Tiny fish makes big splash in aging research at Stanford

Researchers disabled aging-associated genes in the short-lived African killifish, including one for an enzyme called telomerase, whose absence caused humanlike disease in the animal.

Anne Brunet and her colleagues have mapped the location of specific genes involved in aging and age-related diseases along the killifish’s chromosomes.

Gregg Segal

“Live fast, die old” maybe isn’t the catchiest motto. But, for the African turquoise killifish, it’s apt. The life span of the tiny fish can be measured in months, not years, and it does everything quickly: hatch, mature, breed and even age. It’s an example of life on extreme fast-forward.

This accelerated life cycle is a necessity when one makes one’s home in seasonal ponds that regularly evaporate, and the fact that the fish shares many biological characteristics with humans makes it a promising candidate for the study of aging and longevity. But until now, scientists didn’t have the necessary tools and information with which to conduct genetic studies.

Now, researchers at the Stanford University School of Medicine have mapped the location of specific genes involved in aging and age-related diseases along the killifish’s chromosomes. They’ve studied patterns of gene expression in its various tissues, and used genome-editing technology to mutate 13 genes thought to be associated with the aging process.

One gene in particular, a component of an aging-associated enzyme called telomerase, causes fish to develop a constellation of traits similar to those seen in humans lacking the enzyme.

This new biological tool kit, which the researchers have made publicly available, will make it possible to trace the effect of specific genetic changes on aging and the diseases that accompany it. Eventually, it may lead to ways to slow or perhaps even reverse human aging.

‘Best of both worlds’

Although the similarities between fish and humans may not be immediately evident, people have much more in common with the tiny, minnowlike creature than with other short-lived laboratory animals.

“This fish gives us the best of both worlds,” said postdoctoral scholar Itamar Harel, PhD. “As a vertebrate, it shares many critical attributes with humans, including an adaptive immune system, real blood and similar stem cell biology. At the same time, its very short life span mimics those of the laboratory worms, yeast and fruit flies that until now have served as the traditional models of aging research.”

The life span of the tiny fish can be measured in months, not years, and it does everything quickly: hatch, mature, breed and even age.

Gregg Segal

A short life span allows researchers to quickly assess the effect of genetic variations among different strains of fish. It also allows them to breed and raise hundreds of progeny for study within the span of months, rather than the many years required to conduct similar experiments in other vertebrates.

“The life span of a mouse can be as long as three to four years,” said Anne Brunet, PhD, professor of genetics. “This is close to the average length of a postdoctoral or graduate student position. This means that it would be very difficult for a researcher to conduct a meaningful analysis of aging in the mouse within a reasonable time period.”

Harel is the lead author of a paper describing the research. Brunet, an associate director of Stanford’s Paul F. Glenn Center for the Biology of Aging, is the senior author. The paper was published Feb. 12 in Cell.

The killifish is one of the world’s shortest-lived vertebrates, with some varieties living only four months. Old killifish display many characteristics of aging humans: declining fertility and cognitive function, a loss of muscle mass and an increasing likelihood to develop cancerous tumors.

Mapping aging-related genes

Harel and his colleagues benefited from the recent sequencing and assembly of the entire killifish genome by postdoctoral scholar and fellow lab member Berenice Benayoun, PhD, which will be described in a separate, upcoming study. Benayoun is also a co-author of the current study. The availability of the whole genome sequence allowed the researchers to identify and map the location of genes involved in aging and longevity along the animal’s chromosomes. They then used a recently described technique called genome editing to specifically mutate certain genes involved in biological processes that represent hallmarks of aging, including those that control decisions regulating how cells enter or exit the cell cycle, whether and how stem cells self-renew and how organisms maintain their genomes and proteins over time.

The killifish’s rapid life cycle meant that Harel was able to generate fish carrying the mutations within 30-40 days, and stable lines — that is, fish with the mutation stably integrated into all their cells, which they will then pass on to all their progeny — within about two to three months.

Now we have what is essentially a high-throughput vertebrate model for aging research.

One mutation in an enzyme called telomerase was of particular interest. Telomerase helps to maintain the length of protective DNA caps called telomeres found on the end of every chromosome. In most cells, telomeres shorten with every cell division. When they reach a certain minimum length, the cell stops dividing.

“There is a very strong link between telomere shortening and aging,” said Harel, “but the mechanism is not completely understood. Current animal models of aging, such as the mouse, have telomeres that are much longer than humans — about 50,000-100,000 nucleotides — making it difficult to compare with humans.”

Although telomerase is not normally active in most cells of an adult animal, it does work to maintain telomere length in rapidly dividing cells such as male germ cells, which make sperm, as well as tissues with high turnover such as blood and gut. In its absence, the telomeres of those cells shrink rapidly as they repeatedly divide.

Rapid results

In contrast to laboratory mice, the length of killifish telomeres, which average around 6,000-8,000 nucleotides, is similar to that of humans. As a result, Harel and his colleagues were able to quickly see the effect of a telomerase-disabling mutation in the fish. Interestingly, fish in which telomerase activity was disabled displayed a variety of traits that are similar to those seen in humans with a disorder called dyskeratosis congenita, which is also due to a mutation in telomerase.

“Very quickly we began to see an effect on rapidly dividing tissues such as the blood, gut and sperm,” said Harel. “The fish rapidly become sterile, their intestines began to atrophy and they made fewer types of blood cells than their peers.”

The researchers conclude that the killifish is currently the fastest way to study diseases of telomere shortening in vertebrates. They are hopeful that the other mutant strains will be equally useful in their lab and in other labs worldwide.

“Itamar has generated a range of tools necessary to study how genetic changes affect physical characteristics of the killifish,” said Brunet. “It’s a true ‘genotype to phenotype’ platform, and is likely to be transformative. Now we have what is essentially a high-throughput vertebrate model for aging research.”

Other Stanford authors are postdoctoral scholars Param Priya Singh, PhD, and Chi-Kuo Hu, PhD; former postdoctoral scholar Dario Riccardo Valenzano, PhD; graduate student Matthew Pech; undergraduate Elisa Zhang; research assistant Ben Machado; technician Sabrina Sharp; and professor of medicine and of biochemistry Steven Artandi, MD, PhD.

The work was supported by the National Institutes of Health (grant DP1AG044848); the Glenn Laboratories for the Biology of Aging; Damon Runyon Cancer Research Foundation, Rothschild and the Human Frontier Science Program fellowships; and a Stanford Dean’s Fellowship.

Information about Stanford’s Department of Genetics, which also supported the work, is available at http://genetics.stanford.edu.

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