Researchers from Harvard University and Boston Children’s Hospital have discovered that certain gene mutations in zebrafish lead to the formation of limb-like structures in the fish’s pectoral fins, a major development that suggests the same genes likely play a role in human skeletal formation.

In the study published Thursday in Cell, a team led by Michael Brent Hawkins, a recent doctoral recipient from the Department of Organismic and Evolutionary Biology at Harvard Medical School, found that mutations to the waslb and vav2 genes were driving zebrafish to develop joints, muscle and extra layers of bone in their fins. 

Zebrafish are known to have overly simple skeletons — one bone per fin, and never any accompanying joints or muscle.

"It's stunning that with a single mutation, we are able to generate such a coordinated change in the shape of the fin, and likely its function, with new bones, joints and muscles,” said Hawkins.

The waslb and vav2 genes, found in all vertebrates, are studied extensively for cancer and immunology research, but this is the first time they have been shown to play a part in the limb-generation process.

Because humans also possess these genes, Hawkins believes that this discovery could be vital in understanding how to treat human skeletal disease, and potentially even take steps toward human limb regeneration.

"If a fish is able to make more of a limb type of structure, what is possible for humans to be able to do in terms of aging or regeneration or other things?" Hawkins asked.

For the study, Hawkins obtained his subjects from a separate research project led by Katrin Henke, a researcher in the Department of Genetics at Harvard Medical School. She screened around 10,000 lab-cultivated fish, identifying various mutations that were the result of cutting-edge CRISPR-Cas9 gene editing technology.

This tool allows scientists to cut desired DNA strands in a genome and make changes to it. The changes are called mutations, and by comparing a non-mutant animal with its mutant counterpart, scientists can more easily understand the exact behavior of the respective gene that was altered. 

Hawkins zeroed in on the screened zebrafish with evidence of fin-to-limb transformation, and by comparing the mutants with wild-type, or non-mutant, zebrafish, he saw that the two unexpected genes, waslb and vav2, were responsible. 

"With our additional genetic experiments, once we started playing around with other genes we know control different parts of limb patterning, we found that we can actually push this effect even further and make more levels of bones," Hawkins said. "We ended up with a fish that can actually have three and four levels of bone, instead of one."

As per the team’s work, waslb and vav2 mutations directly affect the expression of the Hox gene, which is what typically dictates limb formation or regression. To corroborate the novel discovery further, the researchers experimented with editing the same genes in mice. The result of knocking the genes out, or fully removing their activity, led to dramatic deficiencies in the mice’s skeletal structure. 

"Altogether, this is showing us that the gene is controlling expression of the Hox genes," Hawkins explained. "It's not just a fish thing; this is definitely a mammal thing. It's very related to us."

As discussed in the study, zebrafish and humans have a common ancestor with an intermediate skeletal complexity. Through evolution, the zebrafish lineage simplified the skeleton while the human lineage augmented it. 

Realizing that mutations to latent genes in fish brought about such a significant outcome — the creation of bones, joints and muscle — also brings forth questions about what latent genes the human body might have that are waiting to be activated. 

Hawkins remarked, “What if we have this genetic ability that's latent and hidden, but can become activated with a mutation?"

The article “Latent developmental potential to form limb-like structures in zebrafish” was published Feb. 4 in Cell. The authors of the study were Michael Brent Hawkins, Harvard Medical School; Katrin Henke, Boston Children’s Hospital and Matthew P. Harris, Harvard University. The lead author was Michael Brent Hawkins.