The zebrafish dlx2b 52 4-kb genomic region recapitulates the endogenous dlx2b pattern with expression in pharyngeal tooth germs and pectoral fins, but not oral epithelium. Shown are whole-mount, lateral views of the Tg(dlx2b:EGFP)^wj1 transgenic line, anterior to the left. (A) 58 hpf mRNA in situ hybridization against the GFP reporter message showing expression in a pharyngeal tooth germ (arrow). (B) GFP expression in a living 72 hpf larva in pharyngeal tooth germs (arrow). (C) Reporter expression in the margin of the caudal fin at 31 hpf. (D) Pectoral fin expression at 49 hpf. (Scale bars: A–C, 100 μm; D, 50 μm.)

The zebrafish dlx2b:GFP reporter drives expression in developing A. mexicanus oral teeth, pharyngeal teeth, and pectoral fins. (A) Lateral view of the head at 75 hpf with expression in an oral tooth germ (arrow), upper pharyngeal tooth germ (double arrowhead), and a small ectopic location (arrowhead). (B) Lateral view of pharyngeal tooth (arrow) and pectoral fin (arrowhead) expression at 99 hpf. (C) Oral tooth forming in the upper jaw adjacent to the developing premaxillary bone at 125 hpf. The boundary of the dental epithelium can be seen on one side (double arrowhead) and two groups of dlx2b:GFP reporter expressing cells on the other (arrows). (D) GFP expression in dental epithelial cells (arrow) surrounding a mineralized tooth attached to the dentary bone of the lower jaw (arrowhead) at 125 hpf. (Scale bars: A and B, 100 μm; C and D, 10 μm.)

Dlx gene knockdown perturbs zebrafish pharyngeal tooth morphogenesis. (A) High magnification view of a pharyngeal tooth at 100 hpf in a control injected with the injection solution and no MO. (B) A normal-looking tooth from an identically treated specimen injected with a dlx2b MO. (C) After injection of combined dlx2a and dlx2b MOs, pharyngeal tooth size is reduced (representative example shown). (D) Pharyngeal teeth are even smaller, and are now misshapen, after simultaneous injection with MOs against dlx2a, dlx2b, dlx3b, and dlx5a. (Scale bar: 10 μm.)

Fgf inhibition eliminates dlx2b:GFP reporter expression in zebrafish pharyngeal tooth germs. Tg(dlx2b:EGFP)^wj1 treated from 30–72 hpf with either a DMSO control solution (A) or the Fgf inhibitor SU5402 (B). Pharyngeal tooth expression of the dlx2b:GFP reporter is eliminated with this treatment (arrows). Pectoral fin expression (arrowhead) is also absent, but no pectoral fin bud develops and this therefore may be the result of earlier Fgf requirements in fin development. (Scale bars: 100 μm.)

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