Bioinformatics Identification and Functional Validation of Mitochondrial Homeostasis-Related Genes in Nasopharyngeal Carcinoma Metastasis
Zhen Guo, BinSheng He, DanFeng CaoIntroduction:
Metastatic spread remains the leading cause of death in Nasopharyngeal Carcinoma (NPC); however, the role of mitochondrial homeostasis in this process remains largely unclear.
Objective:
This study aims to identify key genes associated with mitochondrial homeostasis and NPC metastasis, and to further explore their potential regulatory mechanisms, correlations with the immune microenvironment, and prognostic values.
Methods:
Focusing on metastasis- and mitochondrial homeostasis-related genes (MMHRGs), we sequentially conduct screening for hub genes, construct regulatory networks, predict drug binding, explore correlations with immunity and prognosis, and perform in vitro functional verification. Candidate hub genes were prioritized using LASSO regression combined with Random Forests. Putative druggable compounds targeting these genes were identified through the Drug Gene Interaction Database (DGIdb) and further examined by molecular docking. Immune infiltration and Tumor Immune Dysfunction and Exclusion (TIDE) analyses were performed to characterize the tumor immune microenvironment and predict immunotherapy response. A prognostic nomogram incorporating these genes was developed, and its predictive accuracy was assessed using time-dependent ROC curve analysis. Finally, the functional roles of the hub genes were experimentally validated in NPC cell lines using wound-healing assays and transmission electron microscopy.
Results:
A mitochondrial homeostasis-related gene signature, highlighted by four hub genes (FKBP4, ATN1, CDKAL1, GPR25), was identified in NPC metastasis. These genes were embedded within extensive post-transcriptional and transcriptional regulatory networks (ceRNA, RNA-binding proteins, TFs, PPIs) to deeply illustrate the molecular mechanism in NPC. Molecular docking revealed SAR260301 with high binding affinity to FKBP4, suggesting potential binding clues. Consensus clustering stratified NPC into two subtypes (Cluster_A/B), with Cluster_B showing enhanced immune checkpoint expression and TIDE-based features. A prognostic model incorporating HNMT, TAGLN3, CSF2RA, and LYZ demonstrated high predictive accuracy for NPC with AUC values of 0.854, 0.860, and 0.920 for 1-, 2-, and 3-year overall survival in the training cohort, and 0.778, 0.914, and 0.846 in the internal validation cohort. Crucially, functional validation showed that overexpression of GPR25 or knockdown of ATN1, CDKAL1, or FKBP4 significantly inhibited NPC cell migration and disrupted mitochondrial morphology.
Discussion:
The study demonstrated mitochondrial homeostasis-related genes- FKBP4, ATN1, CDKAL1, and GPR25 as candidate hub genes in NPC metastasis.
Conclusion:
This study identified four novel mitochondrial homeostasis-associated hub genes involved in NPC metastasis, established a mitochondria-based molecular subtyping scheme, and provided preliminary prognostic biomarkers and potential therapeutic targets for future mechanistic and translational investigations of metastatic NPC.