Single-nucleotide-resolution genomic maps of O6-methylguanine from the glioblastoma drug temozolomide

Abstract

Temozolomide kills cancer cells by forming O6-methylguanine (O6-MeG), which leads to apoptosis due to mismatch-repair overload. However, O6-MeG repair by O6-methylguanine-DNA methyltransferase (MGMT) contributes to drug resistance. Characterizing genomic profiles of O6-MeG could elucidate how O6-MeG accumulation is influenced by repair, but there are no methods to map genomic locations of O6-MeG. Here, we developed an immunoprecipitation- and polymerase-stalling-based method, termed O6-MeG-seq, to locate O6-MeG across the whole genome at single-nucleotide resolution. We analyzed O6-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 repair O6-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 in O6-MeG accumulation in highly expressed genes, suggesting an additional transcription-associated contribution to its repair. These data provide high resolution insight on how O6-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 O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/571283v2_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@357353org.highwire.dtl.DTLVardef@12688b9org.highwire.dtl.DTLVardef@da3381org.highwire.dtl.DTLVardef@16b7e7e_HPS_FORMAT_FIGEXP M_FIG C_FIG