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Preferential DNA Repair of an Active Gene in Human Cells
Isabel Mellon, Vilhelm A. Bohr, Charles Allen Smith and Philip C. Hanawalt
Proceedings of the National Academy of Sciences of the United States of America
Vol. 83, No. 23 (Dec. 1, 1986), pp. 8878-8882
Published by: National Academy of Sciences
Stable URL: http://www.jstor.org/stable/28850
Page Count: 5
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Removal of pyrimidine dimers was measured in defined sequences in human cells amplified for the dihydrofolate reductase (DHFR) gene. We quantitated repair in specific restriction fragments by using the dimer-specific bacteriophage T4 endonuclease V and analysis by Southern blotting. Within 4 hr after 5- or 10-J/m2 UV irradiation, more than 60% of the dimers had been removed from a 20-kilobase fragment that lies entirely within the transcription unit of the DHFR gene and from a 25-kilobase fragment located in the 5′ flanking region of the gene. Repair in the overall genome was measured by analyzing cellular DNA treated with T4 endonuclease V in alkaline sucrose gradients. Sixty-nine percent of the dimers were removed from the genome overall within 24 hr after irradiation, but only 25% were removed within 4 hr and 38% were removed within 8 hr. These results demonstrate a strong preferential rate of removal of dimers from the 50-kilobase region that includes the transcriptionally active DHFR gene compared to that in total cellular DNA. We confirmed that DHFR-containing DNA is repaired more rapidly than bulk DNA by using an approach that provides a direct comparison between repair in specific sequences and repair in total cellular DNA. We also show that the DHFR-containing sequences are repaired more rapidly than the nontranscribed repetitive α DNA sequences. Our finding of preferential early repair in a transcriptionally active region in overall repair-proficient cells suggests that selective dimer removal from active sequences may be a general characteristic of mammalian DNA repair.
Proceedings of the National Academy of Sciences of the United States of America © 1986 National Academy of Sciences