URG4-silencing suppresses cell proliferation in nasopharyngeal carcinoma through induction of apoptosis.

Abstract

To investigate the expression and function of up-regulator of gene-4 (URG4) in nasopharyngeal carcinoma (NPC). Fresh NPC tumor tissue samples with paired adjacent normal nasopharyngeal tissues samples of 9 NPC patients were collected from NPC curative resection surgery. NPC cell lines (CNE1, CNE2 and HONE1) were cultured. Lentivirus-mediated URG4-specific short hairpin RNA (shRNA) stable transfection was done. The effect of URG4 on CNE4 and HONE1 cells viability was determined via 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT). The plate-colony-formation assay was performed. Apoptosis analysis was done by flow cytometry. The expression levels of protein and RNA were detected by Western blotting and quantitative polymerase chain reaction (qPCR). We determined the expression of URG4/URGCP in NPC tissues and cell lines using qPCR analysis and found it was significantly upregulated in NPC. After that, stable URG4-silencing NPC cells were constructed by transfection with lentivirus-mediated shRNA. Functionally analyses indicated that knockdown of URG4 significantly impaired cell viability and colony formation ability, as confirmed by MTT and colony formation assays. Furthermore, URG4-silencing NPC cells showed more cells in the stage of early and late apoptosis compared with controls by flow cytometry assay. Western blot analysis further confirmed that knockdown of URG4 enhanced the expression of cleaved caspase-3, cleaved PARP and Bax, while decreased the expression of Bcl-2 and survivin. URG4/URGCP might play an essential role in NPC cell growth and proliferation and its silencing might be as a potential therapeutic target for NPC.