A zebrafish. Credit: Marrabbio2
A zebrafish. Credit: Marrabbio2

Coloured Cells Chase Each Other To Make A Fish’s Stripes

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This astonishing GIF shows a microscopic chase scene: A black cell flees from the touch of a yellow cell, and the yellow cell goes after it.

On their own, the two cells go round and round. But if there are lots of them, the yellow cells end up corralling the black ones into long bands. And that, according to Hiroaki Yamanaka and Shigeru Kondo from Osaka University, is why zebrafish gets its stripes. In all of his Just-So Stories, Rudyard Kipling never imagined anything like this—an animal pattern that results from hundreds of cellular pursuits, played out over the landscape of a skin.

Here’s a longer video, complete with the most appropriate music I could find.

Kondo’s work has a long pedigree that began with the English mathematician Alan Turing. When Turing wasn’t changing the face of computer science or breaking German codes in WWII, he was thinking about animal patterns. In 1952, he proposed a simple mathematical model involving two molecules: an activator that produces a pattern, and an inhibitor that blocks it. Both diffuse through the skin, and react with each other. By evolving small changes in how quickly these molecules spread and how strongly they interact, animals can produce radically different patterns, from cheetah spots to zebra stripes (see for yourself with this Java applet).*

Turing’s ideas about “reaction-diffusion systems” were based on abstract maths. But in recent decades, scientists like Kondo have shown that many animal patterns behave exactly as he predicted.

In 1995, Kondo’s team showed that as semicircle angelfish grow up, the stripes on their bodies get further apart and new ones appear in the widening gaps. In 2007, they showed that zebrafish can replace lost cells on its skin, but the new patterns are different to the old ones. In 2010, they showed that a black-spotted fish and a white-spotted one can breed to produce hybrids with bizarre, maze-like patterns. All of this suggests that the patterns aren’t generated according to some fixed blueprint. Instead, they’re produced by something drifting across the fishes’ skins, just as Turing suggested.

Then: a plot twist. Several groups studied mutant zebrafish with leopard-like spots instead of zebra-ish stripes, and identified the genes responsible for these weird patterns. But none of these genes produce the kinds of diffusible molecules that Turing envisioned. Instead, they make proteins that are embedded within the outer membranes of cells. Maybe, instead of drifting molecules, the skin cells themselves are moving around.

To test this idea, Yamanaka and Kondo harvested two types of pigmented cells from the fins of zebrafish: black melanophores and yellow xanthophores. On their own, both types of cell move in random directions. But when Yamanaka and Kondo mixed the cells, they saw something astonishing.

The yellow cells speed up, and actively extended finger-like projections called pseudopodia towards the black ones. Upon contact, the black cells recoil and run away, only to be pursued by the yellow ones. And since the black cells are still slightly faster, the result is a continuous “run-and-chase movement”. (For context, the black cells move at just over 2 micrometres (millionths of a metre) per hour. They’re around 50 micrometres wide, so it takes them a day to cover their own length.)

If hundreds of these chases play out across a crowded skin, Yamanaka and Kondo believe that the yellow cells would collectively push black ones away, resulting in clearly defined stripes of dark and light. And that, of course, is exactly what you see in a normal zebrafish.

And it’s not what happens in mutant zebrafish with weird skin pattern. In the so-called jaguar mutants, which have fuzzy stripes, the black cells are less attractive to the yellow cells and less strongly repulsed by them. The two groups of cells move in the normal way on their own. It’s just their interactions that are different. Their lazier pursuits mean that they don’t segregate as neatly, which leads to fuzzier stripes.

Meanwhile, in the leopard mutants, which have spots instead of stripes, the black cells don’t flee from the yellow cells at all. Instead, they move towards them. The result is an embrace rather than a chase. Isolated black cells are killed off by the yellow ones, while those that randomly cluster together find safety in numbers and survive. The result: black spots in a sea of yellow.

At first glance, this seems very different to what Turing suggested. Rather than moving molecules that activate or inhibit the production of colour, you have moving cells that are themselves coloured. But there are similarities too. In Turing’s model, the two molecules react with one another, and both diffuse at different speeds. Here, the yellow and black cells certainly interact, and they move at different speeds.

But why exactly are the yellow cells attracted to the black ones, and why are the black ones repelled? How do the mutations behind the jaguar and leopard patterns change these movements? And why do the cell chases almost always go in an anticlockwise spiral?

And perhaps most importantly, how do these chases actually play out in the skin of a fish? “In their model, they assume a random initial distribution of xanthophores and melanophores. Such a situation has not been shown to exist in the fish at any time point in the development of the stripes,” says Christiane Nüsslein-Volhard, a Nobel laureate who studies animal patterns among other things. Her team has also found that the stripes on a zebrafish’s body depend on another type of pigmented cell—the iridophores—that Kondo’s team haven’t studied yet.

Still, Nüsslein-Volhard praises the team for developing a way of harvesting pigmented cells from zebrafish and testing their interactions. Their study leaves lots of unanswered questions, but also some tools for addressing them.

Update: I’ve got a late-arriving comment from Florian Maderspacher, who is currently editor of Current Biology but who used to work in this field.

“This basically confirms what we had found ten years ago: the jaguar gene acts mainly in melanophores, while leopard acts is both cell types. What is new here is that they can correlate the mutants with altered cell behaviours in a culture dish. But the fact that it’s in a culture dish [means] we have no real way of knowing that the cell behaviours they document are really what underlies stripe formation. They say they are similar to what they see in the organism, but they don’t show direct evidence for that. In an extreme case, they might just be looking at a behaviour that is the result of these cells being put out of context. It would be interesting to see if the cells can actually form any sort of pattern in the dish; so far there is no indication for that.”

Reference: Yamanaka & Kondo. 2013.  In vitro analysis suggests that difference in cell movement during direct interaction can generate various pigment patterns in vivo. PNAS http://dx.doi.org/10.1073/pnas.1315416111

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