Poster number 20 submitted by Satenik Valesyan
LC-MS/MS-based detection of 5-methyl cytidine in E. coli tRNAs arising from oxidative stress
Satenik Valesyan (University of Cincinnati), Patrick A. Limbach (University of Cincinnati), Balasubrahmanyam Addepalli (University of Cincinnati)
Oxidative stress caused by endogenous and exogenous sources has been associated with a number of human diseases. One exogenous source is ultraviolet radiation (UVR). Our lab earlier found that post-transcriptional modifications in transfer ribonucleic acids (tRNAs) are damaged by the reactive oxygen species (ROS) generated by UVA. Those studies also revealed that some photoproducts were more readily identified during RNA modification mapping by liquid chromatography tandem mass spectrometry (LC-MS/MS) rather than relying solely on their detection during nucleoside analysis. Another approach for generating ROS is the Fenton reaction, whereby Fe2+ catalyzes the generation of hydroxyl radicals (HO.) from hydrogen peroxide (H2O2). Here we have been exposing Escherichia coli (E. coli) cells to ROS conditions generated by the Fenton reaction. Surprisingly, our preliminary nucleoside analysis results reveal that 5-methylcytidine (m5C) is found at significantly higher levels in tRNAs upon exposure to Fenton conditions. Because m5C has not been reported in any published E. coli tRNA sequences, we have focused our RNA modification mapping analyses on discovery possible sequence locations in specific tRNAs where this modification may be occurring. At present, Tyr-QUA appears to be the most likely tRNA containing this additional modification, which appears induced by the Fenton reaction. Results from our analyses along with a discussion into the possible causes for this change in modification status will be presented.
1. Liguori et al. (2018) Clin Interv Aging. 13: 757–772.
2. Bhattacharyya et al. (2014) Physiol Rev. 94(2): 329–354
3. Sun et al. (2018) ACS Chem Biol., 13(3): 567–572.
4. Zhao et al. (2019) Aging Med (Milton). 2(2): 82–87.
5. Dedon et al. (2010) PLoS Genet., 6(12): e1001247.
6. Sun et al. (2020) Biomolecules. 8;10(11):1527
7. Yu et al. (2017) ACS. 89, 20, 10744–10752
8. Lobue et al. (2019) Methods. 156:128-138
Keywords: Fenton reaction, tRNA, 5-methylcytidine (m5C)