Genetic effects of photoadducts and photocross-links in the DNA of phage lambda exposed to 360 nm light and tri-methylpsoralen or khellin.

The furocoumarin, 4,5',8-trimethylpsoralen, sensitizes cells and viruses to 360 nm light, producing cross-links and monoadducts in their DNA. The furanochromone khellin is a less effective sensitizing agent than psoralen, but has been found to induce cross-links and adducts in DNA also. The number of cross-links increases as the square of the time of exposure to light. We found that greater fluences were required for khellin than for psoralen, possibly because of the less favorable angle of the distal unsaturated bonds for corss-linking pyrimidines in adjacent base pairs. By adjusting the time of exposure to 360 nm light, lambda phages were damaged with [3H]psoralen and [3H]khellin so as to produce equal numbers of cross-links. These exposures were found to produce 8-times more [3H]khellin than [3H]psoralen adducts in the DNA of the phages. Similar exposures were made with nonradioactive photosensitizers to determine the effectiveness of lambda phages carrying cross-links and monoadducts in producing genetic recombinants. Lambda phage-prophage genetic corsses were performed with psoralen and khellin-damaged phages under repressed conditions in which replication of the damaged DNA was blocked. It was estimated from the results that cross-links were about 20-times more effective than monoadducts for inducing recombination under repressed conditions. In tests on the survival of plaque forming ability on wild type bacteria, it was estimated that cross-links were about 15-times more effective than the adducts. The results support the conclusion that, in homoimmune crosses with psoralen-damaged lambda phages infecting wild type lysogens, more than three-quarters of the induced recombination can be attributed to cross-links rather than to monoadducts.