Keywords
p53
Epstein-Barr virus
CRISPR-based gene knockout
Transcriptome sequencing
Published : 2026-08-11
Abstract
Introduction: Nasopharyngeal carcinoma (NPC) is a head and neck cancer associated with Epstein-Barr virus (EBV) in Southern China and Southeast Asia. Although mutation of the p53 tumor suppressor gene is a rare event in NPC, NPC has a high frequency of over-expressed/accumulated p53 protein, which is considered to be a dysfunction or inactivation by EBV in NPC. However, the activity and role of p53 in the NPC remain controversial. Methods: The CRISPR-Cas9 gene editing system was used to establish NPC C666-1 cell lines with p53 knockout (KO), and transcriptome sequencing was performed to identify differentially expressed mRNAs and miRNAs in the p53 KO and control C666-1 cell lines. EdU incorporation assay, soft agar colony formation assay, and plate colony formation assay were performed to detect cell proliferation, and flow cytometry was used to analyze cell cycle and apoptosis. Results: 766 differentially expressed genes (DEGs) and 46 differentially expressed miRNAs were identified. The KEGG pathway enrichment analysis of both the DEGs and miRNAs target genes showed that the p53 signaling pathway was significantly enriched. In the enriched p53 signaling pathway, genes involved in cell cycle arrest and apoptosis induction were significantly downregulated by p53 KO, and re-expression of exogenous p53 rescued their expression in the C666-1 cells. Moreover, p53 KO promoted cell proliferation, decreased cell apoptosis, and accelerated G1/S phase progression in the C666-1 cells, and re-expression of exogenous p53 rescued these phenotypes in the C666-1 cells. Conclusions: Our data indicate that p53 retains tumor suppressor function in NPC C666-1 cells.
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