CUGBP1, which is involved in posttranscriptional regulatory networks, may control cell growth, activation and differentiation. Meanwhile, CCAAT/enhancer-binding protein α (C/EBPα) acts as a basic leucine zipper transcription factor which controls differentiation-dependent gene expression and inhibits cell proliferation. To date, very little is known about the association between CUGBP 1 and C/EBPα in regulating cell proliferation and apoptosis in non-small cell lung cancer (NSCLC). CUGBP1 and C/EBPα mRNA expressions were analyzed in NSCLC tumor and adjacent normal tissues, and the relationship in clinicopathological parameters was evaluated. Knockdown of CUGBP1 and C/EBPα regulated by CUGBP1 in NSCLC cell line was identified by real-time PCR and Western blot. The effect of depletion of CUGBP1 was evaluated by MTT assay and Annexin/Propidium Iodide Apoptosis assay. CUGBP1 is highly expressed and expression of C/EBPα is low in NSCLC tissues. The correlation analysis revealed that there was negative correlation between the expression of CUGBP 1 and C/EBPα. Knockdown of CUGBP1 effectively silenced the expression of CUGBP1 and up-regulated C/EBPα. Also, suppression of CUGBP1 inhibits proliferation and induces apoptosis in A549 cells. These observations suggest that the first proof the overexpression of CUGBP1 in NSCLC contributes to tumorigenesis through down-regulation of C/EBPα. Knockdown of CUGBP1 or up-regulation C/EBPα might be a potential therapeutic approach for human non-small cell lung cancers.