Oxidative DNA Damage Exacerbates the Mutagenic Potential of Alternative DNA Structures via Altered DNA Repair Processing

Alternative DNA structure-forming (i.e., non-B) sequences such as H-DNA-forming sequences are enriched at chromosomal translocation hotspots in human cancer genomes, underscoring their role in genomic instability. H-DNA is particularly susceptible to DNA damage by reactive oxygen species (ROS), a common byproduct from both endogenous metabolism and environmental contaminants, thereby exacerbating its mutagenic potential. Oxidative lesions within B-DNA are efficiently processed by base excision repair (BER), whereas H-DNA is processed in a mutagenic fashion by nucleotide excision repair (NER). Thus, we speculate that the repair of oxidative lesions within H-DNA will promote aberrant BER and NER processing, ultimately enhancing mutagenesis. Here, we examine the processing of oxidative damage within H-DNA by measuring the changes in mutation frequencies and spectra, as well as the association with key NER and BER proteins in human cells in the presence or absence of specific DNA repair proteins. Our results demonstrate that oxidatively damaged H-DNA serves as a substrate for both BER and NER and reveals an interplay between BER and NER proteins, which influences mutation outcomes. This novel framework establishes a link between oxidative stress, DNA repair, and H-DNA-associated mutagenesis, providing insight into how environmentally relevant DNA damage can drive sequence-specific genomic instability at cancer-associated hotspots.