Developmental genetic mechanisms of evolutionary tooth loss in cypriniform fishes

Abstract

The fossil record indicates that cypriniform fishes, a group including the zebrafish, lost oral teeth over 50 million years ago. Despite subsequent diversification of feeding modes, no cypriniform has regained oral teeth, suggesting the zebrafish as a model for studying the developmental genetic basis of evolutionary constraint. To investigate the mechanism of cypriniform tooth loss, we compared the oral expression of seven genes whose mammalian orthologs are involved in tooth initiation in the zebrafish and the Mexican tetra, Astyanax mexicanus, a related species retaining oral teeth. The most significant difference we found was an absence in zebrafish oral epithelium of expression of dlx2a and dlx2b, transcription factors that are expressed in early Astyanax odontogenic epithelium. Analysis of orthologous genes in the Japanese medaka (Oryzias latipes) and a catfish (Synodontis multipunctatus) suggests that expression was lost in cypriniforms, rather than gained in Astyanax. Treatment of Astyanax with an inhibitor of Fibroblast growth factor (Fgf) signaling produced a partial phenocopy of the zebrafish oral region, in that oral teeth, and expression of dlx2a and dlx2b, were lost, whereas shh and pitx2, genes whose expression is present in zebrafish oral epithelium, were unaffected. We hypothesize that a loss of Fgf signaling to oral epithelium was associated with cypriniform tooth loss.

Transgenic analysis of Dlx regulation in fish tooth development reveals evolutionary retention of enhancer function despite organ loss

“…The ability of the zebrafish dlx2b:GFP reporter construct to drive expression in A. mexicanus oral tooth germs indicates that loss of oral Dlx2b expression in cypriniforms resulted from changes in one or more trans-acting regulators rather than in the cis-regulatory regions of this gene. This interpretation is consistent with the coordinate loss of the closely related paralog Dlx2a from cypriniform oral epithelium, a gene likely to be regulated by many of the same trans-acting factors (19). Some of these changes might involve other members of the Dlx gene family.…”

“…5) suggests redundancy in the teleost dentition as well. Interestingly, despite the expression of Dlx genes during zebrafish pharyngeal tooth initiation (20) and the fact that dlx2b prefigures the epithelium from which oral teeth arise in A. mexicanus (19), none of the phenotypes we obtained from gene knockdown involved the early arrest of tooth initiation, as seen in the mouse (22). This result might be explained by the more extensive redundancy of Dlx gene expression during tooth initiation in the zebrafish relative to the mouse (20).…”

“…Dlx2 expression is known to be regulated by Bmp4 in mouse tooth development (15), but whether dental Dlx2b expression in teleosts is regulated by Bmp2b remains to be determined. Fgf signaling has been shown to be upstream of Dlx2b expression in both the oral and pharyngeal dentition of teleosts (12,19). In addition, inhibition of Fgf signaling in A. mexicanus prevents oral tooth development and produces a partial phenocopy of the zebrafish oral gene expression profile (19).…”

“…Dlx genes are expressed at numerous stages of tooth development in the zebrafish (20) and the mouse (21), and loss of Dlx function leads to early arrest of tooth development in the mouse (22). We have argued previously that Dlx genes are likely downstream of the cause of cypriniform oral tooth loss (19) and are thus a good model for the fate of developmental pathways once an initiation signal has been modified in evolution.…”

Evolution and developmental diversity of tooth regeneration

“…The zebrafish (Danio rerio) is a highly derived Teleost species of the order Cypriniformes and over the past 30 years has become the top fish developmental model owing to its transparency during embryogenesis, the ease of vast embryo production and the development of genomic tools. The zebrafish has become a useful model for tooth development and manipulation [8] however the tiny teeth of zebrafish and teeth in all Cypriniformes are confined to the back of the gill arches, embedded deep within the pharyngeal cavity, with oral teeth in this order having been lost approximately 50 million years ago [9].…”