Date of Award
Bachelor of Science (B.S.) in Biology and University Honors
Escherichia coli -- Research, DNA repair, Psoralens, Chemotherapy
8-methoxypsoralen is a DNA-intercalating agent, which can photoreact with pyrimidine bases on opposing DNA strands, to form an interstrand crosslink. These lesions completely block replication and transcription, and are widely used in chemotherapies; yet how these lesions are processed in the cell remains poorly understood and insight into these processes could lead to better therapies that evade resistance. Previous studies isolated an Escherichia coli mutant demonstrating hyper-resistance to interstrand crosslink-inducing agents. The mutation was mapped to 57.2 minutes on the chromosome, and potentially encoded a 55-kDa protein induced as part of the SOS response. Although these genes remain unidentified, hscA and hscB map to this location, have a similar size, and are SOS-inducible. To determine if these or other genes might confer interstrand crosslink resistance in E. coli, we characterized how cells survived 8-methoxypsoralen-UVA treatment in the absence of HscAB, and when these gene products were overexpressed. In a second approach, we developed a selection system to isolate hyper-resistant strains through the sequential growth and exposure of wild-type cultures to 8-methoxypsoralen-UVA. We found no effect on cell survival in the hscAB mutant compared to its wild-type parent, suggesting that HscAB may not contribute to interstrand crosslink resistance as previously hypothesized. However, due to the significant cytotoxicity of plasmids containing hscAB even in the absence of 8-methoxypsoralen-UVA treatment, we could not determine whether overexpression of these gene products provided cellular protection. Using iteratively 8-methoxypsoralen-UVA treated cells, we isolated strains that were >104-fold more resistant to this interstrand crosslink-inducing agent compared to the parent strain. This result suggests that E. coli possess mechanisms of interstrand crosslink repair or tolerance and could serve as a model system for understanding the development of drug resistance in human cells.
Acott, Jedidiah D., "Interstrand Crosslink Resistance in Escherichia coli" (2018). University Honors Theses. Paper 577.