Q&A with David Roth, M.D., Ph.D.
The Irene Diamond Professor of Immunology and Chairman of the Department of Pathology
It took scientists nearly 100 years to solve the particular riddle wryly known by the acronym GOD (“generation of diversity”): How can the body generate a seemingly limitless repertoire of antibodies to meet an equally limitless repertoire of possible pathogens? The Nobel Prize-winning answer arrived in the 1970s in the form of a biological Pick-3 Lotto. The immune system randomly selects pieces of DNA from arrays of V (for variable), D (for diversity), and J (for joining) gene segments, clips them from the chromosome, and pastes them together using specific DNA repair machinery to form a new gene. By 1990, scientists understood that this process, called V(D)J recombination, is initiated by two proteins, Rag1 and Rag2.
Dr. David B. Roth has long been pondering how DNA breaks are repaired. Since DNA damage underlies many diseases, including cancer, the immune system is engaging in a risky process. One way developing lymphocytes (precursors to B and T cells) reduce this risk is by having the Rag proteins guide both the cutting and pasting phases of V(D)J recombination. They also allow repair only through the NHEJ (non-homologous end joining) pathway. In a recent study published in Nature the Roth laboratory identified an alternative DNA repair pathway that is not normally accessed by the Rag proteins yet appears to be quite active in the cell. It turns out that Dr. Roth had stumbled onto this alternative pathway as a graduate student in 1985.
Q: Isn’t it ironic that this newly identified pathway is the same one you had studied as a grad student?
A: Yes, it is kind of ironic. But science can be like that. We can get enthusiastic about new data and forget about old data that don't fit. What happened was that genetic experiments identified several proteins involved in the classic NHEJ pathway, and everyone jumped on that bandwagon and sort of didn’t know what to make of the earlier stuff. Now we've come full circle, and are back to studying the first "alternative pathway."
Q: How did you become interested in an alternative pathway?
A: We had known that when you knock out the NHEJ pathway in laboratory animals, they become hypersensitive to radiation, which causes double-strand DNA breaks, and they make no T or B cells [immune cells]. So there were no V(D)J junctions and the field surmised that this must be the one and only pathway for repairing DNA. But there must have been some ability to rejoin DNA, because mice could still develop and survive. However, the mice generally developed lymphoid [immune system] tumors.
Q: So your laboratory created mutations in the Rag proteins to investigate how mice with these malfunctioning proteins still managed to survive?
A: Yes. We showed that clearly there is a novel repair pathway. It looks like this novel pathway generates chromosomal rearrangements and we think this is the pathway that leads to lymphomas in mice. We have to see if this pathway is involved in human leukemias and lymphomas.
Q: Has this novel alternative pathway opened up a new area of research?
A: It appears so. Many people in the repair field are now trying to chase down this alternative pathway. We don’t know its biological significance, but there are hints that it may be upregulated in cancer. It seems that the classical pathway prevents chromosomal translocations. Somehow it can identify DNA strands from the same chromosome and keep them together.
Q: What is the next step?
A: The most logical thing to do is to put a Rag mutation into mice and monitor their lymphoid development. What happens to an absolutely normal mouse that doesn’t use the right pathway? Does it get cancer? Another obvious step is to figure out what the alternative pathway proteins are, and we can use the Rag mutants to figure that out.
Q: Are you ever humbled by nature?
A: Constantly. It is what I like about being a scientist. You always think you know how things work. But almost every time we have a really great model, it turns out to be wrong. Our job is never done.