The role of the redox/miR-6855-3p/PRDX5A axis in reversing SLUG-mediated BRCA2 silencing in breast cancer cells

BACKGROUND: We have previously shown that the zinc finger transcription repressor SNAI2 (SLUG) represses tumor suppressor BRCA2-expression in non-dividing cells by binding to the E2-box upstream of the transcription start site. However, it is unclear how proliferating breast cancer (BC) cells that has higher oxidation state, overcome this repression. In this study, we provide insight into the mechanism of de-silencing of BRCA2 gene expression by PRDX5A, which is the longest member of the peroxiredoxin5 family, in proliferating breast cancer cells. METHODS: We used cell synchronization and DNA affinity pulldown to analyze PRDX5A binding to the BRCA2 silencer. We used oxidative stress and microRNA (miRNA) treatments to study nuclear localization of PRDX5A and its impact on BRCA2-expression. We validated our findings using mutational, reporter assay, and immunofluorescence analyses. RESULTS: Under oxidative stress, proliferating BC cells express PRDX5 isoform A (PRDX5A). In the nucleus, PRDX5A binds to the BRCA2 silencer near the E2-box, displacing SLUG and enhancing BRCA2-expression. Nuclear PRDX5A is translated from the second AUG codon in frame to the first AUG codon in the PRDX5A transcript that retains all exons. Mutation of the first AUG increases nuclear localization of PRDX5A in MDA-MB-231 cells, but mutation of the second AUG decreases it. Increased mitronic hsa-miRNA-6855-3p levels under oxidative stress renders translation from the second AUG preferable. Mutational analysis using reporter assay uncovered a miR-6855-3p binding site between the first and second AUG codon in the PRDX5A transcript. miR-6855-3p mimic increases accumulation of nuclear PRDX5A and inhibits reporter gene translation. CONCLUSION: Oxidative stress increases miR-6855-3p expression and binding to the inter-AUG sequence of the PRDX5A transcript, promoting translation of nuclear PRDX5A. Nuclear PRDX5A relieves SLUG-mediated BRCA2 silencing, resulting in increased BRCA2-expression. GRAPHICAL ABSTRACT: [Image: see text]