Genome-Wide Study Based On Abasic Site Revealed Selective RNA Oxidation In Neuro 2a Cells. Mus musculus

To investigate selectivity of mRNA oxidation, total RNA and oxidized RNA isolated from Neuro 2a cells before and after H2O2 treatment were employed for microarray analysis. It was found that selective oxidation of mRNA already occurs under normal culture conditions but was increased by H2O2, especially in a subset of mRNAs related to certain functions. Moreover, mRNA oxidation level is also related to its abundance or stability (half-life time). This shows for the first time that mRNA oxidation is associated with RNA homeostasis including function, stability and abundance depending on cellular redox status in a genome-wide scale. Overall design: Neuro 2a cells received hydrogen peroxide treatment or no treatment as a control. Samples were applied for RNA extraction and ARP labeling, which could bind with apurinic/apyrimidinic sites, and then a pull-down process to isolate oxidized RNA. Total RNA and oxidized RNA were used for subsequent transcriptomic profiling. 4 types of samples were analyzed: Basal-total: untreated N2a cells labeled with ARP, but not processed for the pull-down assay. Ox-total: hydrogen peroxide-treated N2a cells labeled with ARP, but not processed for the pull-down assay. Basal-ARP: untreated N2a cells labeled with ARP, and processed for the pull-down assay. ARP-derivatized RNA, which is also oxidized RNA, was concentrated and used for the microarray analysis. Ox-ARP: hydrogen peroxide-treated N2a cells labeled with ARP, and processed for the pull-down assay. ARP-derivatized RNA, which is also oxidized RNA, was concentrated and used for the microarray analysis.