Mutant clownfish reveal how nature pushes boundaries

In 1999, the clown fish (Amphiprion ocellaris) is hatched in the waters of a tropical hobbyist in the UK.

These clown fish are very popular with aquarists for their unique pattern of three straight white bars bordered by a thin black line. But this UK fish was special: instead of the usual straight bars, it had a wavy, corrugated pattern, symmetrical on both sides. These patterns are passed down through the generations, leading to a lineage called “Snowflake,” but the mechanism behind this unusual pattern remains a mystery.

Twenty years later, researchers at the Okinawa Institute of Science and Technology (OIST), Academia Sinica in Taiwan, Kyoto University, and the University of Virginia have finally revealed the exact gene responsible for the change, and in the process, have revealed clues to the great mystery of how nature creates patterns. Their findings were published in Nature Communication.

“In theory, it should be simple,” says Professor Vincent Laudet of the Marine Eco-Evo-Devo Unit at OIST. How does a cell know that it should be black, white, or orange, so that it always creates clearly organized patterns on a macroscale?

From single amino acid changes to universal models

Snowflake fish are similar to other clownfish in every way except for their unusual patterning, which makes them a perfect model for genetic pigmentation research. To understand why they look the way they do, researchers turned to another fish. Zebrafish are best studied for their horizontal stripes, which keep equal in size and spacing as the fish grows. Thanks to another mutation, “Leopard” zebrafish, which produces spots instead of stripes, researchers have identified one of the genes involved in the pattern. It includes a special gap protein that acts as a telephone line between cells, allowing them to exchange information in the form of electrical currents and small molecules.

When the team compared Snowflake’s genome to wild clownfish, they found striking similarities. Laudet says: “We saw it right away – Snowflake had exactly the same transformation as Leopard!”

However, this eureka led to more questions than answers. In zebrafish, a gap junction protein was thought to be responsible for driving the so-called growth of the Turing pattern. Named after the famous British mathematician Alan Turing, this model explains how the protein combines the structure of lines by preventing short-term contact while promoting long-term interaction between pigmentation cells, which leads to the development of similar patterns in zebrafish.

But Turing’s model could not account for the clownfish model, as their stripes are fixed in order and position throughout their life, meaning that information is exchanged to explain where and when the bar should be formed. The first author of this study, Dr. Marleen Klann from the Marine Eco-Evo-Devo Unit, continues: “We found that the gap junction protein is not responsible for Turing patterning in zebrafish, and showed that it ensures clear cell-to-cell communication in general.

This primary evidence points researchers to a universal biological mechanism for organizing cells at the border. The team turned to another area to better understand the principles governing the organization of pigmentation cells in clownfish: membrane physics. Here, they found that the so-called Edwards-Wilkinson model is the simplest possible model that correctly accounts for the formation and disruption of the clean boundary between pigmentation cells in clownfish and more.

“The model describes two forces,” explains co-author Professor Simone Pigolotti of the Biocomplexity Unit at OIST. One is surface tension, which favors a smooth layer. Another is noise, which has the opposite effect.

Pigolotti says: “This model is a general tool that allows us to understand what we have observed as well as provide information on where to look next, including all kinds of organisms, which can help us clarify general principles.

Laudet concludes: “It is thanks to Snowflake and our transgenic anemonefish, which we successfully developed with Professor Masato Kinoshita of Kyoto University, that we are now close to understanding the incredibly complex mechanisms of the unusual task of cellular organization.”