The repair of DNA containing interstrand crosslinks induced by psoralen-plus-light in E. coli cells has been investigated. During a 30-minute incubation after psoralen-plus-light treatment, crosslinks were excised and the cellular DNA was cut into discrete pieces. The molecular weight of these pieces corresponds to about twice the single-strand distance between crosslinks, as measured by sedimentation velocity in alkaline sucrose. During further incubation, these DNA fragments were covalently joined into high molecular weight DNA. This joining did not occur in cells carrying a mutation at recA; in these strains the DNA was further degraded to smaller polynucleotides and acid-soluble material. The possibility that repair of crosslinked DNA involves strand exchanges between homologous duplexes was investigated. Cells were grown in '3C,16N-containing medium for several generations, then switched to medium of normal density that also contained [3Hjthymidine for about 0.5 generation.After the crosslinking treatment, the cells were incubated in medium of normal density in order for repair to occur. The DNA was extracted and centrifuged in alkaline CsCl density gradients, where the light and heavy strands were separated. Molecules of intermediate density that contained 3H accumulated during repair in wild-type cells, but not in control cells or treated recA-cells. After molecular weight reduction of the intermediate-density DNA, the IIH could be separated from the heavy strands, demonstrating that covalent joining between heavy and light strands of homologous duplexes accompanies repair. A mechanism involving sequential excision and genetic recombination is proposed for the repair of DNA containing interstrand crosslinks.Covalent linkages between the complementary strands of DNA in vitro, or in cells, are detected after treatment with several agents, including nitrous acid (1), nitrogen or sulfur mustards (2), mitomycin C (3), or psoralen-plus-light (4-6). Crosslinks are also formed in low yields after exposure to ultraviolet light (7), and ionizing radiation (8). Such interstrand crosslinks prevent the complementary strands from diffusing apart after exposure to denaturation conditions, and a crosslinked DNA duplex is therefore reversibly bihelical (1).Crosslinks that remain in cellular DNA should block the separation of complementary strands during semiconservative replication and, if left unrepaired, might prevent cells from dividing normally. Cells, however, survive treatments producing many crosslinks in their DNA, suggesting that DNA that contains crosslinks may.be successfully repaired. Animal cells growing in tissue culture maintain their ability to grow and divide after treatments with bifunctional alkylating agents that produce thousands of interstrand crosslinks per cell (9). Bacteria also retain their colony-forming ability after exposure to crosslinking agents. Wild-type Escherichia coli cells survive treatments producing about 55-70 crosslinks per genome, and strains defective in genetic recombina...

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