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One of the great transformations for descendants of fish was to become creatures that walk on land, with thick, sturdy "toes" replacing their long, elegant fins. Scientists from the University of Chicago now know how the same cells which make fin rays in fish, form fingers and toes in animals.
After three years of painstaking experiments using gene-editing techniques and sensitive fate maps to label and track developing cells in fish, researchers can now describe how the small flexible bones at the ends of fins are related to fingers and toes suitable for life on land.
"When I first saw these results you could have knocked me over with a feather," said the study's senior author, Neil Shubin, PhD, the Robert R. Bensley Distinguished Service Professor of Organismal Biology and Anatomy at the University of Chicago. Shubin is an authority on the transition from fins to limbs.
To unravel how fins might have transformed into wrists and fingers, researchers worked mostly with a standard fish model: zebrafish.
Tetsuya Nakamura, PhD, a postdoctoral scholar in Shubin's lab, used a gene-editing technique, CRISPR/Cas to cut and insert genes into the zebrafish. Clustered regularly interspaced short palindromic repeats (CRISPR, pronounced crisper) are pieces of single-celled organisms. These single celled organisms have all their water-soluble components (proteins, DNA and metabolites) together within the cytoplasm enclosed by one cell membrane, rather than in separate compartments. Short segments of "spacer DNA" can be cut by CAS proteins to isolate genes or gene elements.
At the same time, Andrew Gehrke PhD, a former graduate student in Neil Shubin's lab, refined cell-labelling techniques to map when and where specific embryonic cells migrated as the fish and mice grew and developed.
"It was one of those eureka moments," Gehrke said. "We found that the cells that mark the wrists and fingers of mice and people were exclusively in the fin rays of fish."
Scientists studied the development of cells, beginning in some experiments soon after fertilization, and followed them as they became part of an adult fin.
"What matters is not what happens when you knock out a single gene but when you do it in combination," Nakamura explained. "That's where the magic happens."
Researchers also used a high-energy CT scanner toview minute fin structures within the adult zebrafish. These can be invisible, even to most traditional microscopes.
The authors suspect what happened in Nakamura's mutants was that cells stopped migrating from the base of the fin to the tip. This inability to migrate to the tip meant there were fewer cells to make long fin rays, leaving more cells in the base to become flexible cartilage elements.
Future research includes new expeditions to find more fossil fish — such as Tiktaalik, discovered by Shubin and colleagues in 2006 — to link primitive fish development into the first four-legged animals. They are now planning experiments with Hox genes to learn the extent to which common cells form different structures in fish and people.
The research was funded by the Japan Society for the Promotion of Science Postdoctoral Research Fellowship, the Uehara Memorial Foundation Research Fellowship, the Marine Biological Laboratory, the National Institutes of Health, the National Science Foundation, the Brinson Foundation and the University of Chicago. Additional authors include Justin Lemberg and Julie Szymaszek.
Markers of the wrists and digits in the limb of a mouse (LEFT) are present in fish
and demarcate the fin rays (RIGHT). The wrist and digits of tetrapods
are the cellular and genetic equivalents of the fin rays of fish.
Image Credit: Marie Kmita and Andrew Gehrke