The human DEK proto-oncogene has been found to play an important role in autoimmune disease, viral infection and human carcinogenesis. Although it is transcriptionally up-regulated in cervical cancer, its intracellular function and regulation is still unexplored. In the present study, DEK and IκBα [inhibitor of NF-κB (nuclear factor κB) α] shRNAs (short hairpin RNAs) were constructed and transfected into CaSki cells using Lipofectamine™. The stable cell line CaSki–DEK was obtained after G418 selection. CaSki–IκB cells were observed at 48 h after psiRNA-IκB transfection. The inhibitory efficiency of shRNAs were detected by RT (reverse transcription)–PCR and Western blot analysis. The proliferation activity of cells were measured using an MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide] assay, cell apoptosis was measured using an Annexin V/PI (propidium iodide) kit, the cell cycle was analysed by flow cytometry and cell senescence was detected using senescence β-galactosidase staining. The intracellular expression of NF-κB p65 protein was studied by cytochemistry. The expression levels of NF-κB p65, p50, c-Rel, IκBα and phospho-IκBα protein were analysed by immunoblotting in whole-cell lysates, cytosolic fractions and nuclear extracts. The protein expression and activity of p38 and JNK (c-Jun N-terminal kinase) were also assayed. In addition, the NF-κB p65 DNA-binding activity was measured by ELISA. Following the silencing of DEK and IκBα, cell proliferation was inhibited, apoptosis was increased, the cell cycle was blocked in the G0/G1-phase with a corresponding decrease in the G2/M-phase, and cell senescence was induced. All of these effects may be related to the up-regulation of NF-κB p65 expression and its nuclear translocation.
Silencing of the DEK gene induces apoptosis and senescence in CaSki cervical carcinoma cells via the up-regulation of NF-κB p65
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Kuiran Liu, Tianda Feng, Jie Liu, Ming Zhong, Shulan Zhang; Silencing of the DEK gene induces apoptosis and senescence in CaSki cervical carcinoma cells via the up-regulation of NF-κB p65. Biosci Rep 1 June 2012; 32 (3): 323–332. doi: https://doi.org/10.1042/BSR20100141
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