The genome-wide sequence specificity of DNA cleavage by bleomycin analogues in human cells

Bleomycin (BLM) is a cancer chemotherapeutic agent that cleaves cellular DNA at specific sequences. Using next-generation Illumina sequencing, the genome-wide sequence specificity of DNA cleavage by two BLM analogues, 6'-deoxy-BLM Z and zorbamycin (ZBM), was determined in human HeLa cells and compared with BLM. Over 200 million double-strand breaks were examined for each sample, and the 50,000 highest intensity cleavage sites were analysed. It was found that the DNA sequence specificity of the BLM analogues in human cells was different to BLM, especially at the cleavage site (position "0") and the "+1" position. In human cells, the 6'-deoxy-BLM Z had a preference for 5'-GTGY*MC (where * is the cleavage site, Y is C or T, M is A or C); it was 5'-GTGY*MCA for ZBM; and 5'-GTGT*AC for BLM. With cellular DNA, the highest ranked tetranucleotides were 5'-TGC*C and 5'-TGT*A for 6'-deoxy-BLM Z; 5'-TGC*C, 5'-TGT*A and 5'-TGC*A for ZBM; and 5'-TGT*A for BLM. In purified human genomic DNA, the DNA sequence preference was 5'-TGT*A for 6'-deoxy-BLM, 5'-RTGY*AYR (where R is G or A) for ZBM, and 5'-TGT*A for BLM. Thus, the sequence specificity of the BLM analogue, 6'-deoxy-BLM Z, was similar to BLM in purified human DNA, while ZBM was different.