USP49 inhibits ferroptosis via activation of the SHCBP1-β-catenin-GPX4 pathway, promoting the malignant progression of adenocarcinoma of the esophagogastric junction

Background: Adenocarcinoma of the esophagogastric junction (AEG) has received widespread attention because of its increasing incidence. However, the molecular mechanism underlying tumor progression remains unclear. Methods: Malondialdehyde assay, lipid peroxidation and Transmission electron microscopy were used to evaluate ferroptosis. Cell proliferation was determined using CCK-8, EdU assay and cell colony formation assays. A xenograft animal model in vivo was established by subcutaneous injection of stable cell lines into the flank of nude mice. Western blotting and Immunohistochemistry were performed to evaluate protein expression in cells and patient tissues, respectively. Additionally, the interaction of Ubiquitin-specific peptidase 49 (USP49) and Shc SH2-domain binding protein 1 (SHCBP1) was detected by immunofluorescence(IF), coimmunoprecipitation (Co-IP), cycloheximide and ubiquitination assays. was used to determine USP49 and GPX4 expression in. Results: Here, we report that the downregulation of USP49 promotes ferroptosis in OE33 and OE19 cells, thereby inhibiting cell proliferation in vitro and in vivo, whereas the overexpression of USP49 had the opposite effect. In addition, USP49 downregulation promoted radiotherapy sensitivity in OE33 and OE19 cells. Moreover, the downregulation of USP49 reduced the mRNA and protein levels of glutathione peroxidase 4 (GPX4), and the overexpression of GPX4 rescued the malignant biological behavior induced by USP49 knockdown. Mechanistically, USP49 deubiquitinates and stabilizes Shc SH2-domain binding protein 1 (SHCBP1), subsequently facilitating the entry of β-catenin into the nucleus to enhance GPX4 transcriptional expression. Finally, high USP49 expression was correlated with shorter overall survival in patients with AEG. Conclusions: In summary, our findings identify USP49 as a novel regulator of ferroptosis in AEG cells, indicating that USP49 may be a potential therapeutic target in AEG.