OGT‐Mediated O‐GlcNAcylation Stabilizes c‐Myc Activity and Promotes Chemoresistance in Triple‐Negative Breast Cancer

The transcription factor c-Myc is often overexpressed in chemotherapy-resistant triple-negative breast cancer (TNBC). c-Myc function and stability are considered key factors regulating chemoresistance. Recent studies have revealed a potential link between the O-linked β-N-acetylglucosamine modification (O-GlcNAcylation) of c-Myc and its function and stability; however, the underlying mechanisms remain unexplored. This study aimed to investigate the role of O-GlcNAcylation in promoting chemoresistance and to explore the underlying mechanisms. A cisplatin (DDP)-resistant MDA-MB-231 cell line was established using a dose escalation. CCK-8, flow cytometry, and colony formation assays were used to evaluate cell resistance under different treatments. Western blotting and coimmunoprecipitation analyses were performed to evaluate the expression of c-Myc and its O-GlcNAcylation under different conditions. The possible O-GlcNAcylation sites were predicted using DictyOGlyc 1.1. Inhibition of O-linked N-acetylglucosamine transferase (OGT) significantly suppressed colony formation and promoted apoptosis of DDP-resistant cells. c-Myc expression was downregulated when OGT-mediated O-GlcNAcylation was inhibited. Additionally, OGT interacted with c-Myc, promoting its stability at the Thr58 residue. Mutation of Thr58 not only resulted in lower c-Myc stability, reduced colony formation ability, and increased apoptosis but also resulted in a decrease in both the total expression and O-GlcNAcylation of c-Myc. Therefore, O-GlcNAcylation at Thr-58 regulates c-Myc activity to promote chemoresistance of TNBC cells.