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Chewing on the mysteries of the jaw
Scientists in the University of Southern California (USC) Stem Cell laboratory of Gage Crump have revealed how key genes guide the development of the jaw in zebrafish. These findings may offer clues for understanding craniofacial anomalies in human patients, who sometimes carry a mutation in equivalent genes.
In the study published in Developmental Cell, first author Lindsey Barske and colleagues report that two related genes, Nr2f2 and Nr2f5, regulate stem cell timing in order to generate skeletal cells.
Just like in humans, fish skeletons generally start out as cartilage later to be replaced by bone. However, most zebrafish upper jaw bones develop without a cartilage template. Nr2f genes prevent stem cells in the developing upper jaw from becoming cartilage, keeping them available for making bone later. By contrast in the lower jaw, the Endothelin1 (Edn1) gene prevents Nr2f activity allowing for extensive formation of early cartilage, driving outgrowth of the lower jaw.
"Our study illustrates the idea that development of any organ requires a balance between early maturation and maintenance of stem cells. Without inhibitory signals like those from the Nr2f genes, there probably wouldn't be enough uncommitted precursors left over to make later-forming cell types or maintain adult tissues."
Prior to this work, little was known about the signals that initiate patterning of the upper jaw. The scientists took a novel genomics approach and identified new genes important for upper jaw development by carefully defining all the genes expressed during early jaw development. Then using powerful gene editing, they removed many of those genes from the genome to discover that zebrafish mutants, lacking several Nr2f genes, display a second cartilage-based lower jaw — where the upper jaw should be.
"The power of this approach is that hundreds of genes can be functionally tested in a cost-effective, rapid manner in zebrafish, thus allowing us to assign new functions for the many poorly characterized genes in the genome."
• The upper jaw transforms into a lower-jaw-like structure in Nr2f mutant zebrafish
• Nr2fs globally inhibit lower-jaw gene expression in upper-jaw precursors
• Reducing Nr2f dosage fully rescues lower-jaw defects in endothelin1 mutants
• Nr2fs drive jaw asymmetry by limiting cartilage differentiation in the upper jaw
The jaw is central to the extensive variety of feeding and predatory behaviors across vertebrates. The bones of the lower but not upper jaw form around an early-developing cartilage template. Whereas Endothelin1 patterns the lower jaw, the factors that specify upper-jaw morphology remain elusive. Here, we identify Nuclear Receptor 2f genes (Nr2fs) as enriched in and required for upper-jaw formation in zebrafish. Combinatorial loss of Nr2fs transforms maxillary components of the upper jaw into lower-jaw-like structures. Conversely, nr2f5 misexpression disrupts lower-jaw development. Genome-wide analyses reveal that Nr2fs repress mandibular gene expression and early chondrogenesis in maxillary precursors. Rescue of lower-jaw defects in endothelin1 mutants by reducing Nr2f dosage further demonstrates that Nr2f expression must be suppressed for normal lower-jaw development. We propose that Nr2fs shape the upper jaw by protecting maxillary progenitors from early chondrogenesis, thus preserving cells for later osteogenesis.
Co-authors include: Lindsey Barske, Pauline Rataud, Kasra Behizad, Lisa Del Rio, Samuel G. Cox, J. Gage Crump.
Seventy-five percent of the research was supported by $262,500 in federal funding from the National Institute of Dental and Craniofacial Research (R01 DE018405, R35 DE027550 and K99 DE026239), and 25 percent by $87,500 of non-federal funding from the A.P. Giannini Foundation. The USC Office of Research and the USC Norris Medical Library funded the bioinformatics software and computing resources.
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GREEN labels skeletal stem cells in the embryonic zebrafish head, and MAGENTA labels early-forming cartilaginous facial skeleton. Image credit: Beat Fierz/EPFL