Roughly 500 million years ago, something that would forever change the course of eukaryotic development was brewing in the genome of some lucky organism: a gene called Pax6. The gene is thought to have orchestrated the formation of a primitive visual system, and in organisms today, it initiates a genetic cascade that recruits more than 2,000 genes to build different parts of the eye.
Pax6 is only one of thousands of genes encoding transcription factors that each have the powerful ability to amplify and silence thousands of other genes. While geneticists have made leaps in understanding how genes with relatively simple, direct functions could have evolved, explanations for transcription factors have largely eluded scientists. The problem is that the success of a transcription factor depends on how usefully it targets huge numbers of sites throughout the genome simultaneously; it’s hard to picture how natural selection enables that to happen. The answer may hold the key to understanding how complex evolutionary novelties such as eyes arise, said Cédric Feschotte, a molecular biologist at Cornell University.
For more than a decade, Feschotte has pointed to transposons as the ultimate innovators in eukaryotic genomes. Transposons are genetic elements that can copy themselves and insert those copies throughout the genome using a splicing enzyme they make. Feschotte may have finally found the smoking gun he has been looking for: As he and his colleagues recently reported in Science, these jumping genes have fused with other genes nearly 100 times in tetrapods over the past 300 million years, and many of the resulting genetic mashups are likely to encode transcription factors.
The study provides a plausible explanation for how so-called master regulators like Pax6 could have been born, said Rachel Cosby, the first author of the new study, who was a doctoral student in Feschotte’s lab and is now a postdoc at the National Institutes of Health. Although scientists had theorized that Pax6 arose from a transposon hundreds of millions of years ago, mutations since that time have obscured clues about how it formed. “We could see that it was probably derived from a transposon, but it happened so long ago that we missed the window to see how it evolved,” she said.
David Adelson, chair of bioinformatics and computational genetics at the University of Adelaide in Australia, who was not involved with the study, said, “This study provides a good mechanistic understanding of how these new genes can form, and it squarely implicates the transposon activity itself as the cause.”
Scientists have long known that transposons can fuse with established genes because they have seen the unique genetic signatures of transposons in a handful of them, but the precise mechanism behind these unlikely fusion events has largely been unknown. By analyzing genes with transposon signatures from nearly 600 tetrapods, the researchers found 106 distinct genes that may have fused with a transposon. The human genome carries 44 genes likely to have been born this way.
The structure of genes in eukaryotes is complicated, because their blueprints for making proteins are broken up by introns. These noncoding sequences are transcribed, but they get snipped out of the messenger RNA transcripts before translation into protein occurs. But according to Feschotte’s new study, a transposon can occasionally hop into an intron and change what gets translated. In some of these cases, the protein made by the fusion gene is a mashup of the original product and the transposon’s splicing enzyme (transposase).
Once the fusion protein is created, “it has a ready-made set of potential binding sites scattered all over the genome,” Adelson said, because its transposase part is still drawn to transposons. The more potential binding sites for the fusion protein, the higher the likelihood that it changes gene expression in the cell, potentially giving rise to new functions.
“These aren’t just new genes, but entire new architectures for proteins,” Feschotte said.
Cosby described the 106 fusion genes described in the study as the “tiniest tip of the iceberg.” Adelson agreed and explained why: Events that randomly create fusion genes for functional, non-harmful proteins rely on a series of coincidences and must be exceedingly rare; for the fusion genes to spread throughout a population and withstand the test of time, nature must also positively select for them in some way. For the researchers to have found the examples described in the study so readily, transposons must surely cause fusion events much more often, he said.
“All of these steps are very unlikely to happen, but this is how evolution works,” Feschotte said. “It’s very quirky, opportunistic and very unlikely in the end, yet you see it happen over and over again on the timescales of hundreds of millions of years.”
To test whether the fusion genes acted as transcription factors, Cosby and her colleagues homed in on one that evolved in bats 25 million to 45 million years ago — a blink of an eye in evolutionary time. When they used CRISPR to delete it from the bat genome, the changes were striking: The removal dysregulated hundreds of genes. As soon as they restored it, normal gene activity resumed.
To Adelson, this shows that Cosby and her co-authors practically “caught one of these fusion events in the act.” He added, “It’s especially surprising because you wouldn’t expect a new transcription factor to cause wholesale rewiring of transcriptional networks if it had been acquired relatively recently.”
Although the researchers didn’t determine the function of the other fusion proteins definitively, the genetic hallmarks of transcription factors are there: Around a third of the fusion proteins contain a part called KRAB that is associated with repressing DNA transcription in animals. Why transposases tended to fuse with KRAB-encoding genes is a mystery, Feschotte said.
Transposons comprise a hefty chunk of eukaryotic DNA, yet organisms take extreme measures to carefully regulate their activity and prevent the havoc caused by problems such as genomic instability and harmful mutations. These dangers made Adelson wonder if fusion genes sometimes endanger orderly gene regulation. “Not only are you perturbing one thing, but you’re perturbing this whole cascade of things,” he said. “How is it that you can change expression of all these things and not have a three-headed bat?” Cosby, however, thinks it’s unlikely that a fusion gene leading to harmful morphogenic changes would readily propagate through a population.
Damon Lisch, a plant geneticist at Purdue University who studies transposable elements and was not involved with the study, said he hopes this study pushes back against a widespread but misguided notion that transposons are “junk DNA.” Transposable elements generate tremendous amounts of diversity and have been implicated in the evolution of the placenta and the adaptive immune system, he explained. “These are not junk — they’re living little creatures in your genome that are under very active selection over long periods of time, and what that means is that they evolve new functions to stay in your genome,” he said.
Though this study highlights the mechanism underlying transposase fusion genes, the vast majority of new genetic material is thought to form through genetic duplication, in which genes are accidentally copied and the extras diverge through mutation. But a large quantity of genetic material does not mean that new protein functions will be significant, said Cosby, who is continuing to investigate the function of the fusion proteins.
“Evolution is the ultimate tinkerer and ultimate opportunist,” said David Schatz, a molecular geneticist at Yale University who was not involved with the study. “If you give evolution a tool, it may not use it right away, but sooner or later it will take advantage of it.”