Sloppy Copying--New DNA Polymerase Described

Researchers from University of Southern California (Los Angeles), Rockefeller University (New York), and the National Institute of Child Health and Human Development (Bethesda, MD) report the discovery of a new DNA polymerase, pol V, which, unlike its predecessors (pol I, II, and III), is error-prone. This enzyme is part of the damage-inducible UmuD'₂-C complex, and is believed to be responsible for mutations in DNA caused by exposure to UV light. This system, described in the August 3 issue of the Proceedings of the National Academy of Sciences, may shed light on fundamental cell survival processes involved with aging, cancer, and the generation of diversity in the immune system.

Exposure to ultraviolet light and other stresses, including certain oxidizing chemicals, is known to create mutations. In bacteria, such stresses induce a highly developed emergency defense mechanism to repair damaged DNA, which kicks in within an hour after exposure.

"The cell has an extraordinary ability to repair and restore damaged DNA," Myron F. Goodman, of USC, leader of the team, says. "So the mystery was, if these repair systems were so efficient, why did ultraviolet light do so much damage? Why did we see so many mutations?"

Prior research revealed that the DNA-replication mechanism became inaccurate only when a complex of unusual proteins—given the name umuC and umuD—were present. These genes are induced by a RecA dependent mechanism, termed the "SOS response."

"What everyone assumed," says Goodman, "was that these umu proteins somehow inhibited, or interfered with, the DNA copying enzymes and somehow lowered the copying fidelity of pol III, the enzyme thought to be doing the copying in SOS conditions."

But because of the evanescent nature of the umuC-D complex, it had been difficult to determine how that reduction in copying fidelity actually occurred.

In the latest experiment Goodman and his colleagues reconstituted this process in vitro, using a native UmuD'₂C complex, and found that the highly purified preparation contained DNA polymerase activity intrinsic to UmuD'₂C. This fifth E. coli DNA polymerase activity when combined with RecA, copies ultraviolet-damaged DNA very efficiently—about 100 times as efficient as pol III—and is highly "error prone," making mistakes 100 times more often.

"What this seems to be," says Goodman, "is a last-ditch cell defense. Faced with a choice between possible mutation and death, the cell chooses possible mutation."

If so, the evolutionary consequences are obvious. "Exposing a colony of bacteria to stress provides a mechanism that causes mutations that may produce organisms more suited to the changed environment," Goodman says.

Goodman says the discovery of an inaccurate DNA copying system immediately suggests a possible tie-in to one of the most mysterious aspects of the human immune system, "somatic hypermutability," a crucial part of the effectiveness of the immune system's B-cells. These cells produce immunoglobulins in a bewilderingly "hypervariable" variety of configurations.

This feature has been traced to an area, called the "hypervariable" region, on the B-cell's immunoglobin-producing genes; but the mechanism by which this hypervariability is achieved has thus far remained elusive, with the most recent explanation (a problem in a repair mechanism) recently disproved.

"But an inaccurate copier, like the one we found in E. coli, could produce precisely the effects we observe," says Goodman, predicting that experiments will soon test this hypothesis.

The pol V research was supported by grants from the National Institutes of Health. Besides Goodman, members of the research team were graduate students Xuan Shen and Mengjia Tang of the USC College of Letters, Arts and Sciences; Ekaterina G. Frank and Roger Woodgate of the NIH Section on DNA Replication, Repair, and Mutagenesis; and Mike O'Donnell of Rockefeller University and the Howard Hughes Medical Institute.

For more information: Myron Goodman, Department of Biological Sciences, University of Southern California, Stauffer Hall of Science, Los Angeles, CA 90033. Tel: 213-740-5190 Fax: 213-740-5191. Email:mgoodman@mizer.usc.edu.