van Knippenberg, S. S. F. A.

bioRxiv2025

SMARCA2/4-Dependent Chromatin Remodelling Establishes Gene Regulatory Programs in Early Human Embryos and Blastoids

Establishment of cell lineages during development is regulated by transcription factors binding at cis-regulatory elements to activate gene regulatory programs. Many transcription factors require chromatin remodellers for accessibility at their target sites. Although these key principles of gene regulation in mammalian development have emerged, the contribution of chromatin remodellers to early human embryogenesis remains unknown. Here, we show that the SWI/SNF ATPases SMARCA2 and SMARCA4 are required for establishing the epiblast and trophectoderm fates during human pre-implantation development and facilitate accessibility at regulatory elements. We find that degradation of SWI/SNF ATPases disrupts epiblast formation in blastoids and enhances trophectoderm specification, while also showing transcriptional and chromatin misregulation in TE-like cells. In human embryos, SWI/SNF perturbation impaired blastocyst formation and the establishment of the inner cell mass. Single-nucleus chromatin accessibility and transcriptome profiling in blastoids reveals that the SWI/SNF complex safeguards the naive epiblast and trophectoderm programs and facilitates enhancer and transcription factor motif accessibility. These findings identify SWI/SNF chromatin remodellers as critical regulators of embryonic lineage specification during human pre-implantation development.

developmental biology↗

bioRxiv2025

Feeder-free culture of naive human pluripotent stem cells retaining embryonic, extraembryonic and blastoid generation potential

Conventional human pluripotent stem cells (hPSCs) are widely used to study early embryonic development, generate somatic cells, and model diseases, with differentiation potential aligned to a post-implantation epiblast identity. In the past decade, naive hPSCs, representing a pre-implantation stage, have been derived. Naive hPSCs efficiently differentiate towards embryonic and extraembryonic lineages such as trophectoderm, primitive endoderm, and extraembryonic mesoderm, and also self-organize into blastocyst-like structures called blastoids. However, their culture typically relies on mouse embryonic fibroblasts (MEFs), which are variable, resource-intensive, and can confound analyses. We report the long-term maintenance of naive hPSCs in a feeder-free, serum-coated system. We successfully expanded for up to 25 passages 8 different naive hPSCs lines across 5 laboratories. Growth rate, clonogenicity, and gene expression profiles on serum coating were comparable to MEF-based cultures, but serum coating eliminated fibroblast contamination. Naive hPSCs cultured on serum exhibited more efficient germ layer specification, retained trophectoderm potential and high blastoid formation efficiency. Exome sequencing revealed fewer mutations in serum-cultured cells, and mass spectrometry identified extracellular matrix proteins such as vitronectin, fibronectin, and collagens in the serum coating. Overall, serum coating offers a scalable, cost-effective and therefore widely applicable alternative for naive hPSC culture, maintaining developmental potential, reducing DNA mutations, and eliminating MEF-related confounding factors. We believe serum coating will expand the use of naive hPSCs to large-scale studies and facilitate the investigation of mechanistic insights into developmental and disease modelling.

cell biology↗