Tooth development in frogs: Implications for the re-evolution of lost mandibular teeth and the origin of a morphological innovation

Background:

Many structures are conserved evolutionarily across a wide range of organisms to a surprising degree. One such feature is teeth, calcified structures found in the jaws of many vertebrates. Teeth first evolved over 500 million years ago in jawless fishes. Since then, some animals have evolved toothlessness, such as birds which have developed beaks, and anteaters which use their tongues to eat ants, rendering teeth unnecessary.

Frogs are an interesting part of this group, as they have (mostly) evolved toothlessness on their lower jaw. However, of the nearly 8000 species of living frogs, only one – Guenther’s marsupial frog (Gastrotheca guentheri) – was found to have teeth on both of its jaws. These teeth are thought to have been lost in the common ancestor of the frog more than 200 million years ago. Guenther’s marsupial frog therefore presents an exception to Dollo’s law of irreversibility, which poses that if an organism loses a particular trait during evolution, it will never reverse back to that state even if it finds itself in identical conditions to where it once lived.

Unfortunately, Guenther’s marsupial frog was last sighted in 1996 and is currently feared to be extinct due to habitat loss, and fewer than 50 museum samples exist of this animal across the world. Due to this, it wasn’t possible for the authors of this preprint to study this frog directly, so instead they used the Cuban tree frog.

Like the majority of frogs, the Cuban tree frog develops keratinised mouthparts during their larval (tadpole) stage of development (Figure 1). The development and genes responsible for the formation of these mouthparts are poorly understood, and it’s not known how these mouthparts might affect odontogenesis (tooth development) in frogs.

The authors of this preprint looked at the Cuban tree frog in order to understand how Guenther’s marsupial frog re-evolved mandibular teeth. More specifically, the aims of this preprint were to:

1. Determine the precise timings of odontogenesis in the upper jaw of Cuban tree frogs.
2. Investigate whether odontogenesis is at all initiated in the lower jaw to understand how Guenther’s marsupial frog re-evolved mandibular teeth.
3. See if initial tooth development occurs normally, or whether the keratinised mouthparts of the tadpoles affect this.

Main Findings:

Initiation of tooth development occurs as expected in the upper jaw

The authors found that tooth development occurred normally in the upper jaw of the Cuban tree frog. Genes that are known to mediate tooth development, Pitx2, Shh, Sox2, CTNNB1, and Dlx2 were all found at the final stages of tadpole development, when tooth development is initiated. Interestingly, Pitx2, CTNNB1, and Dlx2  were expressed in cells that support the jaw sheaths and keratodonts of the tadpole feeding apparatus.

At later developmental stages, teeth on the upper jaw developed further as the tadpole feeding apparatus atrophied and detached. Here, the jaw extended posteriorly, moving closer to the eye, and the first tooth buds developed, but did not yet erupt through the overlying tissue.

Is odontogenesis initiated in the lower jaw?

The authors wanted to investigate whether any progression of odontogenesis took place and was subsequently abandoned in the lower jaw. If this was the case, it may provide an explanation as to how Guenther’s marsupial frog re-evolved mandibular teeth.

They found that in the final tadpole stages, the lower jaw epithelia looked similar to that of the upper jaw before tooth development occurred. However, no localised epithelial thickening, a hallmark of tooth development, was observed. However, gene expression in the keratinised mouthparts of the lower jaw were comparable to that of the upper jaw of the same stage. Especially surprising was the presence of Pitx2, as this is one of the most crucial drivers of the tooth gene regulatory network. Further investigation into later stages of development also showed no evidence of odontogenesis taking place in the lower jaw of the Cuban tree frog.

Do the keratinised mouthparts affect tooth development?

Frogs first develop a ‘primary mouth’ as an embryo and then form a unique feeding apparatus during their tadpole stages of development. These structures don’t involve teeth, but most have keratinised feeding structures (with a few exceptions, notably Xenopus). Teeth are later formed during metamorphosis, when their initial feeding structures are atrophied. The authors found that the same genes are present during primary mouth formation and tadpole mouthpart development as in the upper jaw when tooth development starts. This presence may indicate that these genes play a role in the formation of the primary mouth, that they could be involved in the development of the keratinised mouthparts found in tadpole feeding structures, or that they could be involved in making the tissue competent for tooth formation at a very early stage in tadpole development (much earlier than when teeth are seen). More research needs to be done to see what, if any, effect the keratinised structures have on odontogenesis in frogs.

Next steps?

These findings show that the tadpole keratinised mouthparts have an interesting developmental progression, and since this is the first data on the gene expression in these structures, it’s clear that more research should be done to unravel their full involvement in teeth formation in frogs. A more in-depth understanding of how these structures develop may also provide the answer to how mandibular teeth re-evolved in Guenther’s marsupial frog.

Why I am highlighting this preprint:

In school, we learned about evolution on an organism scale, but didn’t really touch on how genes and developmental processes may be conserved across species. The idea that this process was lost and somehow re-emerged is, I think, a really cool concept. Also, I have never really considered tooth development in frogs (I honestly didn’t even know frogs had teeth!), so I found this preprint interesting as it was so far outside of my own field of research.

Questions for the authors:

1. Why did you choose to look at tooth development in the Cuban tree frog, and not a frog that is more closely related to Guenther’s marsupial frog? What differences would you expect to see in a more closely related species rather than the Cuban tree frog?
2. Since the keratinised feeding apparatus of tadpoles has similar signalling as seen in primary tooth development in the upper jaw, do you think it is possible that the ‘original’ tooth development machinery could be found here rather than in the jaw at later stages of frog development?
3. To what extent do different frog species exhibit variation in tooth development pathways? Are there common evolutionary patterns?

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