Read an Excerpt
Tears of the Cheetah
And Other Tales from the Genetic Frontier
By Stephen J. O'Brien St. Martin's Press
Copyright © 2003 Stephen J. O'Brien
All rights reserved.
ISBN: 978-1-250-10231-7
CHAPTER 1
A Mouse That Roared
It is considered China's golden age. From A.D. 960 to 1279 the Song Dynasty ruled over a cultural and technological renaissance that gave rise to seminal discoveries — printing, magnetic force, the compass, and gunpowder — several hundred years prior to parallel innovations in Western Europe. The Song and succeeding Ming dynasties saw a dazzling population expansion that doubled the 80 million Chinese citizens in the fourteenth century to over 160 million by the peak of the Ming Dynasty in 1650. Today, 1.3 billion people live in China.
The population boom was marked by urban sprawl and agricultural improvements. Crops and grain supplies increased steadily through the Song and Ming dynasties with expansive cultivation, growth in productivity per acre, and a geometric increase in irrigation. Through the last millennium China blossomed as an agrarian culture with 90% of its land under cultivation and less than 2% dedicated to pastures for animal grazing.
The stores of grain across China provided a ripe opportunity for rodents, particularly mice and rats as they became what scientists call commensal species — animals that flourish alongside human activities. Dogs, cats, houseflies, mosquitoes, cockroaches, and pigeons are all commensal species. Wild mice do particularly well in barns, silos, and grain stashes, producing litters of up to a dozen pups each month. As China's agriculture flourished, so did its mice, which prospered to numbers in the millions, if not billions.
Then, some time deep in the Middle Ages, a ferocious disease outbreak devastated Chinese mouse populations. Such a plague would likely have been welcome to the peasants and farmers of the time. To the scientists who stumbled upon this obscure history centuries later, the plague would reveal itself as an unusual and profoundly significant evolutionary event. What they uncovered about these mice would change the way biologists and medical detectives perceived the history of wild species.
The devastating virus that hit these mice caused blood cancers, lower limb paralysis, and paraplegia. By the thousands these overcrowded, stressed mouse communities would succumb to the fatal disease. No one is certain where the virus came from — maybe from domestic cats brought in to dispatch the barn mice, or from birds or livestock. The epidemic killed tens of millions of mice before the tide changed. Somehow certain mice survived and continued to reproduce, feed, and spread, once again to be limited only by available food, predators, and opportunities to set up a nest. It was a narrow escape.
How did the survivors avoid the menacing virus that felled so many mice? The puzzle was solved centuries later by a seasoned medical pathologist whose curiosity and scientific acumen uncovered the eerie scenario as if peeling away the layers of an onion until the prize core, the puzzle's solution, was his.
* * *
Dr. Murray Gardner spoke softly but deliberately to the high school boys just after midnight. "The key is stealth. Nothing is illegal here, but be very careful not to be seen and talk to no one. The last thing we need is to wind up in the newspapers; that would stifle the whole operation."
The boys were amateur mouse catchers, equipped with cotton work gloves, dark plastic garbage bags for the take, and miner's helmets with stun lights to freeze/terrorize the mice so they could grab them and toss them in their sacks. The hunting grounds: a squab farm near Lake Casitas in south Ventura County, California, forty miles north of Los Angeles. Squabs are pigeons raised as a delicacy for Chinese restaurants, and the farm had ten thousand of them brooding in small pens. Under the dung-saturated pen beds lived hundreds of house mice, Mus muscúlus domesticus, quietly pilfering the squab's grain feed.
Gardner needed the mice to search for new retroviruses, a nasty type of virus that causes cancers, notably leukemias and lymphomas in chickens, cats, and mice. Retroviruses are unusual in that their genes consist of RNA — nucleic acids that control cellular activity — rather than standard DNA. These viruses use an enzyme to copy their RNA genetic code into DNA form, which it then inserts into its victim's DNA. RNA is usually a product of DNA, not the other way around — hence the prefix retro. Murray Gardner was keen to sample wild mice because like humans, but unlike most lab mice, they had not been bred artificially to shed or minimize genetic diversity. The mouse retroviruses isolated before Murray's hunts all came from inbred strains, mice descended from twenty or more generations of brother-sister incestuous matings.
Murray had the farmer's permission to collect the mice and take them to his laboratory, but only if he kept the operation quiet. The reason for the secrecy was simple: Squabs are raised for human consumption and farms that raise them face a monthly inspection by the State of California Rodent Control Board to certify that the farms are "rodent free." Once the mice were exposed, forced traps and poison would be inevitable. So the boys collected the mice quickly and before dawn dropped their catch in a drum at a rural gasoline station to be intercepted by Gardner.
Murray Gardner is a curious and sometimes impatient fellow. In another life, he may have been a Sherlock Holmes–style detective, a Greek philosopher, or even a charismatic statesman. A real-life Hawkeye Pierce, Murray served as a MASH doctor in the Korean War, patching injuries and administering obstetric care to frightened young Asian women who carried GIs' babies. After the war, he trained as a medical pathologist and joined the University of Southern California faculty as a medical researcher in 1964.
By 1970, Murray was forty-one, professionally successful, and well settled in his academic routine. His clinical and teaching responsibilities were important but not particularly challenging. In his spare time he consumed scientific articles on cancer, infectious diseases, and medical advances with demonic passion. His first research project was an attempt to show that Los Angeles smog caused cancer in lab mice. He had heard about President Nixon's War on Cancer, a kind of "moon shot" ambition to understand and cure the disease. The program infused millions of government dollars from the U.S. National Institutes of Health (NIH) into targeting causes, diagnostics, and new treatments for cancers. A large part of the new cash went to the Virus Cancer Program, an expansive effort launched by the National Cancer Institute (NCI) to discover human viruses that would cause cancer.
The Virus Cancer Program, which lasted from 1968 to 1980, was sadly short-lived because no human cancer-causing viruses were immediately discovered and critics succeeded in pulling the financial plug by arguing that viruses had nothing to do with human cancer. Today we know that several human viruses cause cancers responsible for hundreds of millions of deaths. Papilloma viruses are the prime cause of cervical cancers; hepatitis B virus leads to liver cancer, affecting 300 million people across the world; and HIV leads to lymphoma, Kaposi's sarcoma, and other tumors in AIDS patients. In retrospect, the Virus Cancer Program was hardly misguided but cutting edge — a research effort ahead of its time.
Murray Gardner was busy exposing laboratory mice to smog and auto emissions on interchanges of Los Angeles freeways when Robert Huebner, a leader of NCI's Virus Cancer Program, asked for his help. Huebner's research group had identified many cancer-causing retroviruses in lab mice, but he worried that the intense inbreeding of the lab mice had compromised their discovery.
Huebner knew that outbred species like humans and wild animals had some thirty-five thousand genes as their genetic base and that nearly every gene had some level of genetic variation. If such diversity includes genes that specify immune response to viruses or other infectious diseases, then the inbreeding of laboratory mice may have inadvertently eliminated key genetic regulators for virus replication and virulence. This hunch would offer a plausible explanation for the ease with which multiple tumor viruses were harvested from inbred mice and chickens, while none had been found so far in humans. Perhaps retroviruses were really present in humans, but were actively repressed by our genetic diversity. The idea made evolutionary sense because virus-repressing genes would provide a real benefit for out-bred species: the prevention of viral-induced cancers.
Huebner and Murray reckoned that such cancer-causing viruses might be lurking in outbred species, but suppressed by the species' genes into a latent form. They agreed that a search for such agents in the wild might uncover some very interesting natural microbes in a few rare sensitive mice, ones that were the forebears of the tumor viruses that had been discovered in lab mice. All they needed was to get their hands on some wild mice and take a look using standard virological tools.
The hunt was on — at dairy farms, racetracks, aviaries, alleys, birdseed factories, freeway trellises, and any place wild mice might be lurking. Somewhere between ten thousand and twenty thousand mice were collected by Gardner's clandestine operations over the next decade. He paid the boys ten cents a mouse. After scores of false starts, narrow escapes, and raised eyebrows, Gardner managed to capture mice from fifteen locales in the greater L.A. area. He watched them age and searched for cancer, retroviruses, and other viral diseases. Nearly all the mice were free of cancer, retroviruses, or other infections except for a few sporadic tumors in aging mice. Nor did Gardner find the natural microbes they thought might echo tumor viruses in lab mice. The exception was the squab farm near Lake Casitas.
The mice nestled in the squab bins were different. They were battling a massive epidemic of lethal retroviruses. Nearly 85% of the farm mice carried evidence of exposure to one of two ravaging viruses. The more virulent strain was called ecotropic MuLV, murine (i.e., mouse) leukemia virus. Ecotropic means that the virus grew in mouse cells in the lab, but not in cultured cells from other species like human, rat, or cat. Murray isolated and injected the wild mouse virus into lab mice; it caused a fatal spinal hind limb paralysis and in older animals a blood cancer called "lymphoma." The spinal paralysis was termed spongiform polio-encephalomyelopathy, and it developed in the wild mice as early as ten months after birth. The virus killed the wild mice it infected and was transmitted to offspring through breast-feeding.
At last, a wild mice population laced with a fatal retrovirus had been found. Gardner could now study a cancer-causing virus in a population that more closely resembled the genetic diversity of humans and other outbred species.
In scientific inquiry, it seems that the more you discover, the more new unanswered questions appear. Murray sampled the Lake Casitas mouse population monthly for several years, fully expecting the paralytic virus to sweep through the pens and extirpate the mice. That did not happen. The mice flourished at high density for a decade and disease incidence stayed at around 15%. Infected mice died quickly, but 85% of the mice never succumbed to the virus and were never paralyzed. There seemed to be a powerful check on the impact of the deadly virus within the population.
I first heard this curious saga of the lucky mice from Murray at the 1977 Annual Conference of the Virus Cancer Program. The apparent freeze-frame nature of the epidemic was tantalizing. How could a generic mouse population tucked beneath a squab bed in rural California survive a lethal epidemic indefinitely? To me it seemed like a genetic difference, but in those days we were more used to thinking of variable genes as predispositions to hereditary genetic disease like sickle cell anemia or cystic fibrosis. A gene that could block a fatal virus infection would be particularly interesting, so together we set out to find it.
Murray was anxious to enlist my help because in the early 1970s there were few geneticists who also studied retroviruses. My training as a Drosophila (fruit fly) geneticist at Cornell exposed me to experiments in genetic transmission plus gave me a foundation in population and evolutionary genetics. Population genetic experts were tracking fruit fly populations in cages or in natural settings to view patterns of genetic variation that were the precursors to species adaptation and species formation. My postdoctoral fellowship at NCI brought me shoulder to shoulder with the giants of retrovirology, who showed me the fascinating complexity of tumor virology. Murray asked me to help him find out if Lake Casitas mice harbored a resistance gene to defend against the deadly retrovirus and, if so, to figure out how it worked.
To get at the explanation, we needed first to prove the resistant mice really had such a gene and could transmit it to their progeny. We were not sure how to do this, but we had some tricks from classical genetics that I learned from working with fruit flies. Murray was an attentive and enthusiastic student.
Early on, Murray discovered that Lake Casitas paralytic virus was genetically related to another laboratory retrovirus, one that caused a very high cancer incidence in an inbred mouse strain called AKR. This connection would soon prove to be the key that allowed us to get at the Lake Casitas wild mouse resistance.
AKR mice had been inbred extensively in the 1920s and selected by their breeders for a high incidence of leukemia, so much so that 100% of each generation develop leukemia and die before their first birthday. Some rather elegant experiments in molecular biology in the early 1970s showed why. AKR mice carried three full-length copies of a murine retrovirus, termed AKV (for AKR virus), in their chromosomes, nestled between the normal mouse genes for making a mouse. These three AKV genomes (a genome is a full-length copy of the viruses' genetic endowment) are passed down from parents to offspring on the chromosomes of sperm and eggs just like all the regular genes. In effect, the viruses are hitchhikers on the mouse's chromosomes.
After birth, something triggers these latent endogenous viruses to start replicating themselves and spread through lymphocytes (white blood cells). Endogenous means the virus lives in the host chromosomes and is passed vertically to offspring, in contrast to exogenous viruses like flu or smallpox, which spread horizontally between individuals. What these released endogenous retroviruses do best is cause leukemia. They accomplish this feat by infecting a lymphocyte and inserting themselves into a chromosome adjacent to one of a few hundred mouse genes that, when expressed in the wrong cell, lead to uncontrolled cell division, or cancer. The AKR virus simply activates the adjacent gene by providing the genetic equivalent of a "turn me on!" signal to the cellular machinery that decides which genes get expressed. The process transforms the infected lymphocyte into a wildly uncontrolled dividing cell, the first step in leukemia. The AKR mice have high virus titers (concentrations) in their blood and die of cancer as a consequence. Could the Lake Casitas (LC) resistant mice also block the AKR virus? If so, then we could conclude that the wild mice must have carried an anti-retrovirus resistance gene.
Murray set up two mating crosses, one between AKR mice and the LC mice that were infected with the wild mouse virus and a second cross between AKR mice and LC mice that were virus-free and presumably resistant to the virus and its paralysis. The first cross produced offspring that developed AKV virus production, leukemia, and early death, as in their AKR parents. But the second cross was very different. Several virus-free LC parents, when crossed to AKR mice, produced dozens of offspring with no viremia, no leukemia, no paralysis, and long life. Something carried in the chromosomes from the LC virus-free mice had completely shut down the AKV virus in the hybrid offspring. It had to be a resistance gene in the LC resistant mice that neutralized not only the wild mice virus, but also its close relative, the AKV nested in mouse chromosomes.
Other virus-free LC parents produced two categories of offspring in about equal proportions: one group of pups was highly viremic (that is, riddled with virus particles throughout their bodies and bloodstream) and the second completely virus-free. This pattern made sense to a geneticist. The virus-free LC mice with no viremic offspring carried two copies of the wild mouse resistance gene; the LC virus-free mice that produced both viremic and virus-free offspring had one resistance gene and one sensitive gene.
To nail our conclusion further, Murray then crossed the resistant AKR-LC hybrid progeny back to their AKR parents. This mating produced a 50:50 ratio of virus-free mice to viremia/leukemia-riddled offspring. That 50:50 split is precisely the prediction of Gregor Mendel's first law of genetics, the law of single gene segregation. The crosses left no doubt in our minds that the LC mice had a genetic gold nugget on one of their chromosomes, a powerful force that protected its carriers from retroviremia, leukemia, and early death. Murray and I named the new gene AKVR for "AKV restriction."
(Continues...)
Excerpted from Tears of the Cheetah by Stephen J. O'Brien. Copyright © 2003 Stephen J. O'Brien. Excerpted by permission of St. Martin's Press.
All rights reserved. No part of this excerpt may be reproduced or reprinted without permission in writing from the publisher.
Excerpts are provided by Dial-A-Book Inc. solely for the personal use of visitors to this web site.
