Mapping with Mice

© Luke Bartholomew Tan

It was 1980, in the early days of the molecular biology era, when Nancy Jenkins and her collaborator-and-spouse Neal Copeland accepted their first faculty positions at the Jackson Laboratory in Bar Harbor, Maine. "Everyone told us that going there would ruin our careers," says Jenkins. The lab was populated by geneticists who were used to thinking in terms of mice, not molecules. "We were the first people at Jax who even knew what a restriction enzyme was, let alone used one," she says. So friends feared their science might suffer.

But Jenkins saw things differently. "We thought that if you could combine molecular biology with formal genetics, you could begin to build incredibly interesting models of human disease." Turns out, Jenkins had it right.

"That was their brilliant scientific prescience," says Jeffrey Friedman of Rockefeller University. "They brought their skills to the Jackson Labs, where they were in a position to apply molecular tools to the fabulous genetic resources there. Neal and Nancy really were the founders of modern mouse genetics."

"They have made so many contributions to mouse genetics, it's difficult to really catalog them," agrees Raju Kucherlapati of Harvard Medical School. Among those contributions, Jenkins and Copeland developed new approaches for speeding the identification and cloning of genes through retroviral and transposon integration, and they produced genetic maps of the mouse genome that served as tools for the entire community - all while pursuing their own interests in the genes and mutations involved in development and cancer.

And they did it all together. "When we started at Jax, we had to make a decision," says Jenkins. "Our interests were exactly the same. So we were either going to compete. Forever. Or we were going to collaborate. We decided if we wanted to stay married, we'd better collaborate." So the two have co-authored every one of the 700-plus papers they've published since then. "It's hard to say where Neal ends and Nancy begins," laughs George Carlson of the McLaughlin Research Institute in Montana.

Their partnership began when Nancy and Neal were postdocs at the Dana Farber Cancer Institute in the late 1970s. There they started studying retroviruses and the effect these parasites have on a host organism. They carried this interest in endogenous viruses with them to the Jackson Lab. "The inbred strains of mice they have there are quite a rich repository of endogenous viruses," says Jenkins. "Some of those strains have a high incidence of cancer. Some don't. So the question was: What was each strain's endogenous virus content, and did those viruses contribute to the cancer?"

To answer that question, Jenkins says, "Neal called up the production lab, which is where they maintain all the inbred strains, and said 'I want one of everything.' No one had ever done this before. It was just unheard of." But eventually they got their delivery. "In came these big boxes, and inside were hundreds of little ice cream cartons that each had three air holes poked in top." And in each carton was a single mouse. "It was like doing an Easter egg hunt," says Jenkins. "'Oh, look at this one. It has spots!' We were kids in a candy shop."

The two set out to tally the retroviruses. In the process, they noticed something. "One of the endogenous viruses, for which we had a probe, was tracking along with the dilute mutation in inbred strains that inherited a dilute color coat when we did our crosses," she says. And Jenkins had a hunch that the virus was inserting itself into the dilute gene, giving these mutants their washed-out appearance. After discussing her suspicions at a meeting within Jax, Jenkins says that Eva Eicher, who studies the genetics of sex determination, finally said: "'So just prove it. Prove that the mutation is caused by a virus. I've got the mouse in my mouse room you should use'." For 25 years, Eicher had been keeping a mouse strain that had once been dilute, but had reverted to the wild-type coloring somewhere along the line. "I looked at that mouse and saw the virus was gone," says Jenkins. "So, several centuries ago, the dilute mutant arose because a virus popped into that gene" - a finding they published in Nature in 1981.

The dilute gene, it turns out, encodes a type of myosin that, among other things, corrals pigment granules in the periphery of the melanocyte in which they're synthesized, so they can be incorporated into hair. Without that myosin, "the pigment granules sit in a blob around the melanocyte nucleus" and the animal's coat is subdued in hue, says John Mercer of the McLaughlin Research Institute, who describes himself as "the last postdoc standing when we finished cloning the dilute locus" in the Jenkins/Copeland lab, results that appeared in Nature in 1991.

Jenkins' work on coat color mutations - including dilute and agouti (yellow) - "is really classic stuff that should be in all the genetics textbooks," says David Largaespada of the University of Minnesota, another former postdoc and collaborator. "It's a tour de force of forward genetics and, in the case of dilute, really opened up the whole cell biology of vesicle transport in mammalian cells."

And that's just one example of the discoveries that emerged from Jenkins' and Copeland's work on positional cloning - findings that were accelerated by the couple's efforts to build a molecular map of the mouse genome. "They were major players in the early mapping of genes," says Luis Parada of the University of Texas Southwestern Medical Institute. "I think a significant percentage - if not the major percentage - of genes mapped in the late '80s and early '90s were done through a collaboration with Nancy and Neal."

At the start, the mapping project was simply meant to be a lab resource. "We thought our postdocs would find it useful. But it quickly took on a life of its own," says Jenkins - and consumed much of the lab's attention. "I would venture to guess that a third of our publications are based on gene mapping," says Jenkins, who by then had moved, with Copeland, to the National Cancer Institute in Frederick, Md. "That work could never have been tackled at an academic institution," she adds. "It took time, it took money, it was not focused, and it would never have stood up to an R01. But a lot of exciting science came out of it, so it was worth it."

For example, mapping led them to the lymphoproliferation gene, lpr, which turns out to encode Fas ligand, making it "one of the most important genes in the cell death pathway," says Jenkins. The group also cloned several genes that affect coat color, such as dilute and microphthalmia. And because the pigment system derives from the embryo's neural crest, many of these mutations have interesting effects on other neural tissues. Dilute mutants, for example, are prone to seizures. And mice with mutations in microphthalmia have small eyes (hence the name).

While half the Jenkins-Copeland lab has traditionally focused on mutations that affect development, the other half has mined the mouse genome for genes involved in cancer. In the early days, they relied on insertional mutagenesis using retroviruses, a lab specialty. "This was the first method that could give you, essentially, a saturation analysis of oncogenes in the mouse," says Irving Weissman of Stanford University. "So in terms of people who do gene discovery of cancer genes, that made Nancy and Neal among the best in the world."

For reasons that are not well understood, however, retroviral insertion usually leads to leukemias, lymphomas, and other hematopoietic cancers. "Our lab, along with two or three other big labs in the Netherlands, has probably contributed to 95% of the hematopoietic cancer genes that have been identified in mouse," says Jenkins. But blood cancers account for only a few percent of the cancers that plague humans. "So we really wanted to study solid tumors."

That's where Sleeping Beauty comes in. Largaespada started working with this transposon, which originated in fish, after he left Jenkins' group to start his own lab in Minnesota. "Nancy and Neal had gotten me excited about using insertional mutagenesis as a tool for gene discovery," he says. And he was hoping that transposon insertion might yield a broader spectrum of cancers than retroviruses had. Largaespada's student, Adam Dupuy, then took Sleeping Beauty with him when he left to do his postdoc - with Jenkins and Copeland. "Adam came to the lab specifically to do that science," says Jenkins. "Of course, Neal and I, in our infinite wisdom, said it will never work," she jokes. The frequency with which the transposon moved was just too low. "And we're not talking two-fold too low. We're talking thousands of fold," she says. "So we sat down and pow-wowed and made a list of everything we could think of to make this thing move. And darned if we weren't successful. I love it when I'm wrong and the postdocs are right!"

And once the transposon got moving, it produced mice that developed, predominantly, hematopoietic cancers. "How funny is that? We spent 25 years looking at hematopoietic cancers, thought we'd found a way to look at solid tumors, and what do we get back? Hematopoietic cancers," says Jenkins. "When I looked at the first pathology report, I thought, 'you have got to be kidding!'" But the results - published in Nature in 2005 - indicated that they were heading in the right direction. Since then, Jenkins and Copeland have continued to tweak the Sleeping Beauty system, and they are now able to express the transposase - needed to get the transposon to move - in a tissue-specific manner. Their preliminary results look promising. "We think that the system is going to be generally useful for studying the genetics of any type of cancer that can be studied in mice," Largaespada says.

What's more, it provides the researchers with models that will allow them to identify new targets for cancer screening and even treatment - work that Jenkins and Copeland are continuing in their new digs at Singapore's Institute of Molecular and Cell Biology. With science funding dwindling in the United States, Jenkins says, "we realized we couldn't afford to keep our work at NCI going at the level it needed to be." So when Singapore made an offer, Jenkins says they were happy to make the move.

The science is great, our colleagues are great, we love the area. And the food is excellent, so I'm happy as a clam." Indeed, her love of good food is legendary. "Nancy and Neal are extreme hedonists," says Weissman, who tried to recruit the couple to Stanford before they left for Singapore. "After I'm with them for a few days, I have to go on a diet."

"No two people I know are as interested in going to new places and finding the next great restaurant," adds Friedman. "They're just incredibly adventurous." That spirit should continue to serve them well as they embark on their next great scientific adventure. "The fact that Singapore was able to attract Nancy and Neal was clearly a coup," says Kucherlapati. "And with the resources that they now have available there, it should be win-win for everybody."