Bowfin genome reveals old dogfish can teach researchers new tricks
International team of researchers sequence genome of the enigmatic bowfin fish
Peer-Reviewed PublicationIMAGE: FRESHLY DEPOSITED BOWFIN EGGS ATTACHED TO NEST MATERIAL. MALE BOWFIN BUILD NESTS IN WHICH FEMALES LAY EGGS. AFTER THE MALE FERTILIZES THE EGGS, IT WILL REMAIN WITH THE NEST TO GUARD THE YOUNG view more
CREDIT: M. BRENT HAWKINS
The fish species Amia calva goes by many names including bowfin, freshwater dogfish, grinnel, and mud pike. No matter what you call it, this species is an evolutionary enigma because it embodies a unique combination of ancestral and advanced fish features.
In a paper published August 30 in Nature Genetics an international and collaborative team of researchers, headed by Ingo Braasch and Andrew Thompson of Michigan State University, have begun to unravel the enigma by sequencing the genome of the bowfin fish. Their collaborative analysis yielded unexpected insights into diverse aspects of the biology of this mysterious, ancient lineage.
The bowfin is a bony fish endemic to eastern North America and is the sole surviving member of a once large lineage of many species that are now known only from fossils. Scientists have long been fascinated with the bowfin because it bears a combination of ancestral features, such as lung-like air breathing and a robust fin skeleton, and derived features like simplified scales and a reduced tail. The bowfin also occupies a key position in the fish family tree, where it sits between the teleosts, a large and diverse group that arose recently, and more ancient branches that include sturgeons, paddlefish, and bichirs.
Due to this special position in the fish family tree, the bowfin can help scientists understand how aspects of modern fishes evolved from their ancient antecedents. By examining the bowfin genome, scientists can investigate the genetic basis of the unique set of old and new features of the bowfin. They can also use this genomic information as a framework to better understand the origin of the teleosts, which have duplicated and extensively modified their genomes since separating from the bowfin lineage and emerging as the dominant lineage in most aquatic habitats.
As a doctoral candidate in the Department of Organismic and Evolutionary Biology at Harvard University, study co-author M. Brent Hawkins (PhD ’20) examined the evolution and development of the bowfin pectoral fin. Hawkins’ doctoral thesis, conducted with Professor Matthew P. Harris, Harvard Medical School and Boston Children’s Hospital, and Professor James Hanken, Department of Organismic and Evolutionary Biology at Harvard university, contributed some of the study’s most surprising findings.
Hawkins focused on the pectoral fin of the bowfin because of its ancestral configuration of the skeleton. The bowfin retains the metapterygium, which is a portion of the fin skeleton that is homologous to the limb bones of tetrapods. Model organisms such as the widely used zebrafish and medaka have lost the metapterygium, which makes comparisons between the fin and the limb difficult. By studying the bowfin fin, scientists can use knowledge of bowfin development as a steppingstone to bridge teleost fin development to tetrapod limb development and help explain the evolution of the fin-to-limb transition.
CAPTION
Schematics show the arrangement of bones in fins and limbs. Elements that are derived from the ancestral metapterygium are shown in magenta. The tetrapod limb and a portion of the bowfin fin arose from the metapterygium, while teleosts have lost the metapterygial components
CREDIT
M. Brent Hawkins
With co-authors Emily Funk and Amy McCune, both at Cornell University, Hawkins collected young bowfin embryos from nests in the wild in upstate New York. Hawkins raised the embryos, collecting pectoral fin samples as they developed. He extracted mRNA from the samples and performed Transcriptome Sequencing with the help of the Harvard University Bauer Core to determine which genes are turned on in the developing fin by parsing the transcriptome data using the genomic reference sequence. Once identified, he used in situ hybridization to visualize where these genes are activated during fin outgrowth. Initially, Hawkins expected the bowfin gene data to look very similar to other fins and limbs. “As a field, we have characterized many of the genes involved in appendage patterning. We have a good idea of what the essential fin and limb genes are and where they should be turned on,” said Hawkins. However, when he analyzed the fin data he was shocked by the results.
While the bowfin pectoral fins did express many of the expected appendage growth genes, some of the most critical of these genes were in fact entirely absent. One such gene called fibroblast growth factor 8 (Fgf8) is turned on at the far tip of developing fins and limbs and is required for the outgrowth of these appendages. When Fgf8 is lost appendage outgrowth is impaired, and if extra Fgf8 is applied to an embryo, it can cause a new limb to form. “Every other fin and limb we know of expresses Fgf8 during development,” Hawkins said. “Discovering that bowfin fins don’t express Fgf8 is like finding a car that runs without a gas pedal. That the bowfin has accomplished this rewiring indicates unexpected flexibility in the fin development program. With the genome in hand, we can now unlock how this flexibility evolved.”
While some genes like Fgf8 were mysteriously absent from the bowfin fin, other genes were unexpectedly activated in the fins. The HoxD14 gene is expressed in the fins of fishes from the deeper branches of the fish family tree, such as paddlefish, but this gene was lost in more recent branches including the teleosts. When the authors found this gene in the bowfin genome data, they thought it must not be expressed because the DNA sequence did not encode a functional protein. Surprisingly, Hawkins and colleagues found that bowfin fins made HoxD14 gene transcripts at high levels, even though it did not code for a protein. “The fact that the HoxD14 gene can no longer make a protein, but it still transcribed into mRNA at such high levels suggests that there might be another function that we do not yet understand. We might be seeing a new level of Hox gene regulation at play in the bowfin,” said Hawkins.
CAPTION
A recently hatched bowfin larva facing to the left as seen through a microscope.
CREDIT
M. Brent Hawkins
Taken together the Fgf8 and HoxD14 results indicate that genetic programs, even those that guide the formation of important structures such as fins and limbs, are not as invariable as previously thought. “By studying more species, we learn which rules are hard and fast and which ones evolution can tinker with. Our study shows the importance of sampling a broader swath of natural diversity. We might just find important exceptions to established rules,” said Hawkins.
Hawkins also suggests that the results of the bowfin study serve as a warning against treating members of deeper branches of the tree of life as stand-ins for actual ancestors. “Some people might describe species like the bowfin as a ‘living fossil’ that reliably represents the ancestral condition of a lineage. In reality, these deeper branches have been evolving past that ancestor for just as long as the more recent branches, doing their own thing and changing in their own ways. In evolution, old dogs do learn new tricks.”
Hawkins is currently a postdoctoral researcher in the lab of Matthew P. Harris at Harvard Medical School and Boston Children’s Hospital.
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JOURNAL
Nature Genetics
ARTICLE TITLE
The bowfin genome illuminates the developmental evolution of ray-finned fishes
ARTICLE PUBLICATION DATE
30-Aug-2021
Learning from a ‘living fossil’
A Spartan-led team has probed the genetics of bowfin fish like never before to reveal insights about human health and evolutionary history
Peer-Reviewed PublicationAs we live and breathe, ancient-looking fish known as bowfin are guarding genetic secrets that that can help unravel humanity’s evolutionary history and better understand its health.
Michigan State researchers Ingo Braasch and Andrew Thompson are now decoding some of those secrets. Leading a project that included more than two dozen researchers spanning three continents, the Spartans have assembled the most complete picture of the bowfin genome to date.
“For the first time, we have what’s called a chromosome-level genome assembly for the bowfin,” said Braasch, an assistant professor of integrative biology in the College of Natural Science. “If you think of the genome like a book, what we had in the past was like having all the pages ripped out in pieces. Now, we’ve put them back in the book.”
“And in order,” added Thompson, a postdoctoral researcher in Braasch’s lab and the first author of the new research report, published Aug. 30 in the journal Nature Genetics.
This is really important information for a few reasons, the duo said, and it starts with the bowfin being what Charles Darwin referred to as a “living fossil.” The bowfin, or dogfish, looks like an ancient fish.
This doesn’t mean that the bowfin hasn’t evolved since ancient times, but it has evolved more slowly than most fishes. This means that the bowfin has more in common with the last ancestor shared by fish and humans, hundreds of millions of years ago, than, say, today’s zebrafish.
Zebrafish — which are modern, so-called teleost fishes — are a notable example because they’re widely used by scientists as a model to test and develop theories about human health. Having more genetic information about the bowfin helps make the zebrafish a better model.
“A lot of research on human health and disease is done on model organisms, like mice and zebrafish,” Thompson said. “But once you identify important genes and the elements that regulate those genes in zebrafish, it can be hard to find their equivalents in humans. It’s easier to go from zebrafish to bowfin to human.”
For example, one particularly interesting gene is one that’s used in developing the bowfin’s gas bladder, an organ the fish uses to breathe and store air. Scientists believe that the last common ancestor shared by fish and humans had air-filled organs like these that were evolutionary predecessors to human lungs.
In their new study, the Spartan researchers could see that a certain genetic process in the bowfin’s gas bladder development bore striking similarities to what’s known about human lung development. A similar process is also present in the modern teleost fishes, but it’s been obscured by eons of evolution.
“When you looked for the human genetic elements of this organ development in zebrafish, you couldn’t find it because teleost fishes have higher rates of evolution,” Thompson said. “It’s there in modern fishes, but it’s hidden from view until you see it in bowfin and gar.”
The gar is another air-breathing fish with “living fossil” status that’s studied by Braasch and his team. With both the gar and bowfin genomes, the team was able to show where these genetic elements linked to gas bladder and lung formation were hiding out in the modern teleost fishes. The ancient fish enable researchers to build a better bridge between the established modern fish model organisms and human biology.
“You don’t want to base that bridge on one species,” said Braasch, who added this finding also strengthens the implications for evolutionary history. “This is another piece of the puzzle that suggests the common ancestor of fish and humans had an air-filled organ and used it for breathing at the water surface, quite similar to what you see in bowfin and gar.”
Although these findings have insights that are pertinent to all of humanity, Spartans might feel a special affinity for the bowfin. For starters, male fish turn their fins and throats a bright shade of green during spawning season. Also, famed biologist William Ballard of Dartmouth College studied bowfin development from eggs to larval fish at Michigan State’s W.K. Kellogg Biological Station during the 1980s. This was what he called his “Odyssey of Strange Fish,” and Braasch’s team now uses his work to guide their genomic analyses of bowfin development.
Bowfins are native to Michigan. They could be in the Red Cedar River on MSU’s campus now, according to Thompson, but they also can be quite elusive and, sometimes, very aggressive. This made collaborations essential for securing specimens. With colleagues at Nicholls State University in Louisiana, the team caught bowfins for genome sequencing. Amy McCune, a collaborator and professor at Cornell University, knew where to find bowfin eggs in upstate New York and had a graduate student gifted at securing these unique samples for investigating bowfin development.
The Spartans also had connections at other universities and institutions with experts in bowfin biology, chromosome evolution and more. All told, the team included researchers from six states as well as France, Japan and Switzerland. Back in East Lansing, graduate students Mauricio Losilla and Olivia Fitch, research technologist Brett Racicot, and Kevin Childs, director of the MSU Genomics Core facility, also contributed to the study, which comes with an interesting twist at the end.
Almost all vertebrate creatures that grow paired limbs or fins share a common gene.
“Humans use it, mice use it. All fishes that have been studied so far use it,” Braasch said. “The naïve expectation would be that bowfin do, too.”
But that’s not what the team found. The bowfin, the “living fossil,” has evolved a different way of growing its paired fins.
“For whatever reason, it changed its genetic programming. Even ‘living fossils’ keep evolving. They’re not frozen in time,” Braasch said. “It’s sort of a cautionary tale that we shouldn’t take these things for granted. You have to look trait by trait, gene by gene and across many different species to paint the complete picture.”
JOURNAL
Nature Genetics
ARTICLE TITLE
The bowfin genome illuminates the developmental evolution of ray-finned fishes
ARTICLE PUBLICATION DATE
30-Aug-2021
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