Turning Back the Clock

Coleen Murphy had her first child at 38, her second at 41. More than most people, Murphy, a molecular biologist, understood the risks of waiting until she was in her late 30s to build a family: It can be harder to conceive, and birth defects are more likely. But she was dedicated to an all-consuming job as a rising young scientist. “I used to do experiments around the clock, every four hours for three days straight,” she says. “You can’t do that when you have kids.” So she waited: “It was my idea to postpone and gamble a little bit.”

For her, it turned out to be a good bet. As tenured faculty members, Murphy and her husband, molecular biologist Zemer Gitai, are able to manage their schedules, making it easier to balance scientific careers with raising their first-grade son and 3-year-old daughter. “There are times when you can put in a lot of effort to get your career going and other times when you have more flexibility,” Murphy says. The problem for a woman who hopes to have children is knowing how that schedule will fit with the workings of her own body.

Murphy is striving to lessen the risk of waiting to conceive. She believes that in the not-too-distant future — she can’t say exactly when — it will be possible to give a woman a blood test that could provide an individual biological clock, telling her the time frame in which she is most likely to conceive and with less risk. Perhaps a drug could be developed to slow the aging of a woman’s egg cells, a major cause of declining fertility. “A lot of women would be interested in that!” suggests Murphy, now 44, with enthusiasm. An even bolder prospect: Research might provide treatments to allow people to escape common diseases and conditions of old age, such as Alzheimer’s disease and loss of muscle tone, altogether.

An associate professor in molecular biology and Princeton’s Lewis-Sigler Institute for Integrative Genomics, Murphy studies the processes of cognitive and reproductive aging. She and the members of her research team want to understand what is happening at the molecular and cellular level to govern how quickly we age, and why some cellular processes stop working earlier than others. Murphy’s work won’t let us live forever, but it could postpone or even prevent the deterioration in quality of life that most of us experience as we age.

“We used to think that aging was an unavoidable result of living, simply a matter of cells wearing out over time,” Murphy has written. But studies of different organisms — the roundworm Caenorhabditis elegans that she works with as well as yeast, flies, and mice — have found that a mutation in just one gene can dramatically increase life span, she says. This suggests that aging is not merely an issue of haphazard wear and tear, but that it follows a regulated process — and that the impairments associated with the aging process can be disrupted.

Although there is a genetic basis to aging, scientists don’t think that an individual’s life span is a fait accompli. Our environment, diet, and exercise trump genetics in most cases. Murphy and her research team are trying to understand just what role genetics do play. How does each part of the complex system that is the human body break down with age? How do the parts interact? What can be done to prevent breaks, or to repair them?

The molecular biology of aging is a relatively new field, pioneered 20 years ago by Cynthia Kenyon, a professor at the University of California, San Francisco, who was studying development in C. elegans. Kenyon was working with worms that had a gene mutation that caused them to have few progeny, rather than the hundreds that worms typically bear. Kenyon usually studied the worms during their first week of life and then discarded them. By chance, she left a dish of the worms in an incubator for about a month. And when she returned to it, she found that the worms had a characteristic feature of old age — their movements were stiffer and slower than those of their younger counterparts. “My breath was taken away,” she says. “The fact that worms get old just like people stuck with me and made me want to study aging.”

In those days, most scientists believed that aging was the unstoppable and random process of accumulating damage and wear and tear to our tissues. But Kenyon reasoned that aging, like development, was a regulated process, meaning that there was “some kind of dial that you could turn to make aging go faster or slower that had to do with our the genes and was something more than just random damage.” That thought was almost heresy in scientific circles, Kenyon recalls.

Then, in 1993, Kenyon shattered the prevailing view. Her laboratory presented evidence that a single mutated gene can double the life span of C. elegans while leaving the worm fertile and healthy. The worms’ mutation was in the daf-2 gene, the gene for the insulin receptor. And the cause of the worms’ longevity, Kenyon’s team determined, was a protein regulated by the daf-2 gene.

Murphy was finishing her graduate work at Stanford, studying a protein that is responsible for the contraction of muscle cells, when she heard Kenyon lecture on her research on insulin signaling and aging. Murphy was intrigued, and in 1999, she joined Kenyon’s lab. Using a then-new technology called DNA microarrays that enables the study of large numbers of genes simultaneously, Murphy analyzed all 20,000 C. elegans genes during her time there — a critical achievement that identified differences in gene-expression patterns between normal and long-lived worms.

Could the same processes uncovered in 1-millimeter-long nematodes barely visible to the naked eye also take place in humans? The daf-2 gene counterpart in humans is the insulin receptor that regulates metabolism, growth, and stress. Since Kenyon’s pioneering work began, other studies have shown that mutations in the daf-2 genes of mice, dogs, and other mammals also extend life span. Mutations in these genes have been linked to people who live more than 90 years.

Murphy has been building on those findings since arriving at Princeton in 2005. Like Kenyon, she used the unassuming roundworm in her research. A general premise in science is to start simply to gain understanding and then scale up. C. elegans, originally chosen for the study of neurology and development — the worm is transparent, so its cells are easily observed and tracked — has been a boon for aging studies. The roundworm has a life span of three to four weeks, making it feasible to do aging studies relatively quickly, and is one of the simplest organisms to have a nervous system. With exactly 959 cells in the adult body (the human body is thought to have at least a trillion cells), C. elegans is a simple-enough system to be understood entirely. The genes identified then can be tested in human studies.

“We share a large fraction of our genome and proteins with worms, so often what we find in worms ... ends up having correlates in humans,” Murphy explains. “That has been true of almost everything else that the C. elegans field in general has found, which is why it’s a great model system.”

She found backing for her research from the W.M. Keck Foundation and, more recently, from Paul F. Glenn ’52, who has been interested in the biology of aging since he was a Princeton undergraduate. In 1965, he established the Glenn Foundation for Medical Research to fund studies on aging, with the goal of extending the “healthy and productive” human life span. While most foundations, he says, focused on specific diseases related to aging, he was more interested in the aging process itself. Today, his foundation supports aging research at eight institutions, including Harvard, Stanford, the Mayo Clinic, and Princeton, where a $3 million grant allows Murphy and her team to study the networks of genes that are essential for higher neural activities, including memory and cognition.

Her work has implications for understanding and preventing Alzheimer’s disease and related conditions, but her approach makes her a maverick. Most research on Alzheimer’s, which affects more than 5 million people in the United States, is focused on the buildup of a protein called amyloid-beta in the brain that is thought to lead to cognitive decline. But Murphy thinks amyloid-beta may be a consequence of a genetic cause of the disease — not the cause itself — and that eliminating the buildup of so-called amyloid plaques won’t solve the problem. “I think we need to get back to genetics studies and ask how memory actually works,” she says.

“Aging is such an interesting and philosophical question,” Murphy says, as she sits in her office in the Icahn Laboratory. “It’s still one of the big questions in biology.” In the lab next door to her office, where postdocs, graduate students, and undergrads are at work, shelves hold reagents in glass bottles, plastic bottles with bright blue and Princeton-orange tops, micropipettes, and stacked Petri dishes. Those dishes are the laboratory habitats of the tiny C. elegans worms.

Murphy describes her experiments with the zeal of a graduate student who has just tasted the thrill of finding something new about the world; one of the first graduate students to join Murphy’s lab at Princeton, Amanda Kauffman *10, says she was attracted to the lab not just for the chance to do groundbreaking science but by Murphy’s excitement about it. “Coleen really instilled a need to be fearless, to not be bound by your own field, but to learn something new,” Kauffman says.

At first, Murphy, Kauffmann, and others in her lab were looking only at questions of the life span of the worms. But scientific curiosity leads to unexpected paths, and soon they were studying issues related to quality of life. Roundworms reach young adulthood by the third day of their three-week life. The worms, once fast and nimble, begin to show visible signs of aging by Day 10, similar to the signs we associate with old age in people. The worms become slower and less mobile; ultimately, they cannot move beyond wiggling their heads. It seemed clear, then, that their muscles were deteriorating. But how about their nervous systems, and the effect of aging on cognition? How could researchers even measure cognitive decline in a worm?

Kauffman came up with short- and long-term memory tests for the worms. Previous memory tests for C. elegans had been based on negative learning: The worms would learn to avoid certain chemicals placed in their environment because of their odor. But Kauffman and Murphy wanted to understand whether the worms were capable of making positive associations and developing and retaining long-term memory — something no one had shown before. Kauffman worked with three kinds of worms: those with the regular daf-2 gene, those with the mutant gene, and those that were put on a calorie-restricted diet. (Worms on the diet were known to live longer than regular worms.) She hoped to train the worms to associate a neutral odor with food, respond by moving toward the food source, and remember the association hours or days later.

First, Kauffman fed the worms in the presence of butanone, a chemical that neither repelled nor attracted them. To prompt the worms to form long-term memories, she provided seven “training sessions.” At different points, she would test whether worms of different ages would move toward the chemical — whether they would remember that butanone meant food was present.

Indeed, all the worms were capable of learning new tricks — but there were differences. After one 30-minute training session, the worms without the gene mutation associated the odor with food for about two hours. After seven sessions, the association generally lasted 16 to 24 hours, which is equivalent to a few years in the life of a human. By Day 4, that long-term memory was gone. By Day 9, the worms had lost their ability to form new short-tem memories.

The worms with the mutant daf-2 gene, who lived longer, did better. The short-term memory of these worms lasted about three times longer than those without the mutation, and their long-term memory was better, too. The worms with calorie restrictions were especially intriguing: They developed long-term memory more slowly than the others (requiring more training sessions than the normal worms), but the long-term memory that they did create remained with them longer: By Day 4, when long-term memory was gone for all the regular worms, worms on the diet showed they still had some.

The findings provided a clue about the genes involved in memory function. Comparing the activities of genes of the different worms, Murphy and Kauffman narrowed in on a gene called CREB, which is known to be necessary for long-term memory in many organisms, including humans. Levels of the protein produced by CREB decline with age.

Among the worms, the calorie-restricted worms had started with lower levels of CREB, but maintained their levels of the protein into adulthood — just as they retained long-term memory longer. “We don’t fully understand the role of CREB yet, but there is a molecular reason why these worms behaved differently,” says Murphy. She also is working to connect the memory-related behavior Kauffman observed with neuron activity and the rate at which the neurons degrade with age: “We think that different neurons degrade at different rates, which is also true in us,” she says.

Working with C. elegans, Murphy also has made strides in understanding reproductive aging and how a woman’s reproductive window — the oft-dreaded biological clock that ticks away the time available for conceiving a child — may be extended. Unlike men, who produce sperm continuously throughout their lives, women are born with all the eggs — called oocytes — that they will ever have. As women age, the quality and quantity of those oocytes decline, making it more difficult to become pregnant.

Murphy and graduate student Shijing Luo have shown that oocyte quality in the roundworm is regulated by hormone pathways, including the insulin pathway, and that the same pathways are responsible for the processes in humans. Mutations in these pathways can as much as double the reproductive period of a worm’s life by maintaining healthy oocytes, Murphy says. Now, she is testing which genes decline with age in worms — information that eventually could be used to prevent fertility decline in women.

“Such information would empower couples in making decisions about the timing of major life changes, all of which are today made in the context of profound uncertainty about future reproductive options,” says Steve McCarroll, a professor of genetics at Harvard who collaborates with Murphy on aging and reproduction research. “She has put a finger on an important question. Coleen is a leader in studying the reproductive life-span question, which is not only important but also scientifically tractable,” he says.

What’s next in her lab? The day before her interview, Murphy participated in a triathlon, and as she considers the question, she glances at her legs, where the partly rubbed-off numbers from that event still are visible.

“Now I want to focus on knees!” she says. She laughs, but she’s serious: She wants to understand why the structures of the physical body break down in normal worms but not in the daf-2-mutant worms.

Then she adds: “There is a little part of me as I am getting older that is not sure it wants to be so close to the problem.” 

Anna Azvolinsky *09 is a freelance science writer and frequent PAW contributor.