NSUN3 promotes oral squamous cell carcinoma progression through autophagy activation and FOXO pathway modulation

Background: C) modification, mediated by methyltransferases such as NSUN3, is implicated in tumor progression; however, the specific function of NSUN3 in OSCC remains largely unexplored. Autophagy plays a dual role in cancer, and the FOXO pathway is a key regulator of autophagy. This study aimed to elucidate the function of NSUN3 in OSCC and its potential mechanism involving autophagy and FOXO signaling. Methods: NSUN3 expression profiles were characterized in OSCC tissues and cell lines using immunohistochemistry, qRT-PCR, and Western blot. Clinicopathological correlations and survival analyses were performed on a cohort of 60 OSCC patients. NSUN3 was knocked down in SCC15 and SCC25 cells using lentiviral shRNA. Cell proliferation, migration, and invasion were assessed by CCK-8, wound-healing, and Transwell assays. Autophagy activity was evaluated by immunofluorescence (LC3 puncta formation), electron microscopy (autophagosome quantification), and Western blot (LC3-II/I ratio, Beclin1, P62/SQSTM1). The autophagy agonist rapamycin was used to rescue phenotypic changes. The activity of the FOXO pathway was assessed by detecting phospho-FOXO1/FOXO3. Results: NSUN3 was significantly upregulated in OSCC tissues and cells. Elevated NSUN3 expression, along with advanced pTNM stage and lymph node metastasis, constituted independent risk factors for poor overall survival. NSUN3 knockdown suppressed OSCC cell proliferation, migration, and invasion. Mechanistically, NSUN3 depletion inhibited autophagy, as evidenced by reduced LC3 puncta, decreased autophagosome number, lower Beclin1 expression, a reduced LC3-II/I ratio, and increased P62/SQSTM1 levels. Rapamycin treatment reversed these effects and partially restored malignant phenotypes. Furthermore, NSUN3 knockdown increased the phosphorylation (inactivation) of FOXO1 and FOXO3, thereby suppressing the FOXO signaling pathway. Conclusion: NSUN3 is overexpressed in OSCC and is an independent prognostic factor. It promotes OSCC progression by enhancing autophagy, potentially through modulating the FOXO pathway. Targeting NSUN3 may represent a novel therapeutic strategy for OSCC.