Uhrf1 loss disrupts Ctcf-associated chromatin organization during early mouse embryogenesis

DPF3a couples H3K36me2-dependent chromatin remodeling to genome architecture during myogenesis

The findings uncover a non-canonical mechanism whereby a chromatin remodeler regulates transcription primarily through three-dimensional genome organization rather than local accessibility control, and establish histone modification-guided chromatin remodeling as a key principle in gene regulation.

The CTCF Paralog BORIS Contributes to the Ovarian Cancer Transcriptional Program by Relaxing CTCF-mediated 3D Genome Organization

Disruption of three-dimensional genome architecture is a major driver of cancer initiation and progression, frequently arising from genetic and epigenetic alterations at CTCF- and cohesin-bound chromatin anchors. These same chromatin loop anchors can also be occupied by the germ cell-specific CTCF paralog CTCFL (BORIS), which is aberrantly activated in multiple malignancies. Here, we show that in ovarian cancer cells, BORIS activation establishes a distinct transcriptional program that collapses following loss of BORIS chromatin binding and is accompanied by widespread changes in CTCF and cohesin occupancy, histone modifications, and chromatin accessibility. These BORIS-dependent transcriptional alterations occur in long-range genomic clusters, resulting in the coordinated activation or repression of neighboring genes, including hormonally regulated gene families. BORIS loss also increases topologically associating domain (TAD) insulation, strengthens A/B compartment segregation and chromatin loop interactions, and results in a more compact and constrained chromatin architecture. Together, our findings suggest that aberrant BORIS activation promotes transcriptional reprogramming by weakening CTCF-mediated chromatin insulation and relaxing three-dimensional genome organization in ovarian cancer. Significance High-grade serous ovarian carcinoma, the most common subtype of ovarian cancer, remains one of the deadliest gynecological malignancies because of its late diagnosis, extensive genomic instability, and frequent therapeutic resistance. The lack of reliable biomarkers for early detection underscores the need to identify new molecular drivers of disease initiation and progression. Here, we show that the germline-specific gene CTCFL (BORIS) is aberrantly activated in high-grade serous ovarian carcinoma, where it remodels transcriptional, epigenetic, and three-dimensional genome organization by occupying CTCF-bound chromatin loop anchors and weakening CTCF-mediated chromatin insulation. These findings identify BORIS as a key regulator of transcriptional reprogramming in ovarian cancer and support its further investigation as both a prognostic biomarker and a potential therapeutic target.

Alterations in chromatin organization promote totipotent-like features in a DPPA2/DUX-dependent manner

2-cell like cells (2CLC) are a transiently cycling population of cells with totipotent-associated features. Although CTCF depletion induces 2CLC conversion in mouse ESC, whether this reprogramming is a consequence of disrupted higher-order chromatin organization or of CTCF-specific functions remained unclear. Here, we show that depletion of the cohesin release factor WAPL in ESC also promotes 2CLC reprogramming, which is increased by CTCF co-depletion. Single-cell RNA-seq/ATAC-seq analyses in CTCF/WAPL-depleted ESC revealed that chromatin accessibility precedes 2C-associated gene expression. Moreover, we identified ARID3A as a transcription factor that regulates the extent of 2CLC conversion following WAPL/CTCF depletion. Although WAPL or CTCF depletion induces distinct transcriptional changes in human ESC, these do not resemble transcriptional programs of early human embryogenesis, suggesting limited evolutionary conservation. Finally, we demonstrate that 2CLC conversion mediated by alterations in chromatin organization depends on the DPPA2/DUX axis and correlates with nucleolar integrity. Together, these findings establish a mechanistic link between higher-order chromatin organization and totipotency-like cell identity in mice. Higher-order chromatin organization has been implicated in the regulation of totipotency, but the mechanisms linking chromatin architecture to totipotent-like cell states remain poorly understood. This study shows that disruption of chromatin organization by degron-mediated CTCF and/or WAPL depletion promotes totipotent-like conversion through the DPPA2/DUX axis, accompanied by changes in chromatin accessibility and nucleolar integrity. WAPL depletion promotes 2CLC reprogramming in mouse ESC, and is enhanced by CTCF co-depletion. The transcription factor ARID3A limits the extent of 2CLC conversion following CTCF/WAPL-depletion. Increased chromatin accessibility precedes expression of 2C-associated transcripts following CTCF/WAPL depletion. 2CLC conversion induced by CTCF/WAPL depletion depends on the DPPA2/DUX axis and correlates with nucleolar integrity. WAPL or CTCF depletion in human ESC fails to activate transcriptional programs characteristic of early human embryogenesis, suggesting limited evolutionary conservation. WAPL depletion promotes 2CLC reprogramming in mouse ESC, and is enhanced by CTCF co-depletion. The transcription factor ARID3A limits the extent of 2CLC conversion following CTCF/WAPL-depletion. Increased chromatin accessibility precedes expression of 2C-associated transcripts following CTCF/WAPL depletion. 2CLC conversion induced by CTCF/WAPL depletion depends on the DPPA2/DUX axis and correlates with nucleolar integrity. WAPL or CTCF depletion in human ESC fails to activate transcriptional programs characteristic of early human embryogenesis, suggesting limited evolutionary conservation. Chromatin architectural defects trigger the DPPA2/DUX program and nucleolar remodeling to promote totipotent-like cell conversion in mouse embryonic stem cells.

A barrier to human embryogenesis involves dysregulated 3D genome reprogramming and ZGA collapse.

Early embryonic arrest (EEA) is a critical impediment in assisted reproductive technology, leading to lack of viable embryos. Despite its prevalence, the molecular basis of EEA remains elusive, and the contribution of 3D genome reorganization to this arrest remains largely unexplored. Here, we present an integrated analysis of chromatin architecture and transcriptome dynamics in arrested human embryos during zygotic genome activation (ZGA) stage. Our results reveal extensive chromosomal compartment switching, significant alterations in topologically associating domains (TADs) features, and disrupted chromatin looping in arrested embryos. It suggests that 3D genome reorganization and ZGA failure occur in arrested embryos, marked by reduced abundance of ZGA-critical genes, precocious activation of late-stage developmental pathways, suppression of RNA processing and ribosome biogenesis, and compromised energy metabolism. Additionally, we hypothesize that several transcription factors, including KLF17 and several ZNF family members, serve as regulators in the observed structural or functional changes during developmental arrest. Collectively, our findings suggest that defective 3D genome reprogramming and transcriptomic dysregulation during ZGA are associated with human embryonic arrest at the eight-cell stage. This study advances our understanding of human preimplantation development and its development failures by providing a valuable resource and a conceptual framework for future investigations.

VEZF1 facilitates pluripotency exit by regulating developmental transcriptional programs and CTCF occupancy

Polycomb establishes TAD-scale H3K27me3 mega-domains to safeguard neuronal identity

During development, pluripotent stem cells generate diverse cell types through gene regulatory networks orchestrated by combinations of transcription factors (TFs). Following terminal differentiation, however, cellular identities become remarkably stable and resistant to TF-mediated perturbation, yet the mechanisms underlying this stability remain poorly understood. Here, we identify Polycomb repressive complexes (PRCs) as key regulators of neuronal identity maintenance. Although PRCs are well known for repressing the promoters of developmental genes during early cell fate specification, we unexpectedly find that PRC-mediated H3K27me3 expands into megabase-scale domains during neuronal maturation that align with topologically associating domains (TADs). These H3K27me3 “mega-domains” selectively encompass genes associated with alternative neural and non-neural lineages. While depletion of H3K27me3 in mature neurons has only modest effects on basal gene expression, it significantly increases neuronal activity-dependent c-FOS binding and induction of lineage-inappropriate genes within these mega-domains. Our findings reveal a previously unrecognized role for PRCs in establishing TAD-scale repressive chromatin domains during neuronal maturation, thereby safeguarding neuronal identity from external stimuli through broad silencing of alternative cell fate programs. Graphical Abstract