UCSF

Identifying molecular signals mediating the epithelial-mesenchymal interactions (EMI) required for bone formation in the embryonic jaw skeleton could lead to the discovery of novel proteins with therapeutic potential for regenerating bone in cases of disease and injury. An RNA-seq strategy was used to identify candidate genes involved in the EMI of the mandibular primordia, which give rise to the lower jar. RNA in situ hybridization and RT-qPCR were used to characterize the spatiotemporal expression of these genes qualitatively and quantitatively on the mRNA levels in three avian species (duck, quail, and chicken). In vitro organ cultures were used to test if the expression of these genes in mandibular mesenchyme requires epithelial signaling. Gain- and loss-of-function experiments tested if these genes regulate mandibular osteogenesis. Results from the RNA-seq experiment, RNA in situ, and RT-qPCR reveal that spatiotemporal changes in members of the CXC and WNT signaling pathways are present at the right time, place, and levels to mediate the osteogenic EMI in the mandibular primordia. In vitro organ cultures confirmed that mesenchymal expression of these depends on epithelial signaling. Gain-of-function experiments revealed that CXCL14 is sufficient to augment bone formation compared to the non-treated side after 7 days of culture. Loss-of-function experiments demonstrated that WNT signaling is required for mandibular osteogenesis but that CXCL14 can restore bone formation in the absence of WNT signaling. Moreover, altering the WNT pathway affects CXCL14 expression, suggesting that CXCL14 acts downstream of WNT ligands.