An Immune Cell Activation Signature for Non - Small Cell Lung Cancer Revealed Tumor Microenvironment Heterogeneity and the Role of RORA in Regulating ZNF490/NDUFA12 Axis

INTRODUCTION: The development and progression of non-small cell lung cancer (NSCLC) are intricately linked to immune cell activation, but its related signature has not been reported. METHODS: This study combines in silico and in vitro approaches. TCGA-NSCLC and Gene Expression Omnibus (GEO) datasets were utilized to develop and validate a prognostic signature based on cell activation genes. The signature's validity was assessed through the identification of genomic, transcriptomic, tumor microenvironment (TME), and single-cell infiltration characteristics. The function of the candidate gene RORA was verified using CCK8, apoptosis, colony formation, wound healing, and transwell assays. The detailed mechanism of RORA was investigated through ChIP-PCR, luciferase assays, Western blot, and ROS detection. RESULTS: The prognostic signature was constructed from TCGA-NSCLC datasets and validated in six independent datasets (GSE30219, GSE33072, GSE37745, GSE41271, GSE42127, GSE50081). The signature was associated with LRP1B and RYR2 mutations, NSCLC-related pathways, drug response, and immune cell infiltration. The candidate gene RORA significantly inhibits the proliferation and migration abilities of NSCLC cell lines (A549 and NCI-H1299). Furthermore, the transcription factor RORA promotes ZNF490 expression, which subsequently inhibits NUDFs expression and oxidative phosphorylation (oxphos). DISCUSSION: The signature highlighted its significance with genomic features that were frequently reported as prognostic indicators (LRP1B and RYR2 mutations, cancer-related infiltration and pathway infiltration), and putative treatment response (IC50 in the TCGA dataset). Its detailed mechanism of candidate gene RORA revealed its role in oxphos, highlighting the crosstalk between metabolism and immune activation. CONCLUSION: The model is robust and effectively reflects NSCLC heterogeneity while predicting prognosis. RORA promotes the expression of ZNF490 to inhibit NUDFs and oxidative phosphorylation.