Life On Land Tied To Gene Expansion

"So you get expansion of gene expression--not expression of a new gene--just expansion to a new area," says Dymecki, HMS assistant professor of genetics. She and colleagues Scott Baur and Jia J. Mai have recently identified the structure of the gene and also the genetic switch that may have brought about this expansion.

Until recently, the gene, which codes for a receptor found in the brains and skeletons of all vertebrates living today, was thought to be controlled by a single switch, or promoter. If that were true, a defect in the promoter should affect expression in the brain as well as the skeleton. But the researchers found that while mutant mice carrying such a defect almost totally lacked fingers and toes, their brains appeared, for the most part, normal. On closer inspection, the researchers found that there was not one but two promoters, one controlling gene expression in the brain, the other, which carried the mutation, in the limbs. The defective promoter was farthest away from the gene. "This is the first time anyone has seen this distal promoter," says Dymecki.

She and Baur, who is a graduate student, believe that this more distant promoter evolved more recently, perhaps as a result of a duplication of the one lying closer to the gene. Once formed, the new promoter may have accumulated mutations that enabled it to interact with transcription factors found in developing limb cells. As a consequence, the receptor previously expressed in the brain would have come to be expressed in the limb buds. Baur is currently comparing the two promoters to see if he can find signs of a duplication.

Meanwhile, the findings suggest that the origin of life on land may have entailed not just the invention of new genes but also putting old ones to new uses. It is a process nature has used many times before, Dymecki says. The Hox genes, which were first found to regulate body shape in flies and are now known to regulate body and limb development in vertebrates, produce their wide variety of effects by being expressed in different amounts at different times and places. It is not clear how the newly discovered promoter may have brought about the wide variety of vertebrate digits‹from the frog¹s grasping toes to the stubby toes of a human. Dymecki and her colleagues have been developing a system for identifying how variations between species have evolved.

"These are things we¹re still fleshing out," Dymecki says. "I would have to say it¹s been a real whirlwind just to get this paper out." In fact, their current paper appears back-to-back in Development with a paper by a group at UCLA. The California researchers knocked-out the receptor gene in the brain and digits of mice. Intriguingly, the knockout mice exhibited the same phenotype as the HMS mutants. They lacked digits but their brains were apparently normal. One explanation for why effects in the brain were masked is that the receptor may play such an important role there that nature has provided a genetic backup to make sure that its job gets done.

If it weren¹t for a twist of nature, Dymecki and he colleagues would never have identified the second promoter. Dymecki¹s whole excursion into skeletal development "was definitely serendipity," she says. In the course of studying brain development--her primary interest--Dymecki had generated a series of transgenic mice, each with a piece of DNA wedged into a different part of the genome (see sidebar). When she and her colleagues tried ear tagging one, they discovered that the mouse was unable to grab the table top.

It turned out, the mice had failed to develop digits. Suspecting that the piece of DNA had wedged itself into the middle of a gene for skeletal development, the researchers homed in on the gene, which produced IB bone morphogenetic protein receptor (BMPRIB). The protein was known to play a role in skeletal development, specifically the laying down of the cartilaginous blueprint that eventually develops into the bony skeleton. But no one had actually mapped out the structure of the gene (BmprIB)--that is, how exactly it was broken up into functional units, or exons.

After identifying the structure of BmprIB, Baur was able to determine that the chunk of DNA had integrated and essentially knocked out what appeared to be the promoter. The lack of any apparent defect in the brain of their mutant--at the time they did not know that even knockouts show no brain defects--led them to look for a second regulatory element.

Dymecki and her colleagues plan to use the transgenic system to see how, exactly, the limb-region promoter turns on BmprIB during mouse development‹in what cells and at what times. Comparing BmprIB regulation in mice to other animals could provide a first step to understanding how the extraordinary array of land-dwelling adaptations have evolved in different species. "We want to understand what regulatory elements are involved and what the evolutionary implications of those elements are in terms of species-to-species variation in digit formation," says Dymecki. "It does make me chuckle that it all came out of a simple transgenic insertion."