Scientists find clues to how tiny fish ‘pauses’ its life

Stanford scientists have identified molecular drivers that put the “pause” in “diapause,” a life stage of the African killifish that suspends its development as an embryo.

- By Hanae Armitage

Anne Brunet is the senior author of a study that examines the molecular driver of the African killifish's ability to enter a state of suspended animation.
Gregg Segal

Eight months out of the year, new clutches of African killifish must execute the ultimate act of patience, lest they perish in the sun.

As an embryo, the small, yellowish-turquoise fish, which are native to the African savanna, undergo something called diapause in the dry season. It’s a survival tactic that suspends their barely-formed bodies in time, and mud, as they wait for the right conditions to emerge. Now, researchers at the Stanford University School of Medicine have discovered a key molecular driver of the fish’s unusual early-life interlude, helping them better understand how this tiny animal can pause its own development to procure life later.

The African killifish dwells in ephemeral ponds of the African savanna, which come into being during the rainy season. But when the pools dry up, as they eventually do, the killifish have no place to go. “So over a very, very, long time, these killifish evolved the remarkable ability to survive the loss of water through diapause — akin to hibernation, but for embryos,” said Anne Brunet, PhD, professor of genetics.

 Not really asleep, and not awake either, during diapause, embryonic killifish — which have already grown the beginnings of a full organ system — are essentially frozen in time at a basal level of existence. Brunet’s team set out to decipher the genetics behind the sci-fi-like phenomenon, finding that the developmental lull was actually spurred by a flurry of genomic activity.

A paper detailing the findings of the study was published Feb. 21 in Science. Brunet is the senior author, and postdoctoral fellow Chi-Kuo Hu, PhD, is the lead author.

Within the spurt of molecular activation, Brunet and her team pinpointed a gene responsible for proper muscle maintenance — crucial for the embryo’s ability to pick up where it left off developmentally after diapause is over.

They also found one more interesting nugget: Even after remaining in diapause for months — sometimes years — the killifish in the study experienced no detriment to their health. There was no evolutionary trade-off; the rates of growth, fertility, and aging were the same as killifish that did not undergo diapause. So, essentially a killifish that’s gone through diapause, compared to one that hasn’t, Brunet said, has extended its total life.

“Diapause lasts around five months, about the same as an average African killifish lifespan. But some killifish have stayed in diapause for 2½ years,” she said. “If you think about that in human terms, that’s like if we were to exist, paused as an embryo, for some 400 years, only to resume natural development and live out a full life.”

Stop in the name of life

To understand this diapause state in the laboratory, Brunet and her team set up lab plates filled with dirt and embryonic killifish. Killifish in diapause are not much to look at on the outside; their bodies are encased by a protective sac, making them appear as little whiteish spheres. But through RNA sequencing technology, which provides insight into gene activity, Brunet saw that these embryos were much more interesting on the inside. 

Brunet and her team put embryonic killifish in plates filled with dirt to study the animals in diapause.
Brunet lab

“Overall, we saw a 30% increase in gene activity during diapause,” Brunet said. As the team got into the nitty-gritty, it narrowed its focus to an especially active gene, CBX7, which functions in something called the polycomb complex. In this complex, CBX7 is a sort of master regulator of muscle maintenance throughout diapause. 

In killifish models that lacked CBX7, muscles withered, and fish came out of diapause early, something that would likely prove fatal in the wild. Interestingly, said Brunet, the polycomb complex is found in animals other than killifish, including humans. But it plays different roles.

“It’s possible that other animals — such as mice — have vestigial diapause that could be tapped into,” Brunet said. “Our lab works on stem cells, and we think a lot about stem cell quiescence, a sort of state of stem cell dormancy within tissues. It’s likely that some of the genes that are active in diapause in the killifish are conserved in other species, so you could imagine that tapping into this network may be able to help inform tissue preservation or the preservation of stem cells in a tissue in other animals.” 

Brunet and her team are careful not to draw direct parallels between killifish and humans just yet, but she’s intrigued by the thought. “As time passes, our organs progressively degenerate, especially in disease,” she said. “So identifying the general, fundamental mechanisms of organ preservation could be important to understanding how to counter the normal atrophy of organs over time or under disease conditions.”

Other Stanford authors of the paper are postdoctoral scholar Param Singh, PhD, and graduate student Adam Reeves.

This study was funded by the National Institutes of Health (grants DP1AG044848, T32 CA930235, T32GM00779040 and CIHRPJT-153049), the Life Science Research Foundation and the Glenn Laboratories for the Biology of Aging.

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