Yanding Zhang

Shox2-deficient mice exhibit a rare type of incomplete clefting of the secondary palate

The short stature homeobox gene SHOX is associated with idiopathic short stature in humans, as seen in Turner syndrome and Leri-Weill dyschondrosteosis, while little is known about its close relative SHOX2. We report the restricted expression of Shox2 in the anterior domain of the secondary palate in mice and humans. Shox2-/- mice develop an incomplete cleft that is confined to the anterior region of the palate, an extremely rare type of clefting in humans. The Shox2-/- palatal shelves initiate, grow and elevate normally, but the anterior region fails to contact and fuse at the midline, owing to altered cell proliferation and apoptosis, leading to incomplete clefting within the presumptive hard palate. Accompanied with these cellular alterations is an ectopic expression of Fgf10 and Fgfr2c in the anterior palatal mesenchyme of the mutants. Tissue recombination and bead implantation experiments revealed that signals from the anterior palatal epithelium are responsible for the restricted mesenchymal Shox2 expression. BMP activity is necessary but not sufficient for the induction of palatal Shox2 expression. Our results demonstrate an intrinsic requirement for Shox2 in palatogenesis, and support the idea that palatogenesis is differentially regulated along the anteroposterior axis. Furthermore, our results demonstrate that fusion of the posterior palate can occur independently of fusion in the anterior palate.

Transgenically ectopic expression of Bmp4 to the Msx1 mutant dental mesenchyme restores downstream gene expression but represses Shh and Bmp2 in the enamel knot of wild type tooth germ.

Bmp4 is a downstream gene of Msx1 in early mouse tooth development. In this study, we introduced the Msx1-Bmp4 transgenic allele to the Msx1 mutants in which tooth development is arrested at the bud stage in an effort of rescuing Msx1 mutant tooth phenotype in vivo. Ectopic expression of a Bmp4 transgene driven by the mouse Msx1promoter in the dental mesenchyme restored the expression of Lef-1 and Dlx2 but neither Fgf3 nor syndecan-1 in the Msx1 mutant molar tooth germ. The mutant phenotype of molar but not incisor could be partially rescued to progress to the cap stage. The Msx1-Bmp4 transgene was also able to rescue the alveolar processes and the neonatal lethality of the Msx1 mutants. In contrast, overexpression of Bmp4 in the wild type molar mesenchyme down-regulated Shh and Bmp2 expression in the enamel knot, the putative signaling center for tooth patterning, but did not produce a tooth phenotype. These results indicate that Bmp4 can bypass Msx1 function to partially rescue molar tooth development in vivo, and to support alveolar process formation. Expression of Shh and Bmp2 in the enamel knot may not represent critical signals for tooth patterning.

Msx1 is required for the induction of Patched by Sonic hedgehog in the mammalian tooth germ

We have used the mouse developing tooth germ as a model system to explore the transmission of Sonic hedgehog (Shh) signal in the induction of Patched (Ptc). In the early developing molar tooth germ, Shh is expressed in the dental epithelium, and the transcripts of Shh downstream target genes Ptc and Gli1 are expressed in dental epithelium as well as adjacent mesenchymal tissue. The homeobox gene Msx1 is also expressed in the dental mesenchyme of the molar tooth germ at this time. We show here that the expression of Ptc, but not Gli1, was downregulated in the dental mesenchyme of Msx1 mutants. In wild‐type E11.0 molar tooth mesenchyme SHH‐soaked beads induced the expression of Ptc and Gli1. However, in Msx1 mutant dental mesenchyme SHH‐soaked beads were able to induce Gli1 but failed to induce Ptc expression, indicating a requirement for Msx1 in the induction of Ptc by SHH. Moreover, we show that another signaling molecule, BMP4, was able to induce Ptc expression in wild‐type dental mesenchyme, but induced a distinct expression pattern of Ptc in the Msx1 mutant molar mesenchyme. We conclude that in the context of the tooth germ Msx1 is a component of the Shh signaling pathway that leads to Ptc induction. Our results also suggest that the precise pattern of Ptc expression in the prospective tooth‐forming region is controlled and coordinated by at least two inductive signaling pathways. Dev Dyn 1999;215:45–53.

Timing of odontogenic neural crest cell migration and tooth-forming capability in mice.

The mammalian tooth develops through sequential and reciprocal interactions between cranial neural crest (CNC)‐ derived ectomesenchymal cells and the stomadial epithelium. Classic tissue recombination studies demonstrated that premigratory CNC cells and CNC‐derived ectomesenchymal cells possess odontogenic capacity and can respond to oral epithelial signals to form a tooth, suggesting that the CNC cells contributing to odontogenic tissue are not prespecified. Here we show that, in mice, CNC cells have populated the forming first branchial arch before the 9‐somite stage and continue to migrate into the arch by the 13‐somite stage. Grafts of the first arch from the 10‐somite embryo or earlier yielded membranous bone and cysts but no teeth after subrenal culture. However, grafts of the first arch with its dorsally adjacent tissue containing migrating neural crest cells from the same age embryos gave rise to teeth. In contrast, teeth formed in first arch grafts that do not contain migrating neural crest cells from embryos with 12 or more somites. Interestingly, the acquisition of tooth forming capability in the first arch coincides with the onset of Fgf8 expression in the oral epithelium. These results suggest that there exists a population of odontogenic neural crest cells that migrates into the first arch between the 10‐ and 12‐somite stages. These cells either possess odontogenic potential and are able to initiate tooth development, or can respond to odontogenic signals derived from the oral epithelium to support tooth formation. Developmental Dynamics, 2003.

Expression and regulation of the chicken Nkx-6.2 homeobox gene suggest its possible involvement in the ventral neural patterning and cell fate specification†

Rapid accumulating evidence has suggested that the homeodomain transcription factors of the Nkx family play important roles in controlling vertebrate organ patterning and differentiation. In this study, we report the cloning, expression and regulation of a novel chicken homeobox gene, cNkx‐6.2, whose expression is similar, but not identical, to that of mouse Nkx‐6.2. The earliest expression of cNkx‐6.2 was detected at the neural plate stage in the prospective midbrain and hindbrain regions. As the neural development proceeds, cNkx‐6.2 expression was restricted in the ventral region of the entire neural axis except the forebrain region. At late stages of development, cNkx‐6.2 expression is downregulated in the ventricular neuroepithelial cells, but subsequently upregulated in a subpopulation of cells. Tissue recombination and explant culture experiments demonstrated that expression of cNkx‐6.2 can be induced by the notochord signal and purified SHH protein, and repressed by BMP‐4 and ‐7, indicating that the cNkx‐6.2 expression can be influenced by both ventral and dorsal midline signals. Taken together, these studies have suggested two different roles for the cNkx‐6.2 transcription factor: participating in the Shh‐initiated ventral patterning during early CNS development and controlling cell fate specification and differentiation during late development. Dev Dyn 1999;216:459–468. ©1999 Wiley‐Liss, Inc.