ALOXE3 transcriptionally regulated by activating transcription factor 3 promotes HCC ferroptosis via ERK and JNK signaling pathway.

[BACKGROUND] Ferroptosis, a novel type of regulated cell death driven by iron-dependent lipid peroxide accumulation, represents a promising therapeutic strategy for aggressive cancers. However, the molecular mechanism of ferroptosis in hepatocellular carcinoma (HCC) remains elusive.

[METHODS] RNA sequencing (RNA-seq) identified Activating transcription factor 3 (ATF3) as a key regulator of ferroptosis susceptibility. CCK8 assays, flow cytometry, cell migration assays, colony formation assays, and nanotransmission electron microscopy were performed to investigate the effects of ATF3 in vitro, while subcutaneous xenograft models in nude mice were established to evaluate the biological roles of ATF3 in vivo. Chromatin immunoprecipitation and dual-luciferase assays were conducted to reveal the underlying mechanism of ATF3.

[RESULTS] RSL3-treated HCC cells exhibited characteristic ferroptotic features including elevated lipid peroxidation, mitochondrial shrinkage, and membrane condensation. Clinically, ATF3 expression was significantly higher in adjacent non-tumor tissues compared to HCC tissues and correlated with favorable prognosis, serving as an independent prognostic biomarker. Mechanistically, RSL3-induced ROS accumulation activates ERK/JNK signaling, which upregulates ATF3 expression. As a transcription factor, ATF3 directly binds the ALOXE3 promoter to drive its transcription and subsequent PUFA biosynthesis, thereby sensitizing cells to ferroptosis. Notably, ATF3-overexpressing tumor xenografts showed enhanced sensitivity to RSL3 and significantly enhanced sorafenib's antitumor efficacy.

[CONCLUSION] RSL3-induced ROS accumulation activated ERK/JNK signaling, upregulating downstream ATF3 expression. ATF3 transcriptionally activated ALOXE3, promoting PUFA synthesis to enhance ferroptosis susceptibility in HCC and overcome sorafenib resistance.