Figure 5

Synergistic effect of rspo2 and rspo3 ablation on zebrafish tooth development and Meckel’s cartilage. (A) Flat-mount images of Alcian blue/Alizarin red S stained 9 dpf zebrafish ventral cartilages. Zoom of pharyngeal teeth to right. rspo3^−/− larvae displayed anterior shortening of Meckel’s cartilage, which was exacerbated with rspo3^−/−; rspo2^Δ gRNA disruption (black bars). *indicate absent teeth. Scale bar: 200 μm. (B) Alizarin red S staining of pharyngeal teeth shows that rspo3^−/− are generally normal relative to wild-type while rspo2^Δ larvae have a reduced number of teeth (average of 2 versus 8). Tooth number in rspo2^Δ larvae decreased further with decreasing wild-type alleles of rspo3 (zero teeth detected in rspo3^−/−; rspo2^Δ mutant). (C) Quantification of the anterior–posterior/rostral length of Meckel’s cartilage shows a primary effect in rspo3^−/− larvae, which is exacerbated in rspo3^−/−; rspo2^Δ mutants. (D) Quantification of the anterior–posterior length of ceratohyal cartilage shows no effect in rspo3^−/− larvae, suggesting a cartilage element-specific effect of rspo3 and rspo2. N = 10–16. p < 0.01. *Indicates significance relative to wild-type.