Nitric Oxide Induced S-Nitrosation Causes Base Excision Repair Imbalance

Publication date: Available online 5 May 2018
Source:DNA Repair
Author(s): Marcus C. Parrish, Isaac A. Chaim, Zachary D. Nagel, Steven R. Tannenbaum, Leona D. Samson, Bevin P. Engelward
Nitric oxide, a reactive nitrogen species released by lymphocytes during inflammation, has been shown to react with DNA, proteins, and cells. Protein S-nitrosation, the process by which nitric oxide reacts with cysteine residues on proteins, has been found to modulate DNA repair. However, relatively little is known about the role of S-nitrosation in the context of repair of alkylation damage by the base excision repair pathway (BER). BER of DNA exposed to a methylating agent, such as methylmethane sulfonate (MMS), is initiated by the Alkyladenine DNA Glycosylase (AAG), which removes damaged bases. Here, we analyzed the effects of the transnitrosating peptide S-nitrosoglutathione (GSNO) on the repair of methylated bases. Through the use of the CometChip, a high-throughput version of the comet assay that measures DNA strand breaks, we observed an AAG-dependent increase in BER intermediates in GSNO-exposed mouse embryonic fibroblasts after MMS challenge. Through the use of the Fluorescence-based Multiplexed Host Cell Reactivation Assay (FM-HCR), a high-throughput assay used to measure DNA repair capacity, GSNO exposure was found to alter the activities of BER proteins and reduce overall BER capacity. Furthermore, cells exposed to both GSNO and MMS displayed reduced viability. Given that unrepaired BER intermediates are toxic, these results suggest that the altered activities of BER proteins following GSNO exposure induces an increase in BER intermediates that leads to increased cell death. Taken together, this study shows the detrimental effects of S-nitrosation on the BER pathway and reveals the deleterious impact of altered BER capacity under conditions of co-exposure to an alkylating agent.



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