The Human Genome Project succeeded at the monumental task of assembling a complete parts list for the human body by decoding the DNA sequence of each and every human gene. But an even harder job remains: determining the function of each gene in health and disease.
A new tool for genome research, developed by Tian Xu, Ph.D., professor and vice chair of genetics and Howard Hughes Medical Institute investigator, with colleagues at Fudan University in Shanghai, China, promises to greatly accelerate the work of assigning purpose to thousands of unexplored human genes. The tool is a jumping gene, a small piece of DNA called a transposon that moves around the genome with a preference for settling in other genes and suppressing their activity, which allows scientists to discern their function.
Transposons are active players in many plant and insect genomes, and they helped to make the fruit fly Drosophila the darling of geneticists, as these mobile DNA fragments were used to decipher the role of nearly every gene in that model organism. But for decades scientists could not find an equivalent transposon for mammals.
As reported in the August 12 issue of the journal Cell, Xu and researchers in Shanghai, at Duke University and at the University of Colorado tweaked a transposon called piggyBac, originally identified in the cabbage looper moth by Malcolm J. Fraser, Ph.D., of the University of Notre Dame, so that it can be easily cut and pasted into the genomes of higher organisms, including mice and humans. “With this transposon, we now have the ability to systematically inactivate each and every gene in a model organism like the mouse,” Xu says. David Largaespada, Ph.D., an expert on human genetics at the University of Minnesota who developed another transposon that he and his colleagues recently used in mice to identify genes involved in cancer, agrees: “Researchers now have what is essentially a furry fruit fly.”
Scientists have traditionally relied on mutagenesis, using chemicals to modify mouse genes, but this is painstakingly slow, and it is often difficult to locate the genes that have been mutated. The piggyBac transposon, when injected into fertilized mouse eggs along with an enzyme known as transposase, is remarkably efficient at inserting itself into important coding regions of the genome, and, as its name implies, it can carry genetic tags that allow researchers to locate mutations quickly.
PiggyBac has the added feature of total reversibility, which allows scientists to verify that particular mutations have particular effects. In the presence of transposase, piggyBac easily hops into genes, and it remains stably in place in any mice of subsequent generations that do not inherit the gene for the enzyme. But when these mice are mated with others that produce transposase, piggyBac hops back out of genes, leaving no trace of the mutation in their offspring.
This combination of traits makes piggyBac a “dream tool” for geneticists, Xu says: “This new technology will completely change the game of using mutagenesis to understand the function of mouse genes, and by extension their human counterparts.”
PiggyBac could also be a promising new vehicle for human gene therapy, according to Xu, who says that, in addition to carrying tags that mark mutated genes, piggyBac can be engineered to carry whole blocks of DNA containing one or more new genes into the genome.
To demonstrate this genetic piggybacking, Xu and his colleagues used piggyBac to insert a gene for a protein that glows red under ultraviolet light. As seen in the photo at left, a mother mouse with the gene and any offspring that carry it cast an obvious red hue under the light, but pups without the gene do not. However, many more experiments will be required to know whether the transposon, or some variation of it, could reliably and safely transfer therapeutic genes to humans.
Xu’s immediate goal is to use piggyBac to systematically inactivate every gene in the mouse, one by one, a project that would be unthinkable with traditional mutagenesis methods. “For the past two decades, it has routinely taken about a year to mutate one gene in a mouse, and altogether about 3,000 genes have been knocked out in mice, out of a total of about 25,000 that are in the genome,” Xu explains. With the help of piggyBac, he says, “in three months, with two students, we have done 70 genes. We plan to produce mutant mice inactivating most of the genes in three years.”