Oncologist Leads Research Group To Genetics' Cutting Edge

At the University of Wisconsin's McArdle Laboratory for Cancer Research in Madison, oncology professor Waclaw Szybalski and his team are helping to define the leading edge of genetic engineering. The methods they have developed could have a tremendous impact on the much-discussed Human Genome Project, and it is therefore not surprising that Szybalski and his team of three senior researchers and three graduate students spend long hours in the lab. Indeed, even when they are not working, the researchers can be found in one another's company--at restaurants, on skiing trips, or at other social activities. The success of the team at the 50-year-old McArdle Laboratory may be partly due to this friendly relationship among its members. But it is the atmosphere within the laboratory that seems to spur the researchers on. The team operates without formal structure but with tremendous individual familiarity. Says senior researcher Noaman Hasan, "We are independent in our work, but there is a lot of interaction when there are problems." Adds Szybalski, "I am of the mind that one plus one equals more than two." University of Wisconsin oncologist Waclaw Szybalski and two of his graduate students, Michael Koob and Eric Grimes, borrowed from Greek mythology to describe Achilles' heel cleavage, the bioengineering that allows them to cut long pieces of DNA in one predetermined spot. According to the myth, Achilles was rendered virtually immortal by his mother, Thetis, when she dipped him in the River Styx. She held him by his ankle, leaving only his heel vulnerable. Achilles was killed during the Trojan War when an arrow struck his heel. In the laboratory, a chromosome represents Achilles; the desired restriction site, Achilles' heel. The sequences on the chromosome that are equivalent to the desired cleavage spot are rendered inactive to enzymatic digestion through a process known as methylation. This would be analogous to dipping Achilles (the chromosome) in the River Styx while holding his heel (the restriction site). The sequence that will ultimately be cut is marked by the introduction and binding of a lac repressor, which acts as Thetis' hand. When the lac repressor is removed, the restriction enzyme cuts the DNA in precisely one site, just as the mythical arrow felled Achilles when it struck his heel. Koob and Szybalski have used the technique to cut progressively larger pieces of DNA. Most recently, they cut a 15-megabase pair yeast chromosome (Science, 250:271-3, 1990). Grimes is applying the procedure to bacterial genes. The researchers hope the procedure will prove useful in the study of large genomes that otherwise would have to be cut by digestion with restriction endonucleases. --P.K. Szybalski, a native of Poland who emigrated to the United States in 1950, is aggressively altruistic. "I teach every student," he says. "Since society pays you for your work, which is pleasure, you have a duty to do something back for society. You have to be a preacher. It is not enough to merely discover something--you have to tell people." The team has done both in abundance. Their pursuit of new methods for the manipulation of genes is carried out in an atmosphere charged by the Human Genome Project. The researchers have written almost three dozen papers and abstracts during the last three years. Their technique, known as Achilles' heel cleavage (see story on page 17), has generated the most interest. Szybalski and graduate students Michael Koob (who received his Ph.D. in December) and Eric Grimes described the method in a series of papers (Science, 241:1084-6, 1988; 250:271-3, 1990; and Gene, 90:1-7, 1990). Perhaps the most important facet of the McArdle team's success is that the various members share similar visions for science. Each researcher is dedicated to the development of applicable techniques as opposed to the pursuit of "pure science." Koob, a doctoral candidate in molecular biology, began his project to make restriction sites unique in the fall of 1986. "I was not interested in a biological problem," he says. "I was more interested in technique development." Koob says he was attracted to Wisconsin by Szybalski's emphasis on technique. As a graduate of North Dakota State University in Fargo, he also felt he would be comfortable in the Midwest. Koob's doctoral thesis will explain another new method--one that stabilizes large strands of DNA by isolating them from cells embedded in agarose blocks. According to Szybalski, this "bead" technique is like making huge cell packages for the DNA. These packages provide stability and ease of accessibility in the laboratory. Senior researcher Sun Kim, who holds a doctorate in food chemistry, suggested using an airbrush to form the agarose beads. Koob shares his laboratory with postdoctoral fellow Adam Burkiewicz; Gyorgi, a black hooded nun finch; and a few pet goldfish. "We had Fric and Frac and Fred, but Fric died," Koob says of the fish. The bird, named for a recent Hungarian postdoctoral fellow at the lab, is a widower. "Eva, the female, had a heart attack and died," says Koob. Jokes Szybalski, "If we had a total genome for her, we could have stretched her life another year." Burkiewicz is another team member with the heart of an engineer. "I don't have a motivation for really pure science," he says. "I like things that are more useful." Burkiewicz has devised a faster plasmid mini-prep technique that will be marketed by Promega Corp., headquartered in Madison. The method screens for small DNA while eliminating large DNA, proteins, and contaminants. "This is a science that can help a lot of people's lives," says Burkiewicz. "I like to think of things that will make the Human Genome Project go faster." Grimes agrees with the importance of the group's work but adds that recognition by his peers of his research is the best part of his work. "I like to work on something with a practical application," he says. "And unless it gets out into circulation, it's wasted." The group's projects have been augmented by international collaboration as well, particularly through a partnership with Anna Podhajska, a microbiologist at the University of Gdansk in Poland, who calls the group a "Polish-American corporation." Both laboratories share a grant from the National Science Foundation. During the last two years, the Polish government has supported the joint research effort with funds as well. "It has been a surprisingly pleasant and fruitful collaboration," says Szybalski. "The spirit of the [Polish] workers, their knowledge, and enthusiasm, is great." Most of the group had an opportunity to travel to Europe last September. Koob, Grimes, and Kim flew to Amsterdam and then drove nine hours to Berlin for a poster session at the Second New England Biolabs Workshop on Biological DNA Modification. The three met Szybalski and Podhajska for a sightseeing tour, which included a visit to the Berlin Wall. Says Kim, "Instead of paying five deutsche marks apiece for a rock from the wall, we rented a hammer for five deutsche marks and, in 20 minutes, we had a piece for everybody in the lab." >From Berlin, the group continued to Gdansk, where Koob and Grimes gave a talk. While in Poland, Podhajska and Szybalski introduced their students to the Polish pastime of mushroom picking--an activity the students tried with some reservations. Says Podhajska, "I think Eric [Grimes] was a little nervous about eating them." Kim missed the mushrooms; he did not go to Poland, instead returning to Madison. He is working with Szybalski on the SPEL-6 Sequencing Technique (Gene, 90:177-8, 1990), which proposes using the ligation of hexamers to make sequential elongation primers and sequence DNA "from the top down." Additionally, Kim is pursuing a technique to detect point mutations based on the universal restriction enzyme system developed by Podhajska (Science, 240:504-6, 1988) and a new DNA amplification reaction to combine with the screening for point mutations. Senior researcher Hasan is also actively developing techniques for use in genetic sequencing. So far, he has been able to make plasmids containing vectors with mechanical usefulness. One is a promoter inversion vector to turn genes on and off rapidly (Gene, 56:145-51, 1987). Another is a trimming plasmid, which precisely deletes up to 12 base pairs of DNA from one end of a fragment (Gene, 82:305-11, 1989). His goal is to be able to express a gene for a certain time and then pull it out. Stratagene Cloning Systems in La Jolla, Calif., sells Hasan's handy promoter inversion vector. His gene inversion vector and excision vector are free for the asking. According to Szybalski, Hasan's vectors constitute the groundwork for gene therapy. "Ideally," he says, "you want to develop a plasmid with built-in regulatory circuits that could respond to changes in the body... an artificial homeostatic device. I have had the feeling for a long time that gene therapy has a lot of promise, if the activists allow it." Hasan echoes Szybalski's concerns: "Restriction to science is very harmful. In Wisconsin, the hot issue was bovine growth hormone [BGH]. Scientifically we think the use of BGH is beneficial. The opposition to its use does not have scientifically based arguments." The McArdle Laboratory researchers are aware of the concerns of those who oppose genetic research for ethical reasons. "I was a member of the Recombinant DNA Advisory Committee," says Szybalski. "I remember one lady who was always against everything. I said, `Why?' She said, `Because it makes me uncomfortable.' I said, `Who cares about you being comfortable? There are others suffering.' You have to do what's best for humanity, not what makes ethicists feel good." When challenged, Szybalski takes his message to the streets. He tells of his aborted attempt to debate activist Jeremy Rifkin on the steps of the university's student union. According to Szybalski, he was watching Rifkin address reporters on the BGH controversy when he was approached by Rifkin's supporters. "They said to me, `Are you with the press?'" he recalls. "I said, `Yes, I edit a journal.' [Szybalski is editor-in-chief of Gene.] Then they asked me to step back, and then they lifted me up and carried me away! Is this the American way?" Szybalski adds that he is suspicious of the motives of some ethicists. "I think there are many enemies of the people who are walking in sheep's clothes," he says. In spite of the moral debate, the researchers are optimistic about the Human Genome Project. "It will help the public health tremendously," Hasan says. "The knowledge we have now, in 10 to 15 years, will be a drop in the bucket." Szybalski says he always wanted to be at the forefront of science. As a boy, he thought that supersonic jets would test the limits of technology. But after the Russians invaded Poland and closed the school of aeronautics at his university, Szybalski took up chemistry, his second love. "I always like to be at the forefront of something--something which is brand new," he says. Indeed, Szybalski's group's research has expanded the capabilities of geneticists. He speaks of the Human Genome Project with confidence: "I think it will be completed much sooner than people think. In 10 years, we'll have the first experimental human genome. In 50 years, the leading university medical schools will add it to the patient's chart and, eventually, every newborn baby will have that sequence put in their chart. This will be the most important diagnostic tool." Paul Kefalides is a freelance writer based in Merion, Pa.

Oncologist Leads Research Group To Genetics' Cutting Edge

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