Cytokeratin 18 (CK18), one of the major components of intermediate filaments (IF) in simple epithelial cells, undergoes caspase-mediated cleavage upon epithelial cell necrosis and apoptosis. CK18 has been used as a biomarker of several cancers and has been reported to be dysregulated in cervical cancers. The effects of dysregulated expression of CK18 at a molecular level are, however, unclear. In the present study, the function of CK18 in HeLa cells, a cell line derived from a cervical cancer cells, was investigated using shRNA knockdown. Reduced levels of CK18 led to a significant decrease in cell apoptosis, compared with control cells. Notably, RNA-seq analysis of the transcriptomes of HeLa cells, with or without CK18 knockdown, revealed that genes in the NF-kappa B pathway, and certain apoptosis pathways, were under global transcriptional and alternative splicing regulation. Quantitative RT-PCR confirmed the CK18-regulated transcription of apoptotic genes FAS and FADD, as well as immune genes CXCL2 and CD79B, in addition to alternative splicing of FAS and CTNNB1. Western blot analysis further revealed that CK18 knockdown led to reduced expression of CASP8. In conclusion, the present study indicated that CK18 played a role in apoptosis, which may be mediated via a feed-back regulation loop and may involve regulation of transcription and alternative splicing of a number of genes in apoptotic pathways.