Notch signaling in the embryonic ectoderm promotes periderm cell fate and represses mineralization of vibrissa hair follicles
The Notch signaling pathway is a critical means to regulate cell face choice in animals.  Appropriate regulation of this pathway is also required for human face formation as both loss and gain of function mutations of Notch signaling can cause craniofacial syndromes.  Here we examine the consequences of manipulation of Notch signaling in the early mouse embryonic ectoderm by either removing the transcriptional effector Rbpj or expressing a constitutively active form of the Notch1 intracellular domain.  Loss of Rbpj resulted in cleft secondary palate but strikingly was also associated with the ectopic mineralization of vibrissal follicles.  In contrast, activation of Notch signaling resulted in multiple embryonic defects including a fully penetrant bilateral cleft lip and palate.  Further, single cell RNA-seq data indicated a switch from a basal epithelial cell identity towards periderm when Notch signal transduction was elevated.  These cell fate changes were accompanied by misregulation of genes and pathways known to impact human and mouse orofacial clefting including Grhl3, Irf6, and the Wnt pathway.  Together, these findings provide insight into human craniofacial conditions caused by misregulated Notch activity. Overall design: Single-cell RNA sequencing was performed on microdissected regions of the upper face where the nasal and maxillary prominences are juxtaposed, including the lambdoid junction, nasolacrimal groove, and nasal fin, from E11.5 control and NotchGOF mutant embryos. For each condition, tissue from two embryos with 43 somites was pooled prior to library preparation. Control and mutant samples were processed separately and sequenced as independent datasets. A total of 11,618 cells were obtained for the control dataset (NotchGOF_Control) and 12,220 cells for the mutant dataset (NotchGOF_mutant). Cells were sequenced to an average depth of 38,111 reads per control cell and 36,122 reads per mutant cell.