bims-crepig 2025-02-02 papers

bims-crepig

Biomed News

on Chromatin regulation and epigenetics in cell fate and cancer

Issue of 2025–02–02
twenty-six papers selected by
Connor Rogerson, University of Cambridge

Genomic clustering tendency of transcription factors reflects phase-separated transcriptional condensates at super-enhancers.
Glucocorticoid receptor suppresses GATA6-mediated RNA polymerase II pause release to modulate classical subtype identity in pancreatic cancer.
Two unrelated distal genes activated by a shared enhancer benefit from localizing inside the same small topological domain.
Genetic coupling of enhancer activity and connectivity in gene expression control.
SPT5 regulates RNA polymerase II stability via Cullin 3-ARMC5 recognition.
The α-globin super-enhancer acts in an orientation-dependent manner.
Chemical catalyst manipulating cancer epigenome and transcription.
Synthetic transcription factors establish the function of nine amino acid transactivation domains of Komagataella phaffii Mxr1.
HoxBlinc lncRNA reprograms CTCF-independent TADs to drive leukemic transcription and HSC dysregulation in NUP98-rearranged leukemia.
Telomemore enables single-cell analysis of cell cycle and chromatin condensation.
A multi-modal transformer for cell type-agnostic regulatory predictions.
The common murine retroviral integration site activating Hhex marks a distal regulatory enhancer co-opted in human Early T-cell precursor leukemia.
Pervasive RNA-binding protein enrichment on TAD boundaries regulates TAD organization.
Abundant repressor binding sites in human enhancers are associated with the fine-tuning of gene regulation.
PRC2 promotes canalisation during endodermal differentiation.
scATAC-seq generates more accurate and complete regulatory maps than bulk ATAC-seq.
c-JUN: a chromatin repressor that limits mesoderm differentiation in human pluripotent stem cells.
EZH2 serves as a viable therapeutic target for myeloma-induced osteolytic bone destruction.
Centromeric chromatin clearings demarcate the site of kinetochore formation.
ChromoGen: Diffusion model predicts single-cell chromatin conformations.
Multiplex generation and single-cell analysis of structural variants in mammalian genomes.
Nuclear blebs are associated with destabilized chromatin packing domains.
TRIM28-dependent developmental heterogeneity determines cancer susceptibility through distinct epigenetic states.
The ortholog of human DNAJC9 promotes histone H3-H4 degradation and is counteracted by Asf1 in fission yeast.
SNP rs9364554 Modulates Androgen Receptor Binding and Drug Response in Prostate Cancer.
Trithorax regulates long-term memory in Drosophila through epigenetic maintenance of mushroom body metabolic state and translation capacity.

Nucleic Acids Res. 2025 Jan 24. pii: gkaf015. [Epub ahead of print]53(3):

Genomic clustering tendency of transcription factors reflects phase-separated transcriptional condensates at super-enhancers.

Shengyuan Wang, Zhenjia Wang, Chongzhi Zang.

Many transcription factors (TFs) have been shown to bind to super-enhancers, forming transcriptional condensates to activate transcription in various cellular systems. However, the genomic and epigenomic determinants of phase-separated transcriptional condensate formation remain poorly understood. Questions regarding which TFs tend to associate with transcriptional condensates and what factors influence their association are largely unanswered. Here we systematically analyzed 571 DNA sequence motifs across the human genome and 6650 TF binding profiles across different cell types to identify the molecular features contributing to the formation of transcriptional condensates. We found that the genomic distributions of sequence motifs for different TFs exhibit distinct clustering tendencies. Notably, TF motifs with a high genomic clustering tendency are significantly associated with super-enhancers. TF binding profiles showing a high genomic clustering tendency are further enriched at cell-type-specific super-enhancers. TFs with a high binding clustering tendency also possess high liquid-liquid phase separation abilities. Compared to nonclustered TF binding, densely clustered TF binding sites are more enriched at cell-type-specific super-enhancers with higher chromatin accessibility, elevated chromatin interaction and stronger association with cancer outcomes. Our results indicate that the clustered genomic binding patterns and the phase separation properties of TFs collectively contribute to the formation of transcriptional condensates.

DOI: https://doi.org/10.1093/nar/gkaf015
Gut. 2025 Jan 30. pii: gutjnl-2024-334374. [Epub ahead of print]

Glucocorticoid receptor suppresses GATA6-mediated RNA polymerase II pause release to modulate classical subtype identity in pancreatic cancer.

Thomas L Ekstrom, Raya M Rosok, Amro M Abdelrahman, Christina Parassiadis, Meghana Manjunath, Marianna Y Dittrich, Xin Wang, Ana P Kutschat, Akshay Kanakan, Ashish Rajput, Nadine Schacherer, Teodora Lukic, Danielle M Carlson, Julia Thiel, Waltraut Kopp, Philipp Stroebel, Volker Ellenrieder, Jochen Gaedcke, Meng Dong, Zeynab Najafova, Mark J Truty, Elisabeth Hessmann, Steven A Johnsen.

   BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal cancer with a 5-year survival rate of 12%. It has two major molecular subtypes: classical and basal, regulated by the master transcription factors (MTFs) GATA6 and ΔNp63, respectively.
OBJECTIVE: This study sought to uncover the transcriptional regulatory mechanisms controlling PDAC subtype identity.
DESIGN: We integrated primary tumour single-cell RNA-seq, patient-derived xenograft RNA-seq and multispectral imaging to identify MTF-dependent, subtype-specific markers. We created subtype-specific fluorescent reporter systems and conducted drug screenings to find actionable targets. We analysed chromatin accessibility (ATAC-seq), genome-wide occupancy (ChIP-seq) for epigenetic status (H3K27ac), MTFs (GATA6, ΔNp63), RNA polymerase II (Pol II), H3K4me3-anchored chromatin topology (HiChIP) and nascent RNA capture sequencing (PRO-seq). Additionally, we used nuclease-dead Cas9 (dCas9) to manipulate transcriptional regulatory mechanisms.
RESULTS: Our approach identified glucocorticoid receptor (GR) agonists as agents that suppress the classical transcriptional programme by interacting with GATA6. GATA6 regulates classical-specific transcription through promoter-proximal pause release. Depletion of GATA6 increased Pol II occupancy at GATA6-bound enhancers and transcriptional start sites, stabilising enhancer-promoter interactions. Artificially inducing pausing at GATA6-bound enhancers with dCas9 abrogated target gene expression and induced pausing at both the enhancer and target gene promoter. Conversely, in basal PDAC ΔNp63 promotes Pol II recruitment and stabilises enhancer-promoter interactions.
CONCLUSION: This study provides new insights into the transcriptional control and role of GR agonists in controlling PDAC molecular subtype identity.

Keywords:  GENE REGULATION; MOLECULAR BIOLOGY; PANCREATIC CANCER; RNA EXPRESSION

DOI:  https://doi.org/10.1136/gutjnl-2024-334374
Genes Dev. 2025 Jan 27.

Two unrelated distal genes activated by a shared enhancer benefit from localizing inside the same small topological domain.

Yike Huang, Marjon J A M Verstegen, Sjoerd J D Tjalsma, Peter H L Krijger, Kavvya Gupta, Minhee Park, Alistair Boettiger, Wouter de Laat.

  Enhancers are tissue-specific regulatory DNA elements that can activate transcription of genes over distance. Their target genes most often are located in the same contact domain-chromosomal entities formed by cohesin DNA loop extrusion and typically flanked by CTCF-bound boundaries. Enhancers shared by multiple unrelated genes are underexplored but may be more common than anticipated. Here, we analyzed the interplay between an enhancer and two distal functionally unrelated genes residing at opposite domain boundaries. The enhancer strongly activated their expression and supported their frequent interactions. Cohesin structured the domain and supported their transcription, but the genes did not rely on each other's transcription or show gene competition. Deleting either domain boundary not only extended the contact domain but led to reduced contacts within the original domain and reduction in the expression of both genes. Conversely, by isolating either gene with the enhancer in shorter domains, through insertion of new CTCF boundaries, intradomain contact frequencies increased, and the gene isolated with the enhancer was upregulated. Collectively, this shows that an enhancer can independently activate unrelated distal genes and that long-range gene regulation benefits from operating in small contact domains.

Keywords:  TAD size; chromatin topology; cohesin-dependent gene regulation; coregulated genes; transcription regulatory network

DOI:  https://doi.org/10.1101/gad.352235.124
Nat Commun. 2025 Jan 27. 16(1): 970

Genetic coupling of enhancer activity and connectivity in gene expression control.

Helen Ray-Jones, Chak Kei Sung, Lai Ting Chan, Alexander Haglund, Pavel Artemov, Monica Della Rosa, Luminita Ruje, Frances Burden, Roman Kreuzhuber, Anna Litovskikh, Eline Weyenbergh, Zoï Brusselaers, Vanessa Xue Hui Tan, Mattia Frontini, Chris Wallace, Valeriya Malysheva, Leonardo Bottolo, Elena Vigorito, Mikhail Spivakov.

Gene enhancers often form long-range contacts with promoters, but it remains unclear if the activity of enhancers and their chromosomal contacts are mediated by the same DNA sequences and recruited factors. Here, we study the effects of expression quantitative trait loci (eQTLs) on enhancer activity and promoter contacts in primary monocytes isolated from 34 male individuals. Using eQTL-Capture Hi-C and a Bayesian approach considering both intra- and inter-individual variation, we initially detect 19 eQTLs associated with enhancer-eGene promoter contacts, most of which also associate with enhancer accessibility and activity. Capitalising on these shared effects, we devise a multi-modality Bayesian strategy, identifying 629 "trimodal QTLs" jointly associated with enhancer accessibility, eGene promoter contact, and gene expression. Causal mediation analysis and CRISPR interference reveal causal relationships between these three modalities. Many detected QTLs overlap disease susceptibility loci and influence the predicted binding of myeloid transcription factors, including SPI1, GABPB and STAT3. Additionally, a variant associated with PCK2 promoter contact directly disrupts a CTCF binding motif and impacts promoter insulation from downstream enhancers. Jointly, our findings suggest an inherent genetic coupling of enhancer activity and connectivity in gene expression control relevant to human disease and highlight the regulatory role of genetically determined chromatin boundaries.

DOI: https://doi.org/10.1038/s41467-025-55900-3
Sci Adv. 2025 Jan 24. 11(4): eadt5885

SPT5 regulates RNA polymerase II stability via Cullin 3-ARMC5 recognition.

Yuki Aoi, Leila Iravani, Isabella C Mroczek, Sarah Gold, Benjamin C Howard, Ali Shilatifard.

The stability of RNA polymerase II (Pol II) is tightly regulated during transcriptional elongation for proper control of gene expression. Our recent studies revealed that promoter-proximal Pol II is destabilized via the ubiquitin E3 ligase cullin 3 (CUL3) upon loss of transcription elongation factor SPT5. Here, we investigate how CUL3 recognizes chromatin-bound Pol II as a substrate. Using an unbiased proteomic screening approach, we identify armadillo repeat-containing 5 (ARMC5) as a CUL3 adaptor required for VCP/p97-dependent degradation of SPT5-depleted, chromatin-bound Pol II. Genome-wide analyses indicate that ARMC5 targets promoter-proximal Pol II in a BTB domain-dependent manner. Further biochemical analysis demonstrates that interaction between ARMC5 and Pol II requires the transcriptional cyclin-dependent kinase 9 (CDK9), supporting a phospho-dependent degradation model. We propose that defective, promoter-proximal Pol II that lacks SPT5 is rapidly eliminated from chromatin in a noncanonical early termination pathway that requires CDK9-dependent interaction with the CUL3-ARMC5 ubiquitin ligase complex.

DOI: https://doi.org/10.1126/sciadv.adt5885
Nat Commun. 2025 Jan 25. 16(1): 1033

The α-globin super-enhancer acts in an orientation-dependent manner.

Mira T Kassouf, Helena S Francis, Matthew Gosden, Maria C Suciu, Damien J Downes, Caroline Harrold, Martin Larke, Marieke Oudelaar, Lucy Cornell, Joseph Blayney, Jelena Telenius, Barbara Xella, Yuki Shen, Nikolaos Sousos, Jacqueline A Sharpe, Jacqueline Sloane-Stanley, Andrew J H Smith, Christian Babbs, Jim R Hughes, Douglas R Higgs.

Individual enhancers are defined as short genomic regulatory elements, bound by transcription factors, and able to activate cell-specific gene expression at a distance, in an orientation-independent manner. Within mammalian genomes, enhancer-like elements may be found individually or within clusters referred to as locus control regions or super-enhancers (SEs). While these behave similarly to individual enhancers with respect to cell specificity, distribution and distance, their orientation-dependence has not been formally tested. Here, using the α-globin locus as a model, we show that while an individual enhancer works in an orientation-independent manner, the direction of activity of a SE changes with its orientation. When the SE is inverted within its normal chromosomal context, expression of its normal targets, the α-globin genes, is severely reduced and the normally silent genes lying upstream of the α-globin locus are upregulated. These findings add to our understanding of enhancer-promoter specificity that precisely activate transcription.

DOI: https://doi.org/10.1038/s41467-025-56380-1
Nat Commun. 2025 Jan 24. 16(1): 887

Chemical catalyst manipulating cancer epigenome and transcription.

Yuki Yamanashi, Shinpei Takamaru, Atsushi Okabe, Satoshi Kaito, Yuto Azumaya, Yugo R Kamimura, Kenzo Yamatsugu, Tomoya Kujirai, Hitoshi Kurumizaka, Atsushi Iwama, Atsushi Kaneda, Shigehiro A Kawashima, Motomu Kanai.

The number and variety of identified histone post-translational modifications (PTMs) are continually increasing. However, the specific consequences of each histone PTM remain largely unclear, primarily due to the lack of methods for selectively and rapidly introducing a desired histone PTM in living cells without genetic engineering. Here, we report the development of a cell-permeable histone acetylation catalyst, BAHA-LANA-PEG-CPP44, which selectively enters leukemia cells, binds to chromatin, and acetylates H2BK120 of endogenous histones in a short reaction time. Time-course analyses of this in-cell catalytic reaction revealed that H2BK120 acetylation attenuates the chromatin binding of negative elongation factor E (NELFE), an onco-transcription factor. This H2BK120 acetylation-mediated removal of NELFE from chromatin reshapes transcription, slows leukemia cell viability, and reduces their tumorigenic potential in mice. Therefore, this histone acetylation catalyst provides a unique tool for elucidating the time-resolved consequences of histone PTMs and may offer a modality for cancer chemotherapy.

DOI: https://doi.org/10.1038/s41467-025-56204-2
J Biol Chem. 2025 Jan 22. pii: S0021-9258(25)00058-4. [Epub ahead of print] 108211

Synthetic transcription factors establish the function of nine amino acid transactivation domains of Komagataella phaffii Mxr1.

Prachi Priya, Vedanth Bellad Shivashankar, Pundi N Rangarajan.

  The zinc finger transcription factor Mxr1 (methanol expression regulator 1) of the methylotrophic yeast Komagataella phaffii (formerly Pichia pastoris) harbors a DNA-binding domain (DBD) consisting of two C2H2 zinc fingers (Mxr1ZF) between amino acids 36-101 and a previously identified nine amino acid transactivation domain (9aaTAD) between residues 365-373 (TAD A, QELESSLNA). Beyond this, 21 putative 9aaTADs (designated TAD B-V) located between amino acids 401-1155 remain to be characterized. Here, we demonstrate that a compact synthetic transcription factor composed of Mxr1ZF and three tandem copies of TAD A can activate the transcription of Mxr1 target genes for ethanol and methanol metabolism with specificity and efficiency comparable to the full-length protein. Expression of individual synthetic transcription factors containing Mxr1ZF and each of the 20 putative 9aaTADs in K. phaffii Δmxr1 strain revealed that 10 of these putative TADs are functional, capable of reversing the growth defect of the mutant and activating transcription of target genes required for ethanol and methanol metabolism. Functional analysis indicates that Mxr1 9aaTADs rely on General Control Non-derepressible 5 (Gcn5), a histone acetyltransferase, for transactivation. These findings suggest that recruitment of Gcn5-mediated histone acetylation at target promoters is a critical step in transcriptional activation by Mxr1 9aaTADs. This study represents the first comprehensive characterization of 9aaTADs in a K. phaffii zinc finger transcription factor, providing insights into their mechanism and potential applications in synthetic biology.

Keywords:  9aaTAD; Komagataella phaffii; Mxr1; Pichia pastoris; Synthetic transcription factor; Transactivation; one‐carbon metabolism; transcription factor; yeast metabolism; zinc finger

DOI:  https://doi.org/10.1016/j.jbc.2025.108211
J Clin Invest. 2025 Jan 30. pii: e184743. [Epub ahead of print]

HoxBlinc lncRNA reprograms CTCF-independent TADs to drive leukemic transcription and HSC dysregulation in NUP98-rearranged leukemia.

Karina Hamamoto, Ganqian Zhu, Qian Lai, Julia Lesperance, Huacheng Luo, Ying Li, Nupur Nigam, Arati Sharma, Feng-Chun Yang, David Claxton, Yi Qiu, Peter D Aplan, Mingjiang Xu, Suming Huang.

  Although nucleoporin 98 (NUP98) fusion oncogenes often drive aggressive pediatric leukemia by altering chromatin structure and expression of HOX genes, underlying mechanisms remain elusive. Here, we report that a Hoxb-associated lncRNA HoxBlinc was aberrantly activated in NUP98-PHF23 fusion-driven leukemias. HoxBlinc chromatin occupancies led to elevated MLL1 recruitment and aberrant homeotic topologically associated domains (TADs) that enhanced chromatin accessibilities and activated homeotic/hematopoietic oncogenes. HoxBlinc-depletion in NUP98 fusion-driven leukemia impaired HoxBlinc binding, TAD integrity, MLL1 recruitment, and MLL1-driven chromatin signature within HoxBlinc-defined TADs in a CTCF-independent manner, leading to inhibited homeotic/leukemic oncogenes that mitigated NUP98 fusion-driven leukemogenesis in xenografted mouse models. Mechanistically, HoxBlinc overexpression in mouse hematopoietic compartment induced leukemias resembling those in NUP98-PHF23 knock-in mice via enhancing HoxBlinc chromatin binding, TAD formation, and Hox gene aberration leading to expansion of hematopoietic stem and progenitor cell (HSPC) and myeloid/lymphoid subpopulations. Thus, our studies reveal a CTCF-independent role of HoxBlinc in leukemic TAD organization and oncogene regulatory networks.

Keywords:  Epigenetics; Genetics; Hematology; Hematopoietic stem cells; Oncogenes

DOI:  https://doi.org/10.1172/JCI184743
Nucleic Acids Res. 2025 Jan 24. pii: gkaf031. [Epub ahead of print]53(3):

Telomemore enables single-cell analysis of cell cycle and chromatin condensation.

Iryna Yakovenko, Ionut Sebastian Mihai, Martin Selinger, William Rosenbaum, Andy Dernstedt, Remigius Gröning, Johan Trygg, Laura Carroll, Mattias Forsell, Johan Henriksson.

Single-cell RNA-seq methods can be used to delineate cell types and states at unprecedented resolution but do little to explain why certain genes are expressed. Single-cell ATAC-seq and multiome (ATAC + RNA) have emerged to give a complementary view of the cell state. It is however unclear what additional information can be extracted from ATAC-seq data besides transcription factor binding sites. Here, we show that ATAC-seq telomere-like reads counter-inituively cannot be used to infer telomere length, as they mostly originate from the subtelomere, but can be used as a biomarker for chromatin condensation. Using long-read sequencing, we further show that modern hyperactive Tn5 does not duplicate 9 bp of its target sequence, contrary to common belief. We provide a new tool, Telomemore, which can quantify nonaligning subtelomeric reads. By analyzing several public datasets and generating new multiome fibroblast and B-cell atlases, we show how this new readout can aid single-cell data interpretation. We show how drivers of condensation processes can be inferred, and how it complements common RNA-seq-based cell cycle inference, which fails for monocytes. Telomemore-based analysis of the condensation state is thus a valuable complement to the single-cell analysis toolbox.

DOI: https://doi.org/10.1093/nar/gkaf031
Cell Genom. 2025 Jan 23. pii: S2666-979X(25)00018-7. [Epub ahead of print] 100762

A multi-modal transformer for cell type-agnostic regulatory predictions.

Nauman Javed, Thomas Weingarten, Arijit Sehanobish, Adam Roberts, Avinava Dubey, Krzysztof Choromanski, Bradley E Bernstein.

  Sequence-based deep learning models have emerged as powerful tools for deciphering the cis-regulatory grammar of the human genome but cannot generalize to unobserved cellular contexts. Here, we present EpiBERT, a multi-modal transformer that learns generalizable representations of genomic sequence and cell type-specific chromatin accessibility through a masked accessibility-based pre-training objective. Following pre-training, EpiBERT can be fine-tuned for gene expression prediction, achieving accuracy comparable to the sequence-only Enformer model, while also being able to generalize to unobserved cell states. The learned representations are interpretable and useful for predicting chromatin accessibility quantitative trait loci (caQTLs), regulatory motifs, and enhancer-gene links. Our work represents a step toward improving the generalization of sequence-based deep neural networks in regulatory genomics.

Keywords:  chromatin accessibility; deep learning; gene regulation; genomics; sequence code; transformer

DOI:  https://doi.org/10.1016/j.xgen.2025.100762
J Biol Chem. 2025 Jan 27. pii: S0021-9258(25)00080-8. [Epub ahead of print] 108233

The common murine retroviral integration site activating Hhex marks a distal regulatory enhancer co-opted in human Early T-cell precursor leukemia.

Joyce Hardwick, Javier Rodriguez-Hernaez, Giovanni Gambi, Bryan J Venters, Yan Guo, Liqi Li, Paul E Love, Neal G Copeland, Nancy A Jenkins, Dimitrios Papaioannou, Iannis Aifantis, Aristotelis Tsirigos, Mircea Ivan, Utpal P Davé.

The Hhex gene encodes a transcription factor that is important for both embryonic and post-natal development, especially of hematopoietic tissues. Hhex is one of the most common sites of retroviral integration in mouse models. We found the most common integrations in AKXD (recombinant inbred strains) T-ALLs occur 57-61kb 3' of Hhex and activate Hhex gene expression. The genomic region of murine leukemia virus (MLV) integrations has features of a developmental stage-specific cis regulatory element (CRE), as evidenced by ATAC-seq in murine progenitor cells and high H3K27 acetylation at the syntenic CRE in human hematopoietic cell lines. With ChIP-exonuclease, we describe occupancy of LIM domain binding protein 1 (LDB1), the constitutive partner of the LIM Only-2 (LMO2), GATA1, and TAL1 transcription factors at GATA sites and a composite GATA-E box within the CRE. With virtual 4C analysis, we observed looping between this +65kb CRE and the proximal intron 1 enhancer of HHEX in primary human ETP-ALLs and in normal progenitor cells. Our results show that retroviral integrations at intergenic sites can mark and take advantage of CREs. Specifically, in the case of HHEX activation, this newly described +65kb CRE is co-opted in the pathogenesis of ETP-ALL by the LMO2/LDB1 complex.

DOI: https://doi.org/10.1016/j.jbc.2025.108233
Nucleic Acids Res. 2025 Jan 07. pii: gkae1271. [Epub ahead of print]53(1):

Pervasive RNA-binding protein enrichment on TAD boundaries regulates TAD organization.

Qiang Sun, Qin Zhou, Yulong Qiao, Xiaona Chen, Hao Sun, Huating Wang.

Mammalian genome is hierarchically organized by CTCF and cohesin through loop extrusion mechanism to facilitate the organization of topologically associating domains (TADs). Mounting evidence suggests additional factors/mechanisms exist to orchestrate TAD formation and maintenance. In this study, we investigate the potential role of RNA-binding proteins (RBPs) in TAD organization. By integrated analyses of global RBP binding and 3D genome mapping profiles from both K562 and HepG2 cells, our study unveils the prevalent enrichment of RBPs on TAD boundaries and define boundary-associated RBPs (baRBPs). We found that baRBP binding is correlated with enhanced TAD insulation strength and in a CTCF-independent manner. Moreover, baRBP binding is associated with nascent promoter transcription. Additional experimental testing was performed using RBFox2 as a paradigm. Knockdown of RBFox2 in K562 cells causes mild TAD reorganization. Moreover, RBFox2 enrichment on TAD boundaries is a conserved phenomenon in C2C12 myoblast (MB) cells. RBFox2 is downregulated and its bound boundaries are remodeled during MB differentiation into myotubes. Finally, transcriptional inhibition indeed decreases RBFox2 binding and disrupts TAD boundary insulation. Altogether, our findings demonstrate that RBPs can play an active role in modulating TAD organization through co-transcriptional association and synergistic actions with nascent promoter transcripts.

DOI: https://doi.org/10.1093/nar/gkae1271
iScience. 2025 Jan 17. 28(1): 111658

Abundant repressor binding sites in human enhancers are associated with the fine-tuning of gene regulation.

Wei Song, Ivan Ovcharenko.

  The regulation of gene expression relies on the coordinated action of transcription factors (TFs) at enhancers, including both activator and repressor TFs. We employed deep learning (DL) to dissect HepG2 enhancers into positive (PAR), negative (NAR), and neutral activity regions. Sharpr-MPRA and STARR-seq highlight the dichotomy impact of NARs and PARs on modulating and catalyzing the activity of enhancers, respectively. Approximately 22% of HepG2 enhancers, termed "repressive impact enhancers" (RIEs), are predominantly populated by NARs and transcriptional repression motifs. Genes flanking RIEs exhibit a stage-specific decline in expression during late development, suggesting RIEs' role in trimming enhancer activities. About 16.7% of human NARs emerge from neutral rhesus macaque DNA. This gain of repressor binding sites in RIEs is associated with a 30% decrease in the average expression of flanking genes in humans compared to rhesus macaque. Our work reveals modulated enhancer activity and adaptable gene regulation through the evolutionary dynamics of TF binding sites.

Keywords:  Biocomputational method; In silico biology; Molecular mechanism of gene regulation

DOI:  https://doi.org/10.1016/j.isci.2024.111658
PLoS Genet. 2025 Jan 30. 21(1): e1011584

PRC2 promotes canalisation during endodermal differentiation.

Jurriaan Jochem Hölzenspies, Dipta Sengupta, Wendy Anne Bickmore, Joshua Mark Brickman, Robert Scott Illingworth.

The genetic circuitry that encodes the developmental programme of mammals is regulated by transcription factors and chromatin modifiers. During early gestation, the three embryonic germ layers are established in a process termed gastrulation. The impact of deleterious mutations in chromatin modifiers such as the polycomb proteins manifests during gastrulation, leading to early developmental failure and lethality in mouse models. Embryonic stem cells have provided key insights into the molecular function of polycomb proteins, but it is impossible to fully appreciate the role of these epigenetic factors in development, or how development is perturbed due to their deficiency, in the steady-state. To address this, we have employed a tractable embryonic stem cell differentiation system to model primitive streak formation and early gastrulation. Using this approach, we find that loss of the repressive polycomb mark H3K27me3 is delayed relative to transcriptional activation, indicating a subordinate rather than instructive role in gene repression. Despite this, chemical inhibition of polycomb enhanced endodermal differentiation efficiency, but did so at the cost of lineage fidelity. These findings highlight the importance of the polycomb system in stabilising the developmental transcriptional response and, in so doing, in shoring up cellular specification.

DOI: https://doi.org/10.1371/journal.pgen.1011584
Sci Rep. 2025 Jan 29. 15(1): 3665

scATAC-seq generates more accurate and complete regulatory maps than bulk ATAC-seq.

E Ravza Gur, Jim R Hughes.

  Bulk ATAC-seq assays have been used to map and profile the chromatin accessibility of regulatory elements such as enhancers, promoters, and insulators. This has provided great insight into the regulation of gene expression in many cell types in a variety of organisms. To date, ATAC-seq has most often been used to provide an average evaluation of chromatin accessibility in populations of cells. The development of a single cell approach (scATAC-seq) assay enables researchers to evaluate chromatin accessibility in individual cells and identify sub-groups in mixed populations of cells. To investigate the full potential of single-cell epigenomic data, we have comprehensively compared the information derived from bulk ATAC-seq and scATAC-seq in populations of cells. We found that the chromatin architecture signal is the same using bulk ATAC-seq and scATAC-seq to analyse aliquots of the same cell population. However, scATAC-seq provides substantially higher data quality compared to bulk ATAC-seq improving the sensitivity to detect relatively weak, but functionally important ATAC-seq signals. Furthermore, we found that scATAC-seq identified differences in what was previously assumed to be a homogenous population of cells. Finally, we determined the number of cells required to generate aggregated open chromatin profiles from single cells and to identify biologically meaningful clusters after pseudo-bulking of data. This study illustrates the added value of using scATAC-seq rather than bulk ATAC-seq in evaluating both homogeneous and heterogeneous populations of cells. This paper provides a comprehensive guide on the benefits of using scATAC-seq data to study gene regulation.

Keywords:  Bulk ATAC-seq; Chromatin accessibility; Clustering; Gene regulation; Pseudo-bulking; Regulatory elements; Single-cell epigenomics; scATAC-seq

DOI:  https://doi.org/10.1038/s41598-025-87351-7
Nucleic Acids Res. 2025 Jan 24. pii: gkaf001. [Epub ahead of print]53(3):

c-JUN: a chromatin repressor that limits mesoderm differentiation in human pluripotent stem cells.

Ran Zhang, Guihuan Li, Qi Zhang, Zhenhua Wang, Dan Xiang, Xiaofei Zhang, Jiekai Chen, Andrew P Hutchins, Dajiang Qin, Huanxing Su, Duanqing Pei, Dongwei Li.

Cell fate determination at the chromatin level is not fully comprehended. Here, we report that c-JUN acts on chromatin loci to limit mesoderm cell fate specification as cells exit pluripotency. Although c-JUN is widely expressed across various cell types in early embryogenesis, it is not essential for maintaining pluripotency. Instead, it functions as a repressor to constrain mesoderm development while having a negligible impact on ectoderm differentiation. c-JUN interacts with MBD3-NuRD complex, which helps maintain chromatin in a low accessibility state at mesoderm-related genes during the differentiation of human pluripotent stem cells into mesoderm. Furthermore, c-JUN specifically inhibits the activation of key mesoderm factors, such as EOMES and GATA4. Knocking out c-JUN or inhibiting it with a JNK inhibitor can alleviate this suppression, promoting mesoderm cell differentiation. Consistently, knockdown of MBD3 enhances mesoderm generation, whereas MBD3 overexpression impedes it. Overexpressing c-JUN redirects differentiation toward a fibroblast-like lineage. Collectively, our findings suggest that c-JUN acts as a chromatin regulator to restrict the mesoderm cell fate.

DOI: https://doi.org/10.1093/nar/gkaf001
Nat Commun. 2025 Jan 31. 16(1): 1206

EZH2 serves as a viable therapeutic target for myeloma-induced osteolytic bone destruction.

Rui Liu, Zongwei Li, Rui Chen, Zhihong Fang, Zhiqiang Liu, Huan Liu.

Myelomatous bone disease is a complication characterized by lytic bone lesions, reduced bone formation, bone pain, and increased fracture risk. Understanding these underlying mechanisms is crucial for developing effective therapeutic approaches. Here we show the role of enhancer of zeste homolog 2 (EZH2) in bone lesions induced by myeloma cells. Our research reveals that cytokines produced by myeloma-associated adipocytes activate the expression of EZH2 in myeloma cells. Furthermore, we find that EZH2 forms a transcriptional repression complex with transcription factor AP2α. This complex promotes trimethylation at lysine 27 of histone H3 (H3K27me3) in the promoter region of the tumor suppressor gene EMP1, resulting in transcriptional silencing. EMP1 silencing leads to increased myeloma cell proliferation and the concomitant secretion of osteolytic cytokines that contribute to bone destruction. Importantly, EZH2 inhibitors effectively treat myeloma-induced osteolytic lesions. Thus, targeting EZH2 represents a potential therapeutic strategy for preventing and managing myeloma bone disease.

DOI: https://doi.org/10.1038/s41467-025-56506-5
Cell. 2025 Jan 23. pii: S0092-8674(24)01467-3. [Epub ahead of print]

Centromeric chromatin clearings demarcate the site of kinetochore formation.

Kathryn Kixmoeller, Ekaterina V Tarasovetc, Elie Mer, Yi-Wei Chang, Ben E Black.

  The centromere is the chromosomal locus that recruits the kinetochore, directing faithful propagation of the genome during cell division. Using cryo-ET on human mitotic chromosomes, we reveal a distinctive architecture at the centromere: clustered 20- to 25-nm nucleosome-associated complexes within chromatin clearings that delineate them from surrounding chromatin. Centromere components CENP-C and CENP-N are each required for the integrity of the complexes, while CENP-C is also required to maintain the chromatin clearing. We find that CENP-C is required in mitosis, not just for kinetochore assembly, likely reflecting its role in organizing the inner kinetochore during chromosome segregation. We further visualize the scaffold of the fibrous corona, a structure amplified at unattached kinetochores, revealing crescent-shaped parallel arrays of fibrils extending >1 μm. Thus, we reveal how the organization of centromeric chromatin creates a clearing at the site of kinetochore formation as well as the nature of kinetochore amplification mediated by corona fibrils.

Keywords:  centromere; chromatin; chromosome; cryo-ET; kinetochore; mitosis; mitotic chromosome; nucleosome

DOI:  https://doi.org/10.1016/j.cell.2024.12.025
Sci Adv. 2025 Jan 31. 11(5): eadr8265

ChromoGen: Diffusion model predicts single-cell chromatin conformations.

Greg Schuette, Zhuohan Lao, Bin Zhang.

Breakthroughs in high-throughput sequencing and microscopic imaging technologies have revealed that chromatin structures vary considerably between cells of the same type. However, a thorough characterization of this heterogeneity remains elusive due to the labor-intensive and time-consuming nature of these experiments. To address these challenges, we introduce ChromoGen, a generative model based on state-of-the-art artificial intelligence techniques that efficiently predicts three-dimensional, single-cell chromatin conformations de novo with both region and cell type specificity. These generated conformations accurately reproduce experimental results at both the single-cell and population levels. Moreover, ChromoGen successfully transfers to cell types excluded from the training data using just DNA sequence and widely available DNase-seq data, thus providing access to chromatin structures in myriad cell types. These achievements come at a remarkably low computational cost. Therefore, ChromoGen enables the systematic investigation of single-cell chromatin organization, its heterogeneity, and its relationship to sequencing data, all while remaining economical.

DOI: https://doi.org/10.1126/sciadv.adr8265
Science. 2025 Jan 31. 387(6733): eado5978

Multiplex generation and single-cell analysis of structural variants in mammalian genomes.

Sudarshan Pinglay, Jean-Benoît Lalanne, Riza M Daza, Sanjay Kottapalli, Faaiz Quaisar, Jonas Koeppel, Riddhiman K Garge, Xiaoyi Li, David S Lee, Jay Shendure.

Studying the functional consequences of structural variants (SVs) in mammalian genomes is challenging because (i) SVs arise much less commonly than single-nucleotide variants or small indels and (ii) methods to generate, map, and characterize SVs in model systems are underdeveloped. To address these challenges, we developed Genome-Shuffle-seq, a method that enables the multiplex generation and mapping of thousands of SVs (deletions, inversions, translocations, and extrachromosomal circles) throughout mammalian genomes. We also demonstrate the co-capture of SV identity with single-cell transcriptomes, facilitating the measurement of SV impact on gene expression. We anticipate that Genome-Shuffle-seq will be broadly useful for the systematic exploration of the functional consequences of SVs on gene expression, the chromatin landscape, and three-dimensional nuclear architecture, while also initiating a path toward a minimal mammalian genome.

DOI: https://doi.org/10.1126/science.ado5978
J Cell Sci. 2025 Jan 29. pii: jcs.262161. [Epub ahead of print]

Nuclear blebs are associated with destabilized chromatin packing domains.

Emily M Pujadas Liwag, Nicolas Acosta, Luay Matthew Almassalha, Yuanzhe Patrick Su, Ruyi Gong, Masato T Kanemaki, Andrew D Stephens, Vadim Backman.

  Disrupted nuclear shape is associated with multiple pathological processes including premature aging disorders, cancer-relevant chromosomal rearrangements, and DNA damage. Nuclear blebs (i.e., herniations of the nuclear envelope) have been induced by (1) nuclear compression, (2) nuclear migration (e.g., cancer metastasis), (3) actin contraction, (4) lamin mutation or depletion, and (5) heterochromatin enzyme inhibition. Recent work has shown that chromatin transformation is a hallmark of bleb formation, but the transformation of higher-order structures in blebs is not well understood. As higher-order chromatin has been shown to assemble into nanoscopic packing domains, we investigated if (1) packing domain organization is altered within nuclear blebs and (2) if alteration in packing domain structure contributed to bleb formation. Using Dual-Partial Wave Spectroscopic microscopy, we show that chromatin packing domains within blebs are transformed both by B-type lamin depletion and the inhibition of heterochromatin enzymes compared to the nuclear body. Pairing these results with single-molecule localization microscopy of constitutive heterochromatin, we show fragmentation of nanoscopic heterochromatin domains within bleb domains. Overall, these findings indicate that chromatin within blebs is associated with a fragmented higher-order chromatin structure.

Keywords:  Chromatin Motion; Chromatin Nanodomains; Heterochromatin; Lamins; Nuclear Blebbing

DOI:  https://doi.org/10.1242/jcs.262161
Nat Cancer. 2025 Jan 24.

TRIM28-dependent developmental heterogeneity determines cancer susceptibility through distinct epigenetic states.

PERMUTE

Mutations in cancer risk genes increase susceptibility, but not all carriers develop cancer. Indeed, while DNA mutations are necessary drivers of cancer, only a small subset of mutated cells go on to cause the disease. To date, the mechanisms underlying individual cancer susceptibility remain unclear. Here, we took advantage of a unique mouse model of intrinsic developmental heterogeneity (Trim28+/D9) to investigate whether early-life epigenetic variation influences cancer susceptibility later in life. We found that heterozygosity of Trim28 is sufficient to generate two distinct early-life epigenetic states associated with differing cancer susceptibility. These developmentally primed states exhibit differential methylation patterns at typically silenced heterochromatin, detectable as early as 10 days of age. The differentially methylated loci are enriched for genes with known oncogenic potential, frequently mutated in human cancers and correlated with poor prognosis. This study provides genetic evidence that intrinsic developmental heterogeneity can prime individual, lifelong cancer susceptibility.

DOI: https://doi.org/10.1038/s43018-024-00900-3
Nucleic Acids Res. 2025 Jan 24. pii: gkaf036. [Epub ahead of print]53(3):

The ortholog of human DNAJC9 promotes histone H3-H4 degradation and is counteracted by Asf1 in fission yeast.

Yan Ding, Jun Li, He-Li Jiang, Fang Suo, Guang-Can Shao, Xiao-Ran Zhang, Meng-Qiu Dong, Chao-Pei Liu, Rui-Ming Xu, Li-Lin Du.

Mammalian J-domain protein DNAJC9 interacts with histones H3-H4 and is important for cell proliferation. However, its exact function remains unclear. Here, we show that, in the fission yeast Schizosaccharomyces pombe, loss of Djc9, the ortholog of DNAJC9, renders the histone chaperone Asf1 no longer essential for growth. Utilizing AlphaFold-based structural prediction, we identified a histone-binding surface on Djc9 that binds to helix α3 of H3 in a manner that precludes simultaneous helix α3-binding by Asf1. Djc9 and Asf1 indeed compete for binding to the H3-H4 dimer in vitro, and an H3-α3 mutation impeding Djc9 binding also renders Asf1 non-essential, indicating that the role of Asf1 needed for growth in fission yeast is to prevent histone binding by Djc9. In the absence of Asf1, cell growth is hindered due to unrestrained Djc9-mediated downregulation of H3 and H4. In the presence of Asf1, Djc9 confers resistance to the DNA replication inhibitor hydroxyurea and dominant negative disease-related histone mutants by promoting the degradation of superfluous or dysfunctional histones. Our findings provide new insights into the function and mechanism of this conserved histone-binding protein.

DOI: https://doi.org/10.1093/nar/gkaf036
Biomolecules. 2025 Jan 04. pii: 64. [Epub ahead of print]15(1):

SNP rs9364554 Modulates Androgen Receptor Binding and Drug Response in Prostate Cancer.

Yuqian Yan, Lei Shi, Tao Ma, Liguo Wang, Haojie Huang.

  (1) Background: Prostate cancer treatment efficacy is significantly influenced by androgen receptor (AR) signaling pathways. SLC22A3, a membrane transporter, has been linked to SNP rs9364554 risk loci for drug efficacy in prostate cancer. (2) Methods: We examined the location of SNP rs9364554 in the genome and utilized TCGA and other publicly available datasets to analyze the association of this SNP with SLC22A3 transcription levels. We verified onco-mining findings in prostate cancer cell lines using quantitative PCR and Western blots. Additionally, we employed electrophoretic mobility shift assay (EMSA) to detect the binding affinity of transcription factors to this SNP. The ChIP-Seq was used to analyze the enrichment of H3K27ac on the SLC22A3 promoter. (3) Results: In this study, we revealed that SNP rs9364554 resides in the SLC22A3 gene and affects its transcription. The downregulation of SLC22A3 is associated with drug resistance. More importantly, we found that this SNP has different binding affinities with transcription factors, specifically FOXA1 and AR, which significantly affects their regulation of SLC22A3 transcription. (4) Conclusions: Our findings highlight the potential of using this SNP as a biomarker for predicting chemotherapeutic outcomes and uncover possible mechanisms underlying drug resistance in advanced prostate cancers. More importantly, it provides a clinical foundation for targeting FOXA1 to enhance drug efficacy in prostate cancer patients.

Keywords:  AR; FOXA1; SLC22A3; SNP rs9364554; drug efficacy; prostate cancer

DOI:  https://doi.org/10.3390/biom15010064
PLoS Biol. 2025 Jan 27. 23(1): e3003004

Trithorax regulates long-term memory in Drosophila through epigenetic maintenance of mushroom body metabolic state and translation capacity.

Nicholas Raun, Spencer G Jones, Olivia Kerr, Crystal Keung, Emily F Butler, Kumari Alka, Jonathan D Krupski, Robert A Reid-Taylor, Veyan Ibrahim, MacKayla Williams, Deniz Top, Jamie M Kramer.

The role of epigenetics and chromatin in the maintenance of postmitotic neuronal cell identities is not well understood. Here, we show that the histone methyltransferase Trithorax (Trx) is required in postmitotic memory neurons of the Drosophila mushroom body (MB) to enable their capacity for long-term memory (LTM), but not short-term memory (STM). Using MB-specific RNA-, ChIP-, and ATAC-sequencing, we find that Trx maintains homeostatic expression of several non-canonical MB-enriched transcripts, including the orphan nuclear receptor Hr51, and the metabolic enzyme lactate dehydrogenase (Ldh). Through these key targets, Trx facilitates a metabolic state characterized by high lactate levels in MBγ neurons. This metabolic state supports a high capacity for protein translation, a process that is essential for LTM, but not STM. These data suggest that Trx, a classic regulator of cell type specification during development, has additional functions in maintaining underappreciated aspects of postmitotic neuron identity, such as metabolic state. Our work supports a body of evidence suggesting that a high capacity for energy metabolism is an essential cell identity characteristic for neurons that mediate LTM.

DOI: https://doi.org/10.1371/journal.pbio.3003004