Objective: Plumbagin (PL) is a known quinoid (5-hydroxyl-2-methyl-1,4-napthoquinone) initially isolated from the roots of Plumbago zeylanica L. It is reported to exert anti-proliferative effects in oral squamous cell carci-noma (OSCC) cells, suggesting that PL would be a promising therapeutic anti-cancer drug for OSCC; however, the molecular basis by which PL suppresses cell survival signaling is poorly understood. In this study, we conducted a comprehensive gene expression analysis to identify the molecular basis by which PL suppresses cell survival signaling.Methods: Human OSCC cell lines HSC-3 and SAS were used in this study. Comprehensive gene expression analysis was performed with an Agilent Whole Human Genome DNA microarray 4 x 44 format. Gene Set Enrichment Analysis was conducted to investigate the effect of PL on the oncogenic signaling pathway in the OSCC cells. Results: cDNA microarray and subsequent gene set enrichment analyses showed that PL readily downregulates oxidative phosphorylation, MYC-target genes, and MTORC signaling at 4 h after treatment in HSC-3 and SAS cells. Semi-quantitative PCR analysis showed that gene expression levels of mitochondrial (Mt)-encoded COX-2, COX-3, and ND1 genes are significantly lower in the PL-treated OSCC cells than those in the untreated cells. In addition, an ROS scavenger, NAC, reversed the PL-induced downregulation of expression of Mt-encoded genes. Accordingly, intracellular ATP levels significantly decreased after PL treatment.Conclusions: Collectively, PL downregulates both oxidative phosphorylation and oncogenic signatures and de-creases the survival of OSCC cells, thus highlighting its application as a potent treatment for patients with OSCC.