) plays a pivotal role in the transformation of B cells by Epstein-Barr virus (EBV), functioning independently of EBNA2 to regulate chromatin architecture and gene expression. Our study reveals that EBNA-LP binds to chromatin regions distinct from EBNA2 and facilitates the formation of long-distance chromatin loops by interacting with the cellular factor YY1. This interaction reconfigures the three-dimensional structure of the host genome, enhancing the integrity of topologically associating domains (TADs) and promoting the interaction between enhancers and promoters within these domains. In EBV-infected B cells, EBNA-LP strengthens YY1-mediated chromatin loops within TADs, which helps maintain stable regulatory programs essential for B cell transformation. Notably, EBNA-LP is crucial for establishing EBV-induced enhancers, yet it is not required for their maintenance once formed. Additionally, our data suggest a compensatory increase in CTCF binding in the absence of EBNA-LP, leading to more promiscuous chromatin interactions between TADs and a reduced TAD insulation at their boundaries. These findings provide new insights into the molecular mechanisms by which EBV reshapes the host genome chromatin architecture to support B cell transformation and highlight potential therapeutic targets for disrupting EBV-driven oncogenesis.

DOI: https://doi.org/10.1093/nar/gkaf629
Nature. 2025 Jul 02.

Range extender mediates long-distance enhancer activity.

Grace Bower, Ethan W Hollingsworth, Sandra H Jacinto, Joshua A Alcantara, Benjamin Clock, Kaitlyn Cao, Mandy Liu, Adam Dziulko, Ana Alcaina-Caro, Qianlan Xu, Dorota Skowronska-Krawczyk, Javier Lopez-Rios, Diane E Dickel, Anaïs F Bardet, Len A Pennacchio, Axel Visel, Evgeny Z Kvon.

Although most mammalian transcriptional enhancers regulate their cognate promoters over distances of tens of kilobases, some enhancers act over distances in the megabase range1. The sequence features that enable such long-distance enhancer-promoter interactions remain unclear. Here we used in vivo enhancer-replacement experiments at the mouse Shh locus to show that short- and medium-range limb enhancers cannot initiate gene expression at long-distance range. We identify a cis-acting element, range extender (REX), that confers long-distance regulatory activity and is located next to a long-range limb enhancer of Sall1. The REX element has no endogenous enhancer activity. However, addition of the REX to other short- and mid-range limb enhancers substantially increases their genomic interaction range. In the most extreme example observed, addition of REX increased the range of an enhancer by an order of magnitude from its native 73 kb to 848 kb. The REX element contains highly conserved [C/T]AATTA homeodomain motifs that are critical for its activity. These motifs are enriched in long-range limb enhancers genome-wide, including the ZRS (zone of polarizing activity (ZPA) regulatory sequence), a benchmark long-range limb enhancer of Shh2. The ZRS enhancer with mutated [C/T]AATTA motifs maintains limb activity at short range, but loses its long-range activity, resulting in severe limb reduction in knock-in mice. In summary, we identify a sequence signature associated with long-range enhancer-promoter interactions and describe a prototypical REX element that is necessary and sufficient to confer long-distance activation by remote enhancers.

DOI: https://doi.org/10.1038/s41586-025-09221-6
Nat Cancer. 2025 Jun 30.

Self-amplifying NRF2-EZH2 epigenetic loop converts KRAS-initiated progenitors to invasive pancreatic cancer.

Laura Antonucci, Na Li, Angeles Duran, Isidoro Cobo, Chiara Nicoletti, Kosuke Watari, Shuvro Prokash Nandi, Feng Zhu, Yongmei Zhao, Irene Riahi, Motoyuki Tsuda, Vidhi M Shah, Terry Morgan, Trent Waugh, Luca Caputo, Yuan Liu, Alexandra Rundberg Nilsson, Hongxu Xian, Jelena Todoric, Li Gu, Elsa Sanchez-Lopez, Guido Eibl, Emily A Vucic, Michal Krawczyk, Qianlan Xu, Andrew M Lowy, Georgia Hatzivassiliou, Merone Roose-Girma, Dorota Skowronska-Krawczyk, David A Scott, Dafna Bar-Sagi, Pablo Tamayo, Ying Wu, Rosalie C Sears, Christopher K Glass, Ludmil B Alexandrov, Pier Lorenzo Puri, David W Dawson, Yinling Hu, Maria T Diaz-Meco, Jorge Moscat, Michael Karin.

Pancreatic ductal adenocarcinoma (PDAC) emerges from mutant KRAS-harboring but dormant low-grade pancreatic intraepithelial neoplasia (PanIN). To examine the role of oxidative stress, a putative PDAC risk factor, we established an organoid-based transformation system. Although the prototypic oxidant H2O2 induced organoid transformation, its effect was nonmutational and was mediated by the oxidant-responsive transcription factor NRF2, which induced the histone methyltransferase EZH2. Congruently, nonoxidizing NRF2 activators triggered organoid malignant conversion through NRF2 and EZH2, establishing a hitherto unknown epigenetic mechanism underlying PanIN-to-PDAC progression. While NRF2 induced EZH2 gene transcription in mouse and human PDAC, EZH2, a general repressor, coactivated transcription of NRF2-encoding NFE2L2 and interacted with other transcription factors to induce genes that sustain PDAC metabolic demands. The self-amplifying NRF2-EZH2 epigenetic loop also accounted for inflammation-driven PanIN-to-PDAC progression in vivo and was upregulated in established human PDAC, whose malignancy was maintained by NRF2 binding to the EZH2 promoter.

DOI: https://doi.org/10.1038/s43018-025-01003-3
Nucleic Acids Res. 2025 Jun 20. pii: gkaf572. [Epub ahead of print]53(12):

Differential oligomerization regulates PHF13 chromatin affinity and function.

Francesca Rossi, Alexandre P Magalhaes, Rene Buschow, Tobias Schubert, Laura Glaser, Andrea Fontana, Julia Mai, Hannah Staege, Astrid Grimme, Hans Will, Sabrina Schriener, Denes Hnisz, Martin Vingron, Andrea M Chiariello, Sarah Kinkley.

PHF13 is a H3K4me3 epigenetic reader that modulates key chromatin processes including transcription, DNA damage response, and chromatin architecture. PHF13 is found aberrantly regulated in different cancers and its misexpression alters the epigenetic landscape of key transcription factors that regulate epithelial-to-mesenchymal transition. In this study, we sought to understand how PHF13's chromatin affinity and diverse chromatin functions are intrinsically regulated. Our results show that PHF13 can oligomerize via conserved ordered regions in its N- and C- terminus increasing its chromatin valence and avidity, promoting polymer-polymer phase separation (PPPS) and chromatin inaccessibility. Impressively, a ∼3- to 5-fold overexpression of PHF13 was sufficient to globally compact chromatin visible by optical microscopy, dependent on its ordered dimerizing regions and oligomerization potential. Unexpectedly, we discovered that PHF13 can self-associate independent of its ordered domains via intrinsically disordered regions, which conversely reduced PHF13's chromatin affinity, formed liquid-liquid phase separated (LLPS) condensates, and differentially impacted gene expression. Our findings support that there is an intrinsic balance between PHF13's ordered and disordered regions and that PHF13 can phase transition between polymer-polymer and liquid-liquid phase separation states to impact chromatin structure and function.

DOI: https://doi.org/10.1093/nar/gkaf572
Science. 2025 Jul 03. 389(6755): eadv2861

Zincore, an atypical coregulator, binds zinc finger transcription factors to control gene expression.

Daniëlle Bianchi, Razvan Borza, Erica De Zan, Guizela Huelsz-Prince, Sebastian Gregoricchio, Marleen Dekker, Alex Fish, Abdelghani Mazouzi, Lona J Kroese, Simon Linder, Miguel Hernandez-Quiles, Michiel Vermeulen, Patrick H N Celie, Paul Krimpenfort, Ji-Ying Song, Wilbert Zwart, Lodewyk Wessels, Sebastian M B Nijman, Anastassis Perrakis, Thijn R Brummelkamp.

Zinc finger proteins (ZNFs) are the largest family of transcription factors, yet how they activate gene expression remains unclear. In this study, we identified Zincore, a protein complex consisting of QRICH1 and SEPHS1, as a ZNF-specific coregulator essential for embryonic development in mice and associated with developmental syndromes in humans. We also identified ZFP91 as a representative Zincore client, binding the conserved promoter motif CTTTAAR. Cryo-electron microscopy of a Zincore-ZFP91-DNA complex revealed a SEPHS1 arginine clamp to recognize the DNA-bound zinc finger domains. This mode of binding explains recognition of different ZNFs and stabilizes ZFP91 onto its cognate DNA motif. Thus, our study identified Zincore as a ZNF-specific coregulator essential for development, involving a distinctive mechanism that locks ZNFs onto DNA and regulates transcription.

DOI: https://doi.org/10.1126/science.adv2861
Bioinformatics. 2025 Jun 24. pii: btaf371. [Epub ahead of print]

Adversarial attack of sequence-free enhancer prediction identifies chromatin architecture.

Jamil Gafur, Olivia W Lang, William K M Lai.

MOTIVATION: The wide range of cellular complexity created by multicellular organisms is due in large part to the intricate and synergistic interplay of regulatory complexes throughout the eukaryotic genome. These regulatory elements 'enhance' specific gene programs and have been shown to operate in diverse networks that are distinct across cell states of the same organism. Attempts to characterize and predict enhancers have typically focused on leveraging information-dense DNA sequence in parallel with epigenomic assays. We examined the viability of enhancer prediction using only a minimal set of epigenomic datasets without direct DNA information.
RESULTS: We demonstrate that chromatin datasets are sufficient to identify enhancers genome-wide with high accuracy. By training networks leveraging data from multiple cell types simultaneously, we generated a cell-type invariant enhancer prediction platform that utilized only the patterns of protein binding for inference. We also showed the utility of swarm-based adversarial attacks (APSO) to deconvolute trained genomic neural networks for the first time. Critically, unlike saliency mapping or other game-theory based approaches, APSO is completely network-architecture independent and can be applied to any prediction engine to derive the features that drive inference.
AVAILABILITY: All software and code for data downloading, processing, enhancer inference, XAI, and complete figure generation are publicly available on GitHub at https://github.com/EpiGenomicsCode/ChromEnhancer and Zenodo at https://doi.org/10.5281/zenodo.15652797.
SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

DOI: https://doi.org/10.1093/bioinformatics/btaf371
Proc Natl Acad Sci U S A. 2025 Jul 08. 122(27): e2425984122

TEAD-targeting small molecules induce a cofactor switch to regulate the Hippo pathway.

Alissa D Guarnaccia, Thijs J Hagenbeek, Wendy Lee, Noelyn Kljavin, Meena Choi, Benjamin Tombling, Carsten Peukert, Gözde Ulas, Vasumathi Kameswaran, Daniel Le, Sayantanee Paul, Samir Vaidya, Jason R Zbieg, James J Crawford, Bence Daniel, Anwesha Dey, Jennie R Lill.

  TEAD proteins are the main transcriptional effectors of the Hippo signaling pathway and a clinical-stage pharmacological target in oncology. Most TEAD-targeting small molecules are designed to disrupt interaction between TEAD and the oncogenic transcriptional activators YAP and TAZ. Here, we uncover an alternative mechanism for a subset of TEAD lipid pocket-binding molecules. We report that select sulfonamide-containing TEAD-targeting compounds enhance the interaction between TEAD and the transcriptional repressor VGLL4. Chemically induced VGLL4-TEAD complexes confer an antiproliferative effect by outcompeting YAP-TEAD complexes at chromatin. This cofactor switch from YAP to VGLL4 impacts transcriptional networks, including influencing the expression of genes involved in cellular proliferation and mechanosignaling. We demonstrate that VGLL4 is required for an antiproliferative response to these sulfonamide-containing compounds by counteracting YAP. We show that VGLL4 overexpression can confer sensitivity to these compounds in Hippo-driven cell lines, and we further show that genetic deletion of VGLL4 oblates cellular responsiveness to these molecules in cells and in vivo. Our data reveal a category of TEAD inhibitors that act as "molecular glues" toward the repressive VGLL4-TEAD interaction. These findings open up understandings for curbing the oncogenic activity of Hippo pathway deregulation in cancer, and identify glue-like molecules that promote transcriptional repression.

Keywords:  Hippo; TEAD; VGLL4; cancer therapy; molecular glue

DOI:  https://doi.org/10.1073/pnas.2425984122
Dev Cell. 2025 Jun 25. pii: S1534-5807(25)00361-2. [Epub ahead of print]

A dual enhancer-attenuator element ensures transient Cdx2 expression during mouse posterior body formation.

Irène Amblard, Damir Baranasic, Sheila Q Xie, Benjamin Moyon, Michelle Percharde, Boris Lenhard, Vicki Metzis.

  During development, cells express precise gene expression programs to assemble the trunk of the body plan. Appropriate control over the duration of the transcription factor Cdx2 is critical to achieve this outcome, yet how cells control the onset, maintenance, or termination of Cdx2 has remained unclear. Here, we delineate the cis-regulatory logic orchestrating dynamic Cdx2 expression in mouse caudal epiblast progenitors and their derivatives. Combining CRISPR-mediated deletion of regulatory elements with in vitro models and in vivo validation, we demonstrate that distinct enhancers and a repressive element embedded at the Cdx2 locus act sequentially to drive transient Cdx2 expression. We pinpoint an "attenuator": a minimal region relying on a nuclear receptor to extinguish Cdx2. Changing this single motif converts the attenuator to an enhancer with the opposite regulatory behavior. Our findings establish a dual cis-regulatory logic ensuring precise spatiotemporal control over gene expression for vertebrate body patterning.

Keywords:  CDX; WNT signaling; attenuator; directed differentiation of mouse embryonic stem cells; enhancer; motif composition; retinoic acid nuclear receptor; silencer; spinal cord development

DOI:  https://doi.org/10.1016/j.devcel.2025.06.006
Cell Rep Med. 2025 Jun 27. pii: S2666-3791(25)00288-5. [Epub ahead of print] 102215

Epigenetic profiling identifies markers of endocrine resistance and therapeutic options for metastatic castration-resistant prostate cancer.

Tesa M Severson, Emma Minnee, Yanyun Zhu, Karianne Schuurman, Holly M Nguyen, Lisha G Brown, Sini Hakkola, Renee Menezes, Sebastian Gregoricchio, Yongsoo Kim, Jeroen Kneppers, Simon Linder, Suzan Stelloo, Cor Lieftink, Michiel S van der Heijden, Matti Nykter, Vincent van der Noort, Joyce Sanders, Ben Morris, Guido Jenster, Geert Jlh van Leenders, Mark Pomerantz, Matthew L Freedman, Roderick L Beijersbergen, Alfonso Urbanucci, Lodewyk Wessels, Peter S Nelson, Eva Corey, Stefan Prekovic, Wilbert Zwart, Andries M Bergman.

  Androgen receptor (AR) signaling inhibitors, including enzalutamide, are treatment options for patients with metastatic castration-resistant prostate cancer (mCRPC), but resistance inevitably develops. Using metastatic samples from a prospective phase 2 clinical trial, we epigenetically profile enhancer/promoter activities with acetylation of lysine residue 27 on histone 3 (H3K27ac) chromatin immunoprecipitation followed by sequencing, before and after AR-targeted therapy. We identify a distinct subset of H3K27ac-differentially marked regions that are associated with treatment responsiveness, which we successfully validate in mCRPC patient-derived xenograft (PDX) models. In silico analyses reveal histone deacetylase (HDAC)3 to critically drive resistance to hormonal interventions, which we validate in vitro. Critically, we identify the pan-HDAC inhibitor vorinostat to be effective in decreasing tumor cell proliferation, both in vitro and in vivo. Moreover, we uncover evidence for HDAC3 working together with glucocorticoid receptor (GR) as a potential mechanism for this therapeutic effect. These findings demonstrate the rationale for therapeutic strategies including HDAC inhibitors to improve patient outcome in advanced stages of mCRPC.

Keywords:  H3K27ac; HDAC inhibitors; androgen receptor; biomarkers; drug resistance; enzalutamide; epigenetics; hormone intervention; mCRPC; prostate cancer

DOI:  https://doi.org/10.1016/j.xcrm.2025.102215
Genome Biol. 2025 Jul 03. 26(1): 192

A new IDH-independent hypermethylation phenotype is associated with astrocyte-like cell state in glioblastoma.

Ana Luisa Costa, Daria Doncevic, Yonghe Wu, Lin Yang, Ka Hou Man, Anna-Sophie Spreng, Hannah Winter, Michael Nai Chung Fletcher, Bernhard Radlwimmer, Carl Herrmann.

BACKGROUND: DNA methylation plays a crucial role in cancer development and progression and has been linked to genetically and clinically distinct tumor classes, including IDH-mutated and IDH-wildtype adult-type diffuse gliomas. Here, we identify a CpG-island methylator phenotype (CIMP) that characterizes the receptor tyrosine kinase 2 (RTK2) subtype of IDH-wildtype glioblastoma.
RESULTS: This RTK2-CIMP affects genomic locations and cell functions distinct from those of IDH mutation-associated IDH-CIMP and suppresses the expression of its target genes. The RTK2-CIMP-region chromatin is characterized by a combination of repressive and activating marks, including polycomb-associated H3K27me3 and enhancer-associated H3K4me1, consistent with DNA methylation-mediated silencing of genes with bivalent-state promoters in neural progenitor cells. Functionally, RTK2-CIMP affects neuronal lineage genes and is significantly associated with astrocyte-like glioblastoma, suggesting that RTK2-CIMP is an epigenetic signature of the astrocyte-like cell state. Furthermore, we demonstrate that RTK2-CIMP can be induced by genetic manipulation in glioblastoma cells.
CONCLUSIONS: Our results suggest that RTK2-CIMP is a key contributor to cell-state plasticity in glioblastoma.

DOI: https://doi.org/10.1186/s13059-025-03670-y
PLoS Genet. 2025 Jul 01. 21(7): e1011724

The length and strength of compartmental interactions are modulated by condensin II activity.

Randi Isenhart, Son C Nguyen, Leah Rosin, Weihuan Cao, Patrick Walsh, Haris Muzaffar, Christopher E Ellison, Eric F Joyce.

The spatial organization of the genome is crucial for its function and integrity. Although the ring-like SMC complex condensin II has a well-documented role in organizing mitotic chromosomes, its function in interphase chromatin structure has remained more enigmatic. Using a combination of Oligopaint fluorescence in situ hybridization (FISH) and Hi-C, we show that altering condensin II levels in diploid Drosophila cells significantly changes chromosome architecture at large length scales between chromatin compartments. Notably, condensin II overexpression disrupts the robust boundary between heterochromatin and euchromatin, leading to interactions that span entire chromosomes. These interactions occur independent from transcriptional changes, suggesting that the mechanisms driving compartment formation and their interactions might be distinct aspects of genome organization. Our results provide new insights into the dynamic nature of chromosome organization and underscore the importance of condensin II in maintaining genomic stability.

DOI: https://doi.org/10.1371/journal.pgen.1011724
Mol Cell. 2025 Jul 01. pii: S1097-2765(25)00542-8. [Epub ahead of print]

Chromosomal tethering and mitotic transcription promote ecDNA nuclear inheritance.

Ashley Nichols, Yujin Choi, Roshan Xavier Norman, Yanyang Chen, Josefine Striepen, Eralda Salataj, Eléonore Toufektchan, Richard Koche, John Maciejowski.

  Extrachromosomal DNAs (ecDNAs) are circular DNAs that function in tumor progression and treatment resistance by amplifying oncogenes. ecDNAs lack centromeres and thus are not constrained to Mendelian segregation, enabling unequal and rapid accumulation within daughter cells. Despite intrinsic links to their oncogenic potential, the fidelity and mechanisms of ecDNA inheritance are poorly understood. Using human cancer cell lines, we show that ecDNAs are protected against cytosolic mis-segregation through mitotic clustering and tethering to mitotic chromosome ends. Accurate nuclear segregation of MYC-amplifying ecDNA depends on BRD4 transcriptional co-activation and mitotic transcription of the long non-coding RNA PVT1, which is frequently co-amplified with MYC. Disruption of ecDNA mitotic clustering through BRD4 inhibition, PVT1 depletion, or transcription inhibition causes ecDNA micronucleation and formation of homogeneously staining regions. We propose that nuclear inheritance of ecDNA is facilitated by an RNA-based mechanism that clusters ecDNA during mitosis and protects against cytosolic mis-segregation and chromosomal reintegration.

Keywords:  MYC; PVT1; ecDNA; genome amplification; homogeneously staining region; micronuclei

DOI:  https://doi.org/10.1016/j.molcel.2025.06.013
Genome Biol. 2025 Jul 01. 26(1): 187

Deciphering gene regulatory programs in mouse embryonic skin through single-cell multiomics analysis.

Qiuting Deng, Pengfei Cai, Yingjie Luo, Zhongjin Zhang, Wen Ma, Zijie Huang, Xiaoya Chen, Shijie Hao, Weiguang Ma, Jiangshan Xu, Chunqing Wang, Mengnan Cheng, Xiumei Lin, Ru Zhou, Shanshan Duan, Junjie Chen, Ronghai Li, Xuyang Shi, Chang Liu, Liang Wu, Peng Gao, Jianting Li, Xiao Yang, Xiangdong Wang, Jun Xie, Longqi Liu, Yue Yuan, Chuanyu Liu.

   BACKGROUND: Cell type-specific transcriptional heterogeneity in embryonic mouse skin is well-documented, but few studies have investigated the regulatory mechanisms. Here, we present high-throughput single-cell chromatin accessibility and transcriptome sequencing (HT-scCAT-seq), a method that simultaneously profiles transcriptome and chromatin accessibility. We utilized HT-scCAT-seq to dissect the gene regulatory mechanism governing epidermal stratification, periderm terminal differentiation, and fibroblast specification.
RESULTS: By linking chromatin accessibility to gene expression, we identify candidate cis-regulatory elements (cCREs) and their target genes which are crucial for dermal and epidermal development. We describe cells with similar gene expression profiles that exhibit distinct chromatin accessibility statuses during periderm terminal differentiation. Finally, we characterize the underlying lineage-determining transcription factors and demonstrate that ALX4 and RUNX2 are candidate transcription factors regulators of the dermal papilla lineage development through in silico perturbation analysis and CUT&Tag experiment.
CONCLUSIONS: Overall, HT-scCAT-seq represents a powerful tool for unraveling the spatiotemporal dynamics of gene regulation in single cells. Our results advance the understanding of embryonic skin development while providing a scalable framework for investigating regulatory mechanisms across diverse biological systems and disease contexts.

Keywords:   Cis-regulatory elements; Embryonic skin development; HT-scCAT-seq

DOI:  https://doi.org/10.1186/s13059-025-03652-0
Mol Cell. 2025 Jun 27. pii: S1097-2765(25)00508-8. [Epub ahead of print]

Reconciling competing models on the roles of condensates and soluble complexes in transcription factor function.

Anne Bremer, Walter H Lang, Ryan P Kempen, Kumari Sweta, Aaron B Taylor, Madeleine B Borgia, Aseem Z Ansari, Tanja Mittag.

  Phase separation regulates many biological processes, but the role of transcription factor (TF)-mediated condensates in gene regulation is contentious. We used Gcn4, a prototypical budding yeast TF, to assess two competing models for transcription activation, i.e., mediated via soluble complexes or through transcriptional condensates. We find that the ability of Gcn4 to form soluble complexes with coactivator subunit Med15 closely mirrors its propensity to recruit Med15 into condensates. Both properties are predictive of in vivo activity, cautioning against interpretation of mutational data without direct comparisons. Unexpectedly, Gcn4 variants with the highest affinities for Med15 do not function as per expectation. Instead, their lower activities reflect their ability to phase separate with Med15, suggesting that condensate formation tempers their activity. Our results show that TFs can function as soluble complexes as well as condensates, reconciling two seemingly opposing models, with implications for other phase-separating systems.

Keywords:  GCN4; MED15; Mediator; biomolecular condensate; fuzzy complex; gene regulation; intrinsically disordered protein region; multivalent interaction; phase separation; transactivation domain

DOI:  https://doi.org/10.1016/j.molcel.2025.06.008
Nat Commun. 2025 Jul 01. 16(1): 6042

Mapping the nuclear landscape with multiplexed super-resolution fluorescence microscopy.

Fariha Rahman, Victoria Augoustides, Emma Tyler, Timothy A Daugird, Christian Arthur, Wesley R Legant.

The nucleus coordinates many different processes. Visualizing how these are spatially organized requires imaging protein complexes, epigenetic marks, and DNA across scales from single molecules to the whole nucleus. To accomplish this, we develop a multiplexed imaging protocol to localize 13 different nuclear targets with nanometer precision. Within single cells, we show that nuclear specification into active and repressive states exists along a spectrum of length scales, emerging below one micron and becoming strengthened at the nanoscale with unique organizational principles in both heterochromatin and euchromatin. HP1α was positively correlated with DNA at the microscale but uncorrelated at the nanoscale. RNA Polymerase II, p300, and CDK9 were positively correlated at the microscale but became partitioned below 300 nm. Perturbing histone acetylation or transcription disrupted nanoscale organization but had less effect at the microscale. We envision that our imaging and analysis pipeline will be useful to reveal the organizational principles not only of the cell nucleus but also other cellular compartments.

DOI: https://doi.org/10.1038/s41467-025-61358-0
Nat Commun. 2025 Jul 01. 16(1): 5945

Spatial profiling of chromatin accessibility in formalin-fixed paraffin-embedded tissues.

Pengfei Guo, Yufan Chen, Liran Mao, Angelysia Cardilla, Chin Nien Lee, Yan Cui, Dengge Jin, Yucong Hua, Xiaowei Xu, Yanxiang Deng.

Formalin-fixed paraffin-embedded (FFPE) samples represent a vast, untapped resource for epigenomic research, yet molecular tools for deep analysis of these specimens remain limited. We introduce spatial FFPE-ATAC-seq, an approach for in situ profiling chromatin accessibility within archived tissues. This approach overcomes formalin-induced crosslinking challenges, allowing high-resolution mapping of chromatin landscapes while preserving tissue architecture. Applying spatial FFPE-ATAC-seq to mouse and human tissues, including brain and thymus, reveals intricate spatial organization and distinct cell types in alignment with tissue morphology. Integration with single-cell RNA sequencing validates the precision of our chromatin profiles in identifying key cell types and regulatory elements. We further apply this method to human melanoma, comprehensively characterizing chromatin accessibility across both tumor and non-tumor regions. This method significantly expands the toolkit for epigenomic research, unlocking the potential of an extensive collection of archived FFPE samples for studying gene regulation and disease mechanisms with spatial context.

DOI: https://doi.org/10.1038/s41467-025-60882-3
Nat Commun. 2025 Jul 01. 16(1): 5959

O-GlcNAcylation of FOXK1 co-opts BAP1 to orchestrate the E2F pathway and promotes oncogenesis.

Oumaima Ahmed, Louis Masclef, Nicholas Iannantuono, Jessica Gagnon, Mila Gushul-Leclaire, Karine Boulay, Benjamin Estavoyer, Mohamed Echbicheb, Kalidou Ali Boubacar, Marty Poy, Amina Boubekeur, Saad Menggad, Alejandro Schcolnik-Cabrera, Aurelio Balsalobre, Eric Bonneil, Pierre Thibault, Laura Hulea, Frédérick A Mallette, Jacques Drouin, Yoshiaki Tanaka, El Bachir Affar.

The E2F transcription factors constitute a core transcriptional network that governs cell division and oncogenesis in multi-cellular organisms, although their molecular mechanisms remain incompletely understood. Here, we show that elevated expression of the transcription factor FOXK1 promotes transcription of E2F target genes and cellular transformation. High expression of FOXK1 in patient tumors is also strongly correlated with E2F gene expression. Mechanistically, we demonstrate that FOXK1 is O-GlcNAcylated, and loss of this modification impairs FOXK1 ability to promote cell proliferation and tumor growth. We also show that expression of FOXK1 O-GlcNAcylation-defective mutants results in reduced recruitment of the H2AK119 deubiquitinase and tumor suppressor BAP1 to E2F target genes. This event is associated with a transcriptional repressive chromatin environment and reduced cell proliferation. Our results define an essential role of FOXK1 O-GlcNAcylation in co-opting the tumor suppressor BAP1 to promote cancer cell progression through orchestration of the E2F pathway.

DOI: https://doi.org/10.1038/s41467-025-61022-7
Nat Commun. 2025 Jul 01. 16(1): 5737

Histone lysine methyltransferases MLL3 and MLL4 direct gene expression to produce platelets efficiently.

Guozhen Gao, Josimar Dornelas Moreira, Prosun Das, Kevin Lin, Kai Ge, Taiping Chen, Yue Lu, Margarida A Santos.

Circulating blood platelets are responsible for maintaining hemostasis. They are released into blood vessels from mature megakaryocytes. Although several transcription factors have been reported to orchestrate the transcriptional programs required for platelet production, how chromatin regulators control these processes is still poorly understood. MLL3 and MLL4 are the main lysine methyltransferases responsible for the deposition of H3K4me1 histone marks at enhancers. MLL3 and MLL4 typically form complexes with other co-factors, such as PTIP. Recently, we showed that loss of PTIP leads to decreased platelet numbers in mice. Here, we find that, although MLL3/4 double deficiency does not alter megakaryopoiesis and endomitosis, the final step of megakaryocyte maturation is affected due to an abnormal cytoskeleton and demarcation membrane system. MLL3/4 double-deficient mice develop macrothrombocytopenia; platelets are preactive and pro-apoptotic, leading to their rapid clearance from the circulation. Increased megakaryopoeisis in the bone marrow and spleen cannot compensate for these abnormalities. Mechanistically, the expression of genes responsible for normal megakaryocyte function and platelet production is altered in MLL3/4-deficient megakaryocytes, partly due to impaired enhancer functions associated with these genes. Our findings provide insights into the epigenetic programs that are important for platelet biogenesis.

DOI: https://doi.org/10.1038/s41467-025-61247-6
Sci Rep. 2025 Jul 02. 15(1): 23119

Deep learning deciphers the related role of master regulators and G-quadruplexes in tissue specification.

Artem Bashkatov, Andrey Andreasyan, Dmitry Konovalov, Alan Herbert, Maria Poptsova.

  G-quadruplexes (GQs) are non-canonical DNA structures encoded by G-flipons with potential roles in gene regulation and chromatin structure. Here, we explore the role of G-flipons in tissue specification. We present a deep learning-based framework for the genome-wide G-flipon predictions across 14 human tissue types. The model was trained using high-confidence experimental maps of GQ-forming sequences and ATAC-seq peaks, conjoined with the location of RNA polymerase, histone marks, and transcription factor binding sites. The training dataset for the DeepGQ model was derived from EndoQuad level 4-6 GQs. Model predictions were subsequently validated against the comprehensive EndoQuad dataset (levels 1-6) to optimize the whole-genome prediction threshold. To identify tissue-specific regulatory patterns, we classified GQ promoter predictions as either 'core' or 'tissue-specific'. We identified a notable overlap between predicted unique tissue-specific GQ sites and master regulatory genes (MRGs), tissue-specific DNase-hypersensitivity sites, and proteins that modulate R-loop formation. Collectively, the findings highlight the transactions between MRG and G-flipons intermediated by RNA: DNA hybrids associated with tissue specification.

Keywords:  Chromatin; Deep learning; Flipons; G-quadruplex; R-loops; Tissue differentiation

DOI:  https://doi.org/10.1038/s41598-025-07579-1
Nat Commun. 2025 Jul 01. 16(1): 5834

BMAL1 and ARNT enable circadian HIF2α responses in clear cell renal cell carcinoma.

Rebecca M Mello, Diego Gomez Ceballos, Colby R Sandate, Sicong Wang, Celine Jouffe, Daniel Agudelo, Nina Henriette Uhlenhaut, Nicolas H Thomä, M Celeste Simon, Katja A Lamia.

Circadian disruption enhances cancer risk, and many tumors exhibit disordered circadian gene expression. We show rhythmic gene expression is unexpectedly robust in clear cell renal cell carcinoma (ccRCC). The core circadian transcription factor BMAL1 is closely related to ARNT, and we show that BMAL1-HIF2α regulates a subset of HIF2α target genes in ccRCC cells. Depletion of BMAL1 selectively reduces HIF2α chromatin association and target gene expression and reduces ccRCC growth in culture and in xenografts. Analysis of pre-existing data reveals higher BMAL1 in patient-derived xenografts that are sensitive to growth suppression by a HIF2α antagonist (PT2399). BMAL1-HIF2α is more sensitive than ARNT-HIF2α is to suppression by PT2399, and the effectiveness of PT2399 for suppressing xenograft tumor growth in vivo depends on the time of day at which it is delivered. Together, these findings indicate that an alternate HIF2α heterodimer containing the circadian partner BMAL1 influences HIF2α activity, growth, and sensitivity to HIF2α antagonist drugs in ccRCC cells.

DOI: https://doi.org/10.1038/s41467-025-60904-0
Nat Genet. 2025 Jun 30.

Disruption of TAD hierarchy promotes LTR co-option in cancer.

Elissa W P Wong, Merve Sahin, Rui Yang, UkJin Lee, Dan Li, Yingqian A Zhan, Rohan Misra, Fanny Tomas, Nawaf Alomran, Alexander Polyzos, Cindy J Lee, Tuan Trieu, Alexander Martinez-Fundichely, Thomas Wiesner, Andrew Rosowicz, Shuyuan Cheng, Christina Liu, Morgan Lallo, Alexander N Shoushtari, Taha Merghoub, Pierre-Jacques Hamard, Richard Koche, Ekta Khurana, Effie Apostolou, Deyou Zheng, Yu Chen, Christina S Leslie, Ping Chi.

Transposable elements (TEs) are abundant in the human genome, and they provide the source for genetic and functional diversity. Previous studies have suggested that TEs are repressed by DNA methylation and chromatin modifications. Here through integrating transcriptome and 3D genome architecture studies, we showed that haploinsufficient loss of NIPBL selectively activates alternative promoters (altPs) at the long terminal repeats (LTRs) of the TE subclasses. This activation occurs through the reorganization of topologically associating domain (TAD) hierarchical structures and the recruitment of proximal enhancers. These observations indicate that TAD hierarchy restricts transcriptional activation of LTRs that already possess open chromatin features. Perturbation of hierarchical chromatin topology can lead to co-option of LTRs as functional altPs, driving aberrant transcriptional activation of oncogenes. These data uncovered a new layer of regulatory mechanisms of TE expression and posit TAD hierarchy dysregulation as a new mechanism for altP-mediated oncogene activation and transcriptional diversity in cancer.

DOI: https://doi.org/10.1038/s41588-025-02239-6
Nat Commun. 2025 Jul 04. 16(1): 6194

Precision-edited histone tails disrupt polycistronic gene expression controls in trypanosomes.

Markéta Novotná, Michele Tinti, Joana R C Faria, David Horn.

Transcription of protein coding genes in trypanosomatids is atypical and almost exclusively polycistronic. In Trypanosoma brucei, for example, approximately 150 polycistrons, and 8000 genes, are constitutively transcribed by RNA polymerase II. The RNA pol-II promoters are also unconventional and characterised by regions of chromatin enriched for histones with specific patterns of post-translational modification on their divergent N-terminal tails. To investigate the roles of histone tail-residues in gene expression control in T. brucei, we engineered strains exclusively expressing mutant histones. We used an inducible CRISPR-Cas9 system to delete >40 histone H4 genes, complementing the defect with a single ectopic H4 gene. The resulting "histoneH4" strains were validated using whole-genome sequencing and transcriptome analysis. We then performed saturation mutagenesis of six histone H4 N-terminal tail lysine residues, that are either acetylated or methylated, and profiled relative fitness of 384 distinct precision-edited mutants. H4lys10 mutations were not tolerated, but we derived nineteen strains exclusively expressing distinct H4lys4 or H4lys14 mutants. Proteomic and transcriptomic analysis of H4lys4 glutamine mutants revealed significantly reduced expression of genes adjacent to RNA pol-II promoters, where glutamine mimics abnormally elevated acetylation. Thus, we present direct evidence for polycistronic expression control by modified histone H4 N-terminal tail residues in trypanosomes.

DOI: https://doi.org/10.1038/s41467-025-61480-z
Nucleic Acids Res. 2025 Jun 20. pii: gkaf611. [Epub ahead of print]53(12):

De novo design of insulated cis-regulatory elements based on deep learning-predicted fitness landscape.

Haochen Wang, Yanhui Xiang, Ziming Liu, Wen Yin, Boyan Li, Long Qian, Xiaowo Wang, Chunbo Lou.

Precise control of gene activity within a host cell is crucial in bioengineering applications. Despite significant advancements in cis-regulatory sequence activity prediction and reverse engineering, the context-dependent effects of host cellular environment have long been neglected, leading to ongoing challenges in accurately modeling regulatory processes. Here, we introduce an insulated design strategy to purify and model host-independent transcriptional activity. By integrating heterologous paired cis- and trans-regulatory modules into an orthogonal host cell, we established a controllable transcriptional regulatory system. Using a deep learning-based algorithm combined with an experimental data purification process, we achieved the de novo design full-length transcriptional promoter sequences driven by a host-independent activity landscape. Notably, this landscape accurately captured the transcriptional activity of the insulated system, enabling the generation of cis-regulatory sequences with desirable sequence and functional diversity for two distinct trans-RNA polymerases. Importantly, their activities are precisely predictable in both bacterial (Escherichia coli) and mammalian (Chinese hamster ovary) cell lines. We anticipated that de novo design strategy can be expanded to other complex cis-regulatory elements by integrating the deep learning-based algorithm with the construction of paired cis- and trans-regulatory modules in orthogonal host systems.

DOI: https://doi.org/10.1093/nar/gkaf611
Nat Commun. 2025 Jul 01. 16(1): 5606

MORC2 is a phosphorylation-dependent DNA compaction machine.

Winnie Tan, Jeongveen Park, Hariprasad Venugopal, Jieqiong Lou, Prabavi Shayana Dias, Pedro L Baldoni, Kyoung-Wook Moon, Toby A Dite, Christine R Keenan, Alexandra D Gurzau, Joonyoung Lee, Timothy M Johanson, Andrew Leis, Jumana Yousef, Vineet Vaibhav, Laura F Dagley, Ching-Seng Ang, Laura D Corso, Chen Davidovich, Stephin J Vervoort, Gordon K Smyth, Marnie E Blewitt, Rhys S Allan, Elizabeth Hinde, Sheena D'Arcy, Je-Kyung Ryu, Shabih Shakeel.

The Microrchidia (MORC) family of chromatin-remodelling ATPases is pivotal in forming higher-order chromatin structures that suppress transcription. The exact mechanisms of MORC-induced chromatin remodelling have been elusive. Here, we report an in vitro reconstitution of full-length MORC2, the most commonly mutated MORC member, linked to various cancers and neurological disorders. MORC2 possesses multiple DNA-binding sites that undergo structural rearrangement upon DNA binding. MORC2 locks onto the DNA using its C-terminal domain (CTD) and acts as a clamp. A conserved phosphate-interacting motif within the CTD was found to regulate ATP hydrolysis and cooperative DNA binding. Importantly, MORC2 mediates chromatin remodelling via ATP hydrolysis-dependent DNA compaction in vitro, regulated by the phosphorylation state of its CTD. These findings position MORC2 CTD phosphorylation as a critical regulator of chromatin remodelling and a promising therapeutic target.

DOI: https://doi.org/10.1038/s41467-025-60751-z