Elucidation of non-B DNA-induced mutagenesis mechanisms : DNA repair proteins are required for the processing of H-DNA and Z-DNA in eukaryotes

The vast majority of all cancers result from some form of genetic instability, thus it is important to study the mechanisms involved. The integrity of DNA can be influenced by secondary structure, and DNA can adopt alternative structures that do not conform to the Watson-Crick B-DNA helix (i.e. non-B DNA). To date, >12 different types of non-B DNA structures have been described including H-DNA and Z-DNA, and these structure-forming sequences are abundant in the human genome, occurring 1/3,000 and 1/50,000 base-pairs for Z-DNA and H-DNA, respectively. Non-B DNA can alter DNA metabolism and contribute to the development of many human diseases. Specific to this project, translocations occurring in the c-MYC and BCL-2 genes, which are shown to contain non-B DNA-forming sequences at translocation breakpoint “hotspots”, are characteristic of certain leukemias and lymphomas. However, the mechanism(s) involved in this process remains undefined. Previously, we found these structures to be mutagenic in bacteria, human cells, and mice, largely by stimulating the formation of DNA double strand breaks (DSBs). We speculated that the helical distortions produced by non-B DNA may be recognized as “damage” by the cell, eliciting an error-prone repair response, resulting in genomic instability. Using genetic-based mutation-reporter assays, we have shown for the first time that these structures are mutagenic in yeast. Furthermore, we have identified DNA repair proteins from both the nucleotide excision repair (NER) and mismatch repair (MMR) pathways to be involved in H-DNA and Z-DNA-induced mutagenesis via distinct mechanisms in both yeast and human cells. We further characterized the functions of these proteins using biochemical and molecular biology assays and found that they are enriched at sites of H-DNA and/or Z-DNA, and have cleavage activity at or near the structure. Taken together, these results suggest that non-B structures are processed in an error-prone fashion via various novel structure-specific repair pathways in which repair proteins from multiple pathways cooperate. The results obtained have enhanced our knowledge of DNA structure-induced genetic instability in disease etiology, and will guide future studies in the development of novel strategies to treat and/or prevent genetic diseases.