TRAF3 loss protects glioblastoma cells from lipid peroxidation and immune elimination via dysregulated lipid metabolism
Yu Zeng, … , Ye Song, Aidong Zhou
Yu Zeng, … , Ye Song, Aidong Zhou
Published February 11, 2025
Citation Information: J Clin Invest. 2025;135(7):e178550. https://doi.org/10.1172/JCI178550.
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Research Article Cell biology Metabolism

TRAF3 loss protects glioblastoma cells from lipid peroxidation and immune elimination via dysregulated lipid metabolism

Abstract

Glioblastoma (GBM) is a highly aggressive form of brain tumor characterized by dysregulated metabolism. Increased fatty acid oxidation (FAO) protects tumor cells from lipid peroxidation–induced cell death, although the precise mechanisms involved remain unclear. Here, we report that loss of TNF receptor–associated factor 3 (TRAF3) in GBM critically regulated lipid peroxidation and tumorigenesis by controlling the oxidation of polyunsaturated fatty acids (PUFAs). TRAF3 was frequently repressed in GBM due to promoter hypermethylation. TRAF3 interacted with enoyl-CoA hydratase 1 (ECH1), an enzyme that catalyzes the isomerization of unsaturated FAs (UFAs) and mediates K63-linked ubiquitination of ECH1 at Lys214. ECH1 ubiquitination impeded TOMM20-dependent mitochondrial translocation of ECH1, which otherwise promoted the oxidation of UFAs, preferentially the PUFAs, and limited lipid peroxidation. Overexpression of TRAF3 enhanced the sensitivity of GBM to ferroptosis and anti–programmed death–ligand 1 (anti–PD-L1) immunotherapy in mice. Thus, the TRAF3/ECH1 axis played a key role in the metabolism of PUFAs and was crucial for lipid peroxidation damage and immune elimination in GBM.

Authors

Yu Zeng, Liqian Zhao, Kunlin Zeng, Ziling Zhan, Zhengming Zhan, Shangbiao Li, Hongchao Zhan, Peng Chai, Cheng Xie, Shengfeng Ding, Yuxin Xie, Li Wang, Cuiying Li, Xiaoxia Chen, Daogang Guan, Enguang Bi, Jianyou Liao, Fan Deng, Xiaochun Bai, Ye Song, Aidong Zhou

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Figure 2

TRAF3 promotes ROS-induced mitochondrial damage and inhibits GBM tumorigenesis.

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TRAF3 promotes ROS-induced mitochondrial damage and inhibits GBM tumorig...
(A) GSEA was performed to discern gene expression changes following TRAF3 overexpression (oeTRAF3) in GBM0709 cells. (B) Oxidative stress–related gene heatmap in oeTRAF3 GBM0709 cells. (C) qRT-qPCR of selected genes in oeTRAF3 cells, normalized to GAPDH (n = 3). (D and E) Flow cytometry of cellular (CellROX) and mitochondrial ROS (MitoSOX) median fluorescence intensity (MFI) in oeTRAF3 cells (n = 3). (F) GSH/GSSG ratio in oeTRAF3 cells (n = 3). (G) JC-1 staining of oeTRAF3 cells. Scale bar: 100 μm. The monomer/dimer ratios of JC-1 were statistically analyzed (n = 6). (H) TEM images of mitochondria in oeTRAF3 cells. Scale bar: 500 nm (original magnification, ×40,000; enlarged magnification, ×80,000). The proportion of damaged mitochondria was statistically analyzed (n = 6). (I) SA-β-gal+ cell percentages in oeTRAF3 cells treated or not with GSH-EE (n = 6). (J) Cell viability of TRAF3-OE GBM0709 or GBM0108 cells treated or not with GSH-EE by cell counting kit-8 (CCK8). Absorbance values were normalized to the control (n = 4). (K) GBM0709 or GBM0108 cells (5 × 105 cells/mouse) stably expressing TRAF3 were intracranially (i.c.) injected into nude mice, and tumor growth was monitored by MRI. Tumor volumes in each group were statistically analyzed (n = 5). (L) The survival of mice bearing GBM0709 and GBM0108 GBM tumors was evaluated (n = 5). (M) Consecutive mouse GBM tissues derived from GBM0709 cells were stained for TRAF3, Ki-67, 8-oxoG, and β-gal, respectively. Scale bars: 100 μm. Staining scores were compared (n = 5). Statistical significance was determined using an unpaired, 2-tailed Student’s t test (C–H, K, and M), 1-way ANOVA with Tukey’s post hoc test (I and J), or a log-rank test (L). Ctrl, control. Repeated data are presented as the mean ± SD. *P < 0.05, **P < 0.01, and ***P < 0.001.