Out of the Frying Pan

© Jon Enoch

As a graduate student at the University of Birmingham in the United Kingdom, Amanda Fisher says she was incredibly naïve. "I thought science was just great fun. And it is. But I was very unworldly." So her thesis advisor, Geoff Brown, suggested that she do a postdoc in a lab that might expand her horizons: Robert Gallo's at the National Institutes of Health. It was 1983, at the height of Gallo's race with the French to isolate the virus that causes AIDS. "Now that was bang-on worldly!" she says.

Fisher got the experience she needed—and then some. "It was a tough lab," says Fisher, who was "shocked," for example, by Gallo's encouragement of competition within the lab. "It was very different from what I was used to. But it was a fantastic experience. I left there a very much more confident person, because I knew I could hold my own with the Americans. I knew I could cut it in a very tough lab—because I survived."

Judging by her publications, she did more than survive; in three years, Fisher racked up three papers in Science and another three in Nature. She then returned to England, where she's uncovered key molecular mechanisms in cell differentiation and cell fate, finding a zone within the nucleus where silenced genes are sequestered, and determining which genes are crucial for nuclear reprogramming.

"I think she's among the leaders of British biological sciences," says Rick Young of Whitehead Institute at Massachusetts Institute of Technology. "It's because of people like Mandy that the British, despite having a much smaller collection of scientists, can stay competitive with a much larger group of Americans working on the same problems."

Fisher's foray into HIV research was largely accidental. She went to Gallo's lab hoping to work not on HIV, but on the viral oncogenes Myc and Myb to understand how they influence proliferation and differentiation in hematopoietic cells. Gallo was disappointed, she says, "but he had so many postdocs arriving, he let me do more or less what I wanted."

To do what she wanted, Fisher first had to figure out how to get those genes into human T cells. "We were using electric shock to deliver genes into cells back before there were machines to do it, so we had to build our own," she says. Using protoplast fusion, which involves loading bacteria with the DNA of interest, stripping off their cell walls, and coaxing them into fusing with T cells, Fisher was able to introduce Myc and Myb. She thought it might be worth trying the same on the HIV clones that were floating around the lab.

"People at the time thought it was a mad idea," she recalls. Her labmates had been trying to get whole, human proviral sequences into T cells, without much success. Until Fisher gave it a go with a new clone of HIV. "Lo and behold, we were able to recover replicating, cytopathic virus. That was really important," she says. "Because it meant that you had the virus in its entirety, and it was biologically active and capable of being transmitted." It also provided her colleagues with the material they needed to begin dissecting the functions of individual viral genes.

It was an exciting time, says Fisher, "but HIV research was so cutthroat and political. I just wanted to come back to England and go back to doing straight cell biology." But she brought back with her the enthusiasm she picked up in the United States. "If you talked about an experiment in the States, people would say, 'Yeah, go for it!' But if you talked about the same experiment in England, you'd get, 'Ooh, that's going to be terribly difficult.' It was incredibly frustrating," she says. "I wanted to keep up the pace and the excitement of doing high-risk experiments."

After a three-year stint at the Pierre Chambon Institute in Strasbourg, Fisher and Matthias Merkenschlager, her partner in the lab and in life, were offered an opportunity to set up shop at the fledgling MRC Clinical Sciences Center (CSC). They decided to use their partnership to approach the question of cell fate in a unique way. "I'm a cell biologist, interested in cell commitment and lineage decision-making," says Fisher, who is now CSC's director. "Matthias is an immunologist. We wanted to find a way to combine those two interests, so we started using lymphocytes as models for cell biology."

That's part of what makes their results so powerful, says Thomas Jenuwein of the Max-Planck Institute in Freiburg, Germany. "They know their system inside and out. They have a deep understanding of the steps that lead to the commitment of B cells and T cells. They also have a full catalog of target genes that are activated or repressed as lymphocytes differentiate. So they have the right tools to do the sharpest analysis" of the genetic programs—and epigenetic changes—that drive cell fate.

One of their first big discoveries was that genes that are silenced during differentiation get sequestered in a sort of transcription-free zone within the nucleus. "We were working on a protein called Ikaros, which was known to be required for lymphocytic development," she says. The discovery that silenced genes are physically ushered into areas high in heterochromatin, the form of DNA that is transcriptionally silent, "came from very simply looking at where Ikaros bound." Using a technique called 3D immunofluorescence in situ hybridization, Fisher and her colleagues found that genes which bind Ikaros were located in different places in the nucleus depending on whether they were actively expressed. Genes that were permanently shut down during lymphocyte development were tied to heterochromatin.

"That was a real breakthrough in nuclear cell biology," says Martin Raff from University College London. "It showed that there's a whole geography inside the nucleus that we knew very little about. That field has just exploded in last five years or so. And Mandy was at the forefront."

Those pioneering observations also suggested that cells have yet another level at which they can regulate gene expression, particularly during differentiation, says Luis Aragon, one of Fisher's CSC colleagues. "So when cells commit to a particular lineage, they might put genes in different compartments, depending on whether or not they're going to use them."

For Fisher, putting away genes is just as interesting as using them. "We're really interested in lineage restriction," she says. "So, an [embryonic stem] cell keeps open all its options, a hematopoietic stem cell has limited options, and a lymphocyte has only one option." Understanding how this happens raises the possibility that scientists could learn to reverse the process, reprogramming differentiated cells into regaining their former potential.

To that end, Fisher and her lab have been making heterokaryons—fusing together mouse ES cells and human B cells. "Lymphocytes are wimps, so in that situation, the ES cell dictates what the B cell is going to do," says Fisher. Within days, the hybrid cells switch on a suite of genes associated with pluripotency, and turn off the genes that make a lymphocyte.

Now Fisher is using those cells to determine exactly which genes are responsible for that nuclear reprogramming. So far she's found that pluripotent stem cells need Oct4, but can do without Sox2—factors that US scientists have used to induce pluripotency in human fibroblasts. "It's a nice system for asking, 'Is gene A important for the reprogramming process, is gene B important?'" says Young. And it's another example of how "Mandy is always taking a unique angle on things;" in this case, using heterokaryons to determine which genes are key for nuclear reprogramming.

"She also has the courage to follow new ideas, even when they go against popular wisdom," says Fisher's postdoc Helle Jorgensen. For example, biologists believed that the genes that replicate early during the cell division cycle are those that are being actively expressed. But when postdoc Veronique Azuara observed the opposite, she and Fisher decided to pursue it. In following up, the team discovered that regulatory genes that are going to be used later in development—but that are not actively expressed in ES cells—are replicated early. What's more, they sport a special kind of chromatin marking, which includes histone modifications that signal both activation and repression. In the stem cell's progeny, "if the gene is going to be actively expressed, it will lose the repressive marks. And if it's going to be silent, it will lose the other marks," says Fisher. "So in ES cells, these very important regulatory genes are essentially poised to be expressed later on." Fisher's results, along with similar findings from Young and another group in the United States, were published in 2006.

"This is an entirely new kind of mark that simply hadn't been seen before," says CSC colleague Niall Dillon. "It has the potential to confer plasticity on these key genes." And it allows them to be transcribed quickly when needed. In fact, RNA polymerases are lined up on these genes "like a queue of London buses"—primed and ready for the signal that will tell them to go, says Fisher.

It's that attention to the visuals—like the image of a backup of buses—that "marks Mandy out as being a good scientist and a good communicator," says former postdoc Julie Webb, who recruited Fisher to work on an exhibit designed to get the public excited about, and in some cases involved in, genome sequencing. The two set up a sequencing lab at the Institute for Contemporary Art in London, where visitors could speak with researchers—recruits from Fisher's real lab—who were demonstrating how to sequence a gene thought to be involved in language impairment. They even got to help read the sequence and try to identify the mutation in children affected by the disease. "It got an amazing response from the public," says Fisher. "I had people coming up to me on the bus saying, 'Did I get it? Did I get the mutation?' It was wonderful."

"It got me thinking about how to explain my work to people outside my field," says Jorgensen, who was part of the exhibit. And some of Fisher's projects even provide ready-made ways to explain that work. For example, the Fabrics of Life workshop, which paired design students with life scientists, produced "epigami"—a way to use origami to explain epigenetics. "The unfolded piece of paper represents a naïve cell that hasn't made any developmental decisions," says CSC's Brona McVittie, who helped Fisher bring the project to life. "Once you make a fold, it limits the other folds you can make. And if you uncrease the paper, the memory of the fold is still there—just as mature cells retain the memory of their developmental decisions."

Fisher is "really interested in flying the flag for scientists," says McVittie. Especially when that flag is a folded square of paper that tells a story about cell determination and fate.