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SARS, Malaria, and the Microarray

It was the first Saturday of Spring 2003, and Joe DeRisi and his postdoc David Wang were staked out at either end of the University of California, San Francisco's Genentech Hall waiting for the FedEx truck.

By | November 21, 2005

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Photo: Felix Aburto

It was the first Saturday of Spring 2003, and Joe DeRisi and his postdoc David Wang were staked out at either end of the University of California, San Francisco's Genentech Hall waiting for the FedEx truck. DeRisi, had recently moved his lab to the university's new Mission Bay campus. With the lot still surrounded by chain link fencing and the loading dock closed for the weekend, DeRisi and Wang feared that the FedEx driver might get discouraged and depart without delivering their package: samples of genetic material extracted from the causative agent of SARS.

At the time, SARS was killing patients and healthcare workers in Southeast Asia. Diagnostic tests had until that point failed to identify the culprit. DeRisi thought he could help. Over the previous year, DeRisi, Wang, HHMI investigator Don Ganem, and their UCSF colleagues had engineered a microarray that sported sequences from all known viruses – some 12,000 spots representing 1,000 different viral species. By including highly conserved sequences from all the completely sequenced viruses in Gen-Bank, the researchers hoped that they would be able to detect known viruses and to identify viruses that had not been previously characterized, but would be related enough to cross hybridize with the probes on their chip.

By the time SARS struck, DeRisi and colleagues had used the array to successfully identify 10 or 20 viruses from cultured cells and patients. "But we hadn't really applied the technology to try to discover any novel viruses," says Wang, now at the Washington University School of Medicine in St. Louis. "SARS was the perfect test case," says DeRisi. "We literally begged the CDC to send us samples."

When the FedEx truck arrived, DeRisi and Wang whisked the samples back to the lab, where they set out preparing them for microarray analysis. By 1:00 a.m., the researchers were ready to set up the overnight hybridization reaction. In the morning they returned to analyze the data, scanning the 12,000 spots and examining the ones that were giving off a signal. "By Sunday afternoon we'd convinced ourselves that we were looking at a novel coronavirus," says Wang. Monday morning the CDC held a press conference to announce DeRisi's results, which corroborated their own findings. "It was an intense, exciting experience," says DeRisi.

FROM VIRUSES TO PARASITES

The discovery brought DeRisi full circle back to his roots in virology. As a graduate student, DeRisi joined Pat Brown's lab at Stanford University to study how retroviruses integrate into the host genome. But he soon found himself sidetracked by more technological challenges. The Brown lab had been working on a technique for identifying stretches of DNA passed from parent to offspring. DeRisi got caught up in figuring out how to spot the necessary DNA fragments onto glass chips. In short order, he and labmate Vishy Iyer assembled a chip containing the entire yeast genome, a feat that enabled them to do gene-expression studies on a massive scale. "In the end my thesis became about whole-genome analysis of gene expression rather than retrovirology," says DeRisi.

The work earned DeRisi a position as a fellow at UCSF, and within a year, he was offered a assistant professorship. When DeRisi was recruited, "he was barely out of swaddling clothes," notes UCSF chancellor J. Michael Bishop. "But there was a quick initiative to get him on the faculty so we wouldn't lose him," he says. "Joe sets a spectacular example for students and postdocs because of the way he does science: full throttle, daring, and socially driven."

Among DeRisi's daring, socially driven projects is a study of the biology of Plasmodium falciparum (see related story, p. 19). The first step toward understanding Plasmodium physiology involved tracking the parasite's gene-expression profile as it proceeds through the 48-hour blood stage of its life cycle: invading red blood cells, consuming all their hemoglobin, replicating, and then reinvading a raft of fresh erythrocytes. To do this, DeRisi and his team set up a bioreactor containing five liters of dilute human blood infected with Plasmodium that had been synchronized so that all the organisms would be at the same stage of their life cycles. The researchers then withdrew samples every hour for nearly two-and-a-half days and analyzed the extracted transcripts on a malaria microarray they had designed.

They discovered that Plasmodium has "a unique transcriptional profile unlike any eukaryotic cell we've ever looked at before," says DeRisi. "It's one continuous rolling cascade of expression." The results, he says, could point to "some chinks in the parasite's armor" that researchers can exploit in designing new antimalarial therapies.

In collaboration with Kip Guy, then at UCSF, DeRisi and his colleagues have also synthesized libraries of compounds based on the quinoline backbone structure of antimalarial drugs such as chloroquine. Using a high-throughput technique that employs flow cytometry to assess the growth of parasites in 96-well plates, the researchers have isolated a half-dozen small molecules that seem to inhibit Plasmodium replication. DeRisi says he is testing these compounds in mice to see if they can prevent infection without being toxic.

FIVE LITERS OF BLOOD

"I remember when Joe was really excited about growing malaria in five-liter cultures so he could do a full time course of infection," says Jim Wilhelm of the Carnegie Institution in Baltimore. "He found some source of blood at the hospital, but it turned out malaria didn't like it," says Wilhelm. "So he had to switch to bleeding people in the lab." That degree of perseverance is "typical Joe," says Wilhelm, who collaborated with DeRisi to develop a technique for immuno-precipitating cellular proteins and identifying any RNAs associated with them. "When he sees something worth doing, he does it. He doesn't let anything stand in his way. If he needs five liters of blood, he finds five liters of blood. I had no doubt that experiment was going to get done."

Last year, DeRisi, now an HHMI investigator, was named a MacArthur fellow. The award was "an amazing gift," says DeRisi, who is using a portion of the prize money to support malaria studies in Uganda. Over the summer, DeRisi sent a pair of students to work with UCFS's Phil Rosenthal to help set up a lab for culturing Plasmodium and to collect samples of the parasite from infected patients. "The strain that we study in lab has been in culture 30 years," says Wilson, who was part of the advance team. "By getting a fresh new source of parasites we can start to ask questions about what's different between a strain that's been adapted to culture for 30 years and one that's fairly new."

The first trip was a success. "When we left, the lab was up and running," says Wilson, and they brought back a handful new strains they've succesfully adapted to culture. What's more, Wilson says the trip to Africa allowed him to encounter the disease close up. "I'd never seen a person with malaria," he says. "I'd never slept under a mosquito net." Such firsthand experience helps to remind everyone involved of their long-term goals. "I think Joe has a real commitment to trying to make the world a better place," says Jennifer Gerton of the Stowers Institute for Medical Research in Kansas City, Mo. "He's really motivated by a desire to help humanity."

The projects are technically challenging. "It has always been really fun for me to build a robot and then write the code that drives it," says DeRisi. Of course, in the end, the biology is the thing. "We're not trying to build the most efficient machine ever. We take a brute-force approach and make something that's going to work day in, day out," says DeRisi. "We have to be result-driven here, because we're going to get grants for the biology we produce, not for the elegance of the machines that we make."

OPEN-ACCESS ARRAYS

At the same time, DeRisi labors to make microarray technology available to the masses. "He's like a missionary in terms of bringing genomics approaches to other people," says Wilhelm. In addition to establishing the UCSF core facility, DeRisi has run courses in Cold Spring Harbor and Santa Cruz to teach participants how to make their own arrays. The classes are "a tour de force for Joe and pretty grueling for the students," says Gerton. "But if they're motivated, I think the students can learn pretty much everything they need to know to take that technology back to wherever they hail from and get it going." For those who don't sign up for the course, DeRisi posts all the secrets of microarrays on his website, where he offers software, protocols, and the instructions for building an arrayer, including a complete list of parts and prices.

"A lot of people can't afford to spend $500 per experiment on their microarrays," says Gerton. By enabling individual labs to churn out their own arrays, she says, DeRisi encourages researchers to repeat experiments and collect data points in triplicate, an approach that makes for stronger results. All in all, says Wilhelm, "Joe really enables people around him to do a lot better science."

Although he rarely lifts a pipette these days, DeRisi still engages actively in dozens of collaborative projects. "He just can't help himself," says Gerton, who has worked with DeRisi to map the double-strand break sites that lead to meiotic recombination in yeast. "His interests are very broad and if he can think of an interesting way to attack a problem, off he goes. He must have close to 100 publications with collaborators."

"He has an insatiable curiosity," agrees Bishop. "But he also has a laser focus. No matter how many interests Joe manifests, he's sufficiently focused to be productive and to stick with something until he's got what he wants out of it."

DeRisi legendarily limitless energy reserves certainly don't hurt. "Joe is like a little pocket rocket," says Wilson. "He's got a ton of ideas and more energy than anyone I've ever known." Wang agrees. "A description I've heard many times is that Joe is like the Energizer Bunny on crack," he laughs. "I can't remember who said it first, but I think it's quite accurate."

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