The Deubiquitinating Enzyme UCHL1 Promotes Radiotherapy Resistance in Glioma by Stabilizing Keratin 18

Maintaining stemness in glioma stem cells (GSCs), coupled with robust DNA damage repair capacity, is a key determinant of radioresistance in glioma. Although the deubiquitinating enzyme ubiquitin carboxy-terminal hydrolase L1 (UCHL1) has been implicated in regulating tumor cell functions and facilitating malignant progression, its specific role in sustaining GSC stemness and in promoting radioresistance remains poorly understood. In this study, we combined secondary analyses of public datasets with in–house experimental investigations. Specifically, we first analyzed UCHL1 expression and its prognostic significance in glioma using the GEO2R, GEPIA2, and CGGA databases. Radiosensitivity, DNA repair capability, and GSC stemness were evaluated using clonogenic survival, comet, sphere formation, extreme limiting dilution assay, and Western blot analyses. To identify UCHL1–interacting proteins, co-immunoprecipitation coupled with mass spectrometry was performed, followed by deubiquitination and rescue experiments to validate its substrates and functional dependence. Our results revealed that UCHL1 was markedly overexpressed in radioresistant glioblastoma cells and glioma tissues, and its expression levels positively correlated with poor prognosis in patients with glioma. UCHL1 knockdown attenuated the sphere-forming capacity and self–renewal frequency of GSCs and impaired the cellular response to DNA damage following irradiation, thereby attenuating the radioresistant phenotype. Mechanistically, UCHL1 stabilizes keratin 18 (KRT18) protein via deubiquitination. The effects of UCHL1 on radioresistance and stemness properties were largely diminished upon KRT18 knockdown, accompanied by reduced expression of the stemness marker SOX2. Furthermore, KRT18 knockdown attenuated the radioresistant phenotype induced by UCHL1 overexpression. Collectively, these findings suggest a mechanistic axis through which UCHL1 may regulate GSC stemness and radioresistance and identify UCHL1 as a candidate therapeutic target for reversing therapeutic resistance in glioma, warranting further validation.