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Evolution’s Fast Track

Eyeless cavefish reveal mechanisms of cryptic genetic variation

By STEPHANIE DUTCHEN and MATT FEARER
December 16, 2013

Surface (top) and cave (bottom) populations of the fish Astyanax mexicanus differ in many physical traits, including the cavefish’s loss of eyes and pigmentation. Image: Nicolas Rohner

Thousands of years ago, a population of Astyanax mexicanus fish in northeastern Mexico swam or was swept from its hospitable river home into harsh underwater caves and became trapped. Facing a dramatically different environment of near total darkness and hardly any food, the fish had to adapt—fast.

Among other changes, the “cavefish” swiftly (in evolutionary terms, within just a few thousand years) lost their pigmentation—and their eyes.

A research team led by Harvard Medical School geneticists argues that the loss of eyes was beneficial, helping the cavefish reallocate their limited metabolic resources to biological functions that would be more helpful in their new setting. Indeed, even as their sight and pigmentation diminished, the fish developed heightened senses of smell and taste and grew more whisker-like structures called neuromasts that detect changes in water movement.

How did the cavefish adapt so quickly? According to the researchers, part of the answer is that the fish were able to take an evolutionary shortcut by calling on a phenomenon known as “cryptic” or “standing” genetic variation.

The team reports Dec. 12 in Science that the shock of being exposed to a new environment depleted a stress-response protein called HSP90 in the cavefish, which in turn unmasked existing but previously silent genetic variants for traits such as smaller eye size. That one proved adaptive, the researchers say, and even after HSP90 returned to full power, subsequent generations of fish inherited smaller and smaller eyes until their eyes disappeared altogether.

The work provides the first evidence that HSP90 can facilitate morphological evolution in a natural setting rather than only in a lab.

“It’s a very cool story in terms of the speed of evolution,” said Nicolas Rohner, a postdoctoral researcher in the lab of HMS Department of Genetics head Clifford Tabin and first author of the paper. “By studying these exceptional fish, we believe we’ve identified an additional mechanism explaining how evolution can work.”

Fast track

The classical view of evolution holds that organisms experience spontaneous, or de novo, genetic mutations that produce various novel traits. Nature then selects for the most beneficial, and those get passed along to subsequent generations. It’s an elegant model, but it’s also extremely time-consuming and doesn’t help species that need to cope with sudden, potentially life-threatening changes in their environments.

With standing genetic variation, by contrast, genetic mutations arise and are passed along within a given population but are normally kept silent. The physical manifestations of the mutations don’t emerge unless a population encounters stressful conditions, like being forced to live in a dark cave instead of a vibrant river.

De novo mutations occur after an organism arrives in its new environment. They’re slow, and they favor dominant traits,” explained Rohner. “Standing variation provides a pool of mutations that are already available in the whole population. When organisms find themselves in a new environment, the silent variations are released and nature can select the ones that help.”

How standing mutations collect in a population and become available for use, and how much those mutations contribute to natural selection in complex organisms, have remained in dispute.

Whitehead Institute Member Susan Lindquist had discovered that heat shock protein 90, or HSP90 for short, can buffer and release standing genetic variation in organisms, including fruit flies, yeast and plants. She showed that the normally robust cellular reservoir of HSP90 becomes taxed in response to environmental stress. Its depletion allows phenotypic changes to emerge rapidly. Some emergent traits found in her lab were not adaptive, while others clearly were.

Meanwhile, Tabin’s lab, which is interested in developmental and evolutionary biology, was studying the genetics of eye loss in cavefish. One recent finding showed that certain populations of cavefish lost schooling behavior and that this could be traced to both sight-dependent and sight-independent factors.

The labs formed a partnership to find out whether HSP90 helped spur cavefish evolution.

The experiments

Along with other collaborators, the team conducted a series of experiments with cavefish and surface fish of the same species.

First, the researchers mimicked environmental stress among developing fish by reducing HSP90 using the inhibitor Radicicol. Surface fish raised in the presence of Radicicol displayed significantly more variation in eye size than untreated fish, as did treated fish crossbred from surface and cave parents, indicating that reducing HSP90 did reveal standing genetic variants for eye size among the surface fish.

The team then looked at the effects of Radicicol exposure on adult surface fish and cavefish. Although they don’t have eyes, cavefish still have eye sockets, so the researchers measured those. Once again, the surface fish displayed a strikingly wider than usual range of eye and eye socket sizes. The cavefish, however, showed no difference in socket size range. This supported the team’s idea that the cavefish had undergone natural selection for smaller eye size. At the same time, the treated cavefish’s average socket size was smaller. This showed that the genetics governing eye size remained responsive to HSP90.

To test whether chemically induced HSP90 reduction mimicked what would happen in a natural setting, the researchers compared a host of conditions—including pH, oxygen content and temperature—in the surface and cave waters that are home to these fish. They discovered a considerable difference in conductivity, as measured by salinity, between cave and surface. Because low conductivity can trigger a heat shock response, they raised surface fish in water with low conductivity comparable to that in one of the caves.

The fish showed a comparable HSP90 response as those exposed to Radicicol and once more displayed significant variation in eye size—demonstrating that an environmental stressor could indeed have the same effects as chemical inhibition of HSP90.

“This is the first time that we can see in a natural setting where the stress came from and observe the variation that results,” said Tabin, who is also the HMS George Jacob and Jacqueline Hazel Leder Professor of Genetics.

Additional experiments showed that raising the fish in the dark without other interventions did not affect eye size, and that reducing HSP90 did not reveal standing genetic variation in body size or neuromast number.

“Geneticists are really interested in evolution and how it occurs,” said Rohner. “This work offers more evidence about one mechanism that will hopefully help us better understand the whole process.”

Swimming ahead

The Tabin lab continues to pursue other cavefish studies. One avenue involves investigating the genetic basis of differences in metabolism.

“Cavefish have adapted to starvation,” said Rohner. “There’s basically no food in these caves unless a flood sweeps some in or bat guano falls into the water. These fish can go months without eating and will hardly lose any body fat. They don’t go to sleep; they still move, slowly. No one knows how they’re storing fat so efficiently.”

The lab also wants to look at the cavefish’s brains to see whether, because the fish have lost their optic lobes along with their eyesight, other areas may have grown larger over time.

In addition to revealing more about these odd creatures, such research could illuminate broader principles about how organisms—including humans—adapt to new environments, said Rohner.

“We’re all evolving. We and our environments are changing,” said Rohner. “The impact of environment and ecology on speciation is something we have to understand more about, for the conservation of animals and possibly also for ourselves.”

This work was supported by the National Institutes of Health (R01 HD047360).

Adapted from a Whitehead Institute news release.

 

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