Non-small cell lung cancer (NSCLC) one of the most prevalent and severe malignancies globally and the molecular mechanisms of NSCLC are poor understood, limiting the development of diagnostic biomarkers and targeted therapies. Circular RNAs (circRNAs) have been identified as a sort of critical regulator in cancer progression. In this study, we identities the epigenetic regulation function of circular RNA circRHOT1 in promoting NSCLC cell proliferation. We found that circRHOT1 were elevated in the clinical tumor tissues relative to that in the peritumor tissues from NSCLC patients. circRHOT1 was up-regulated in human lung cancer cell lines compared with normal human lung epithelial cell line. MTT assays revealed that the silencing of circRHOT1 by siRNA suppressed cell viabilities of NSCLC cells. Colony formation and Edu assays confirmed that circRHOT1 knockdown attenuated NSCLC cell proliferation in vitro. Meanwhile, the depletion of circRHOT1 induced NSCLC cell apoptosis and cell cycle arrest in vitro. Mechanically, the depletion of circRHOT1 remarkably reduced c-MYC mRNA and protein expression in NSCLC cells. Inhibition of circRHOT1 reduced the enrichment of transcription active marker histone H3 lysine 27 acetylation (H3K27ac) and RNA polymerase II on the promoter of c-MYC. RNA pull down analysis showed that circRHOT1 was able to directly interact with acetyltransferase KAT5 in NSCLC cells. In summary, we concluded that circRHOT1 contributed to pathogenesis of NSCLC by epigenetically enhancing c-MYC expression through recruiting KAT5. CircRHOT1 and KAT5 may be used as the potential targets for NSCLC therapy.