Role of LADs and Epigenetic Pathways in Nuclear Architecture During Early Mouse Embryo Development

By Pooja Toshniwal PahariaReviewed by Lexie CornerMay 6 2025

A recent study published in Cell investigates how lamina-associated domains (LADs) contribute to nuclear genome organization in early-stage mouse embryos.

Focusing on the zygote and two-cell stages, the researchers examined the interplay between chromatin structure, nuclear architecture, and epigenetic pathways. Their findings show how the nuclear lamina influences genome positioning and stability during early development.

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Background

The three-dimensional (3D) organization of the genome within the nucleus is essential for regulating key DNA-dependent processes, including transcription. LADs are large genomic regions, ranging from approximately 100 kilobases to 10 megabases, that interact with the nuclear lamina, contributing to the structural integrity of the nucleus and influencing genome function.

LADs generally replicate late during the synthesis (S) phase of the cell cycle, in contrast to inter-LAD regions (iLADs), which replicate earlier. While LADs are absent in mature oocytes, they form rapidly after fertilization. By the zygote stage, around 40 % of the mouse genome exhibits stable LAD positioning, suggesting that large portions of the genome are spatially organized early in development.

Study Overview

In the present study, researchers investigated the role of LADs in shaping nuclear architecture during early mouse embryonic development, focusing on the zygote and two-cell stages. They also explored the epigenetic factors influencing LAD formation and their involvement in restructuring LADs during the maternal-to-zygotic transition (MZT).

To identify molecular components involved in LAD development, the team conducted a gain-of-function screen targeting nuclear proteins in mouse embryos. They first removed the zona pellucida using pronase and then separated polar bodies with trypsin, isolating 10–20 blastomeres per embryo for analysis. These samples were processed for Illumina sequencing, with raw reads trimmed and demultiplexed using specialized software.

Sequencing data were aligned to the GRCm38 mouse genome, and low-quality reads were filtered out. Unique reads were binned for LAD analysis. Chromatin profiling was performed using several techniques, including CUT&RUN, CUT&Tag, and LaminB1-DamID.

The resulting datasets were submitted to the GEO database for public access and further analysis. To examine the spatial organization of chromatin and confirm changes in LAD positioning, the researchers used fluorescence in situ hybridization (FISH).

Ribonucleic acid (RNA) sequencing libraries revealed gene expression profiles in the embryos. Two-state hidden Markov models (HMM) distinguished LAD domains from inter-LADs. Principal component analysis (PCA) grouped embryos based on their developmental stage and perturbations. Researchers calculated LaminB1-DamID scores to categorize LAD phenotypes and analyze genome-wide correlations. They investigated whether LAD phenotypes from chromatin modifiers relate to ZGA and assessed modifier effects on LAD establishment during the early and late two-cell stage.

Results

The study found that mouse embryos retain the capacity to reorganize LADs at the two-cell stage, even after earlier disruptions. This suggests that LAD formation in the zygote is not essential for maintaining nuclear organization in later stages.

Perturbations that affect chromatin structure, such as histone modifications and nuclear protein overexpression, produce a variety of altered LAD phenotypes. Methylation of histones H3K27 and H3K9 influenced LAD reorganization, with increased methylation associated with chromatin compaction and gene silencing.

Trimethylation of H3K27 (H3K27me3) emerged as a key factor in defining LAD boundaries, with evidence that H3K4me3 restricts the spread of H3K9me3, thereby helping to establish LAD limits.

Overexpression of H3K9me3 writers and readers disrupted normal LAD structure, causing abnormal fragmentation or, in some cases, more stable LAD formation. Most chromatin modifiers—except HP1γ—led to increased lamina association in zygotes, indicating that heterochromatin expansion promotes aberrant LAD interactions.

The study also revealed how changes in LaminB1-Dam methylation levels affect LAD positioning. Regions gaining methylation moved toward the nuclear lamina, while those losing it shifted inward. LADs that either collapsed or converted into iLADs exhibited weaker interactions with the lamina, indicating reduced structural stability.

RNA sequencing suggested that ZGA contributes to maintaining nuclear architecture in two-cell embryos. The expression of genes during ZGA correlated with specific histone modifications, reinforcing the link between epigenetic regulation and spatial genome organization.

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Conclusion

The findings show that LADs in mouse embryos are dynamic and can be re-established even after initial disruptions. While LAD formation begins shortly after fertilization, it is not strictly required for subsequent nuclear organization, as embryos retain the capacity to restructure these domains during the two-cell stage.

Epigenetic mechanisms—particularly histone modifications like H3K27me3—play a key role in shaping LAD boundaries and guiding their interactions with the nuclear lamina. The study highlights how chromatin modifiers and histone content influence nuclear organization differently in zygotes and two-cell embryos.

Ultimately, multiple chromatin pathways contribute to LAD formation and restructuring during early development and zygotic gene activation.

Journal reference

Pal et al., The establishment of nuclear organization in mouse embryos is orchestrated by multiple epigenetic pathways, Cell, 2025, 188: 1–20, DOI: 10.1016/j.cell.2025.03.044, https://www.cell.com/cell/fulltext/S0092-8674(25)00396-4