Single-Cell Rna Sequencing Revealed Heterogeneity Of Suture Progenitors

Suture mesenchymal stem cells (SMSCs) residing within cranial suture niches are critical for skull development and regeneration. However, their cellular heterogeneity and downstream differentiation trajectories remain poorly characterized due to the limited accessibility of suture tissues and low abundance of osteogenic progenitor cells. We employed Gli1CreERT2;Ai14 mouse models with tamoxifen induction at postnatal day 4 (P4), followed by flow cytometry sorting of tdTomato+ cells at P7 (n=10). Single-cell RNA sequencing (scRNA-seq) was performed using both SMART-Seq and 10× Genomics platforms. In situ hybridization validated spatial expression patterns. Comparative analysis evaluated sequencing depth, and CytoTrace2 predicted differentiation hierarchies. CellChat and SCENIC inferred intercellular communication and transcription factor (TF) networks, respectively. SMART-Seq detected 4,500 genes/cell (vs. 3,200 by 10×), enabling more accurate differentiation state predictions via CytoTrace2. SCENIC identified Erg as a key TF in SMSCs, with downstream targets enriched in mesenchymal stem cell development. E2F7/8 emerged as critical regulators of proliferative progenitors, linked to cell cycle progression. This study identifies SMART-Seq demonstrates superior resolution for rare cell population analysis, offering novel insights into suture biology and regenerative medicine, and predict Erg and E2F7/8 as master regulators of SMSC and proliferative progenitors, respectively.