Single-cell transcriptomic analysis of craniogenesis in control and SOXC-mutant mice

SOXC genes (i.e., Sox4, Sox11 and Sox12) are important members of the SOX transcription factor family. They are involved in several developmental processes, including skeleton formation. However, their role in intramembranous ossification and, particularly, in skull development has not beed investigated yet. Here, we employed single-cell RNA-seq technology to better understand the roles of the SOXC genes in craniogenesis and how their inactivation could affect this process. We identified the expression of the SOXC genes in several cranial populations, including osteodermal and meningeal progenitors. Removal of the SOXC genes in the cranial mesenchyme with a Prx1Cre transgene showed changes in the proportions of progenitor clusters and deriving populations (dermal, hypodermal, periosteal, suture and bone cells). Mutant progenitors showed the downregulation of genes involved in cytoskeleton organization, cell adhesion and migration, cell proliferation and steorl biosynthesis. As a result, SOXC-mutant fetuses have underdeveloped bones, sutures, dermis and hypodermis, particularly in the apical regions. 10X Genomics single-cell RNA-sequencing profiling of cranial tissues from control and SOXC-mutant embryos at different developmental stages (i.e., E11.5, E12.5, 13.5, E15.5 and E17.5). For the E17.5 stage the dermis and hypodermis were removed and only cranial bones were used.SOXC基因（即Sox4、Sox11与Sox12）是SOX转录因子家族（SOX transcription factor family）的重要成员。它们参与包括骨骼形成在内的多种发育过程。然而，其在膜内成骨（intramembranous ossification）中，尤其是颅骨发育中的作用尚未被研究。本研究采用单细胞RNA测序（single-cell RNA-seq）技术，以更深入解析SOXC基因在颅发生（craniogenesis）中的功能，以及其失活如何影响该过程。我们在多个颅骨细胞群中检测到SOXC基因的表达，包括骨-真皮祖细胞（osteodermal progenitors）与脑膜祖细胞（meningeal progenitors）。利用Prx1Cre转基因（Prx1Cre transgene）元件敲除颅间充质（cranial mesenchyme）中的SOXC基因后，可观察到祖细胞簇及其衍生细胞群（真皮、皮下组织、骨膜、骨缝与骨细胞）的比例发生改变。突变祖细胞中，参与细胞骨架组织、细胞黏附与迁移、细胞增殖以及类固醇生物合成（sterol biosynthesis）的基因表达下调。因此，SOXC突变胎儿的骨骼、骨缝、真皮与皮下组织发育不全，尤其在顶区更为显著。本数据集包含对不同发育阶段（即E11.5、E12.5、E13.5、E15.5与E17.5）对照组与SOXC突变胚胎的颅骨组织进行的10X Genomics单细胞RNA测序分析。在E17.5阶段，我们移除了真皮与皮下组织，仅采用颅骨样本开展实验。