FoxN3 knock-down alters spatio-temporal expression patterns of genes involved in cranial cartilage and muscle development in the African clawed-frog Xenopus laevis

What is the gene regulatory network underlying the morphogenesis of the vertebrate head skeleton? A good candidate to approach this question is the FoxN3 gene. Morpholino-mediated FoxN3 knock-down leads to a delay in cranial cartilage and muscle formation, to reduced size and to malformations of both cartilages and muscles in Xenopus laevis . Additionally, muscle fiber development and joint formation are incomplete. We used qRT-PCR and whole-mount in situ hybridization to analyse potential target genes of FoxN3. The spatio-temporal expressions of different cartilage, muscle, and joint markers as well as cell adhesion molecules are changed following FoxN3 knock-down. Expression of N-CAM and N-Cad is decreased throughout development and expression of genes important for cartilage formation (Sox9, Col2α1, Runx2) is delayed. Joint markers (Gdf5/6) and genes (Dlx5/6) important for regional specification are also down-regulated and bagpipe genes show decreased expression at consecutive stages. Expression levels of key myogenic genes (Myf5, MyoD, MHC) are at 30 - 40% of control expression. These results indicate that FoxN3 occupies a key position in the gene regulatory network maintaining normal development of cranial cartilages and muscles as well as jaw joint formation.