DTYMK Functions as a ceRNA to Promote Glioblastoma Progression and Mesenchymal Phenotype Through miR-7-5p/RAF1/MAPK Signaling

<title>Abstract</title> <p> <bold>Objective</bold> Glioblastoma (GBM) carries a dismal prognosis, and the molecular mechanisms governing its malignant progression remain poorly defined. DTYMK, encoding deoxythymidylate kinase, has been implicated as an oncogenic factor across multiple cancer types, yet its functional role and regulatory mechanism in GBM are unknown. This study aimed to characterize the oncogenic function of DTYMK in GBM, define its contribution to proliferation and mesenchymal (MES) transition, and elucidate the underlying molecular mechanism through which DTYMK exerts its effects. <bold>Methods</bold> A six-gene prognostic risk model integrating mitochondrial dysfunction, oxidative stress, and apoptosis-related gene signatures was constructed using LASSO regression and multivariate Cox regression analyses across the Atlas of Cancer Genome (TCGA) and the Chinese Glioma Genome Atlas (CGGA) cohorts. Single-cell transcriptomic profiling was performed to characterize the cellular localization and subtype specificity of DTYMK expression. DTYMK knockdown was conducted in glioma cell lines to evaluate effects on proliferation, apoptosis, invasion, MES transition, and intracellular reactive oxygen species (ROS) levels. In vivo validation employed both subcutaneous and orthotopic intracranial xenograft models. The competing endogenous RNA (ceRNA) mechanism was investigated through bioinformatic prediction, dual-luciferase reporter assays, and rescue experiments using epidermal growth factor (EGF) to reactivate MAPK signaling. <bold>Results</bold> The six-gene prognostic model exhibited robust predictive performance in both cohorts, with DTYMK contributing the highest risk coefficient. Single-cell analysis revealed that DTYMK was predominantly enriched in glioma tumor cells, with significant overrepresentation in MES-like subpopulations. DTYMK knockdown suppressed glioma cell proliferation, migration, and invasion, inhibited MES marker expression, and induced apoptosis accompanied by excessive intracellular ROS accumulation. In vivo, DTYMK depletion significantly reduced tumor burden and prolonged survival. Mechanistically, DTYMK functioned as a ceRNA by competitively sequestering miR-7-5p, thereby relieving translational repression of RAF1 and activating the downstream MEK-ERK signaling cascade. EGF-mediated reactivation of the MAPK pathway partially rescued the malignant phenotypes suppressed by DTYMK knockdown, confirming that MAPK activation is the principal effector mechanism of this regulatory axis. <bold>Conclusion</bold> This study identifies DTYMK as a key driver of GBM malignancy and MES transition operating through the DTYMK-miR-7-5p-RAF1-MAPK axis, and positions DTYMK as a potential prognostic biomarker and therapeutic target for precision intervention in glioma. </p>