UC Davis

Orofacial clefts are the most common birth defect worldwide. One key component of craniofacial development is the formation of the palate separating the oral and nasal cavities, which is a complex process that requires extensive coordination of many factors and signaling pathways that if disrupted may result in cleft palate. I employed single-cell RNA sequencing to describe the transcriptome of the palate and identify the various cell populations that comprise the developing secondary palatal shelves, to gain a deeper insight into their regulatory mechanisms. Through these efforts, I identified key cell populations that contribute to the medial edge epithelium, which facilitates fusion between the paired palatal shelves. I further described novel marker genes of the MEE and found that the Tgf-β receptors I and II are necessary for their specification. Wnt5a is an essential palatogenetic regulator necessary in both palate epithelium and mesenchyme. I show that Wnt5a antagonizes proliferation at the anterior palate, but in contrast with prior reports, does not promote proliferation at the posterior. I then worked to describe several of the key Wnt5a effectors that are dysregulated due to its loss of function, both at the transcriptional level using single-cell data, and at the protein level through immunostaining and immunoblot techniques. Finally, I demonstrated that mesenchymal Wnt5a contributes to activation of Rho family GTPases in the developing palate, and that their ectopic activation can partially rescue the Wnt5a mutant phenotype in culture. I also show that epithelial β-catenin is not required for secondary palate development in multiple models. Contrary to prior reports, the MEE is appropriately specified, and the palate completely fuses in its absence.