Single-nucleotide-resolution genomic maps ofO6-methylguanine from the glioblastoma drug temozolomide

Abstract

ABSTRACTTemozolomide kills cancer cells by formingO6-methylguanine (O6-MeG), which leads to apoptosis due to mismatch-repair overload. However,O6-MeG repair byO6-methylguanine-DNA methyltransferase (MGMT) contributes to drug resistance. Characterizing genomic profiles ofO6-MeG could elucidate howO6-MeG accumulation is influenced by repair, but there are no methods to map genomic locations ofO6-MeG. Here, we developed an immunoprecipitation- and polymerase-stalling-based method, termedO6-MeG-seq, to locateO6-MeG across the whole genome at single-nucleotide resolution. We analyzedO6-MeG formation and repair with regards to sequence contexts and functional genomic regions in glioblastoma-derived cell lines and evaluated the impact of MGMT.O6-MeG signatures were highly similar to mutational signatures from patients previously treated with temozolomide. Furthermore, MGMT did not preferentially repairO6-MeG with respect to sequence context, chromatin state or gene expression level, however, may protect oncogenes from mutations. Finally, we found an MGMT-independent strand bias inO6-MeG accumulation in highly expressed genes, suggesting an additional transcription-associated contribution to its repair. These data provide high resolution insight on howO6-MeG formation and repair is impacted by genome structure and regulation. Further,O6-MeG-seq is expected to enable future studies of DNA modification signatures as diagnostic markers for addressing drug resistance and preventing secondary cancers.GRAPHICAL ABSTRACT