Spatiotemporal single-cell atlas of suture stem cell dynamics in craniosynostosis [scRNA-Seq]

Craniosynostosis is a congenital craniofacial disorder marked by premature suture fusion and aberrant skull morphogenesis. Yet the cellular dynamics and regulatory mechanisms of suture mesenchymal stem cells (SuSCs) in this disease remain largely unknown. Here, we present a comprehensive spatiotemporal atlas of SuSCs integrating single-cell and 2 μm-resolution spatial transcriptomics in Fgfr2C342Y/+, a model that recapitulates features of human Crouzon syndrome, alongside wild-type controls across three key developmental stages (E14.5, E18.5, and P3). By integrating morphology-based cell segmentation with machine-learning classification, we achieved single-cell spatial resolution in mapping Visium HD data. Leveraging this resource, we reveal disruptions in SuSC fate, niche composition, and intercellular signaling across developmental stages in craniosynostotic sutures. Downstream analyses identify Foxa3 as a master transcriptional regulator of these fate shifts and implicate specific immune- and suture meningeal fibroblast–derived signals in driving pathological ossification. Together, our work fills a gap in disease-model single-cell spatial-omics, providing a high-resolution roadmap of SuSC dynamics in craniosynostosis and uncovering candidate molecular targets for future therapeutic strategies.