Abstract
BackgroundTo investigate the expression of cyclin-dependent kinase 8 (CDK8) and β-catenin in colon cancer and evaluate the role of CDK8 in the proliferation, apoptosis and cell cycle progression of colon cancer cells, especially in HCT116 cell line.MethodsColon cancer cell line HCT116 was transfected with small interfering RNA (siRNA) targeting on CDK8. After CDK8-siRNA transfection, mRNA and protein expression levels of CDK8 and β-catenin were determined by reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blot assay in HCT116 cells. Cell proliferation was measured by 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide Methylthiazolyl tetrazolium (MTT) assay, and cell cycle distribution and apoptosis were analyzed by flow cytometry analysis (FACS). CDK8 and β-catenin protein levels were also examined by real-time PCR and immunohistochemistry (IHC) in colon cancer tissues and adjacent normal tissues.ResultsAfter CDK8 specific siRNA transfection, mRNA and protein expression levels of CDK8 and β-catenin in HCT116 cells were noticeably decreased (P < 0.05). CDK8 specific siRNA transfection inhibited HCT116 cells' proliferation and facilitated their apoptosis significantly (P < 0.05). In addition, the proportion of HCT116 cells in the G0/G1 phase was remarkably increased after CDK8-siRNA transfection (P < 0.05). The expression levels of CDK8 and β-catenin in adjacent normal tissues were lower than in tumor tissues (P < 0.05). Moreover, the expression of CDK8 was correlated with the expression of β-catenin in both tumor and adjacent normal tissues (P < 0.05).ConclusionsCDK8 and β-catenin were expressed in colon cancer at a high frequency. CDK8 specific siRNA transfection down-regulated the expression of CDK8 in colon cancer cells, which was also associated with a decrease in the expression of β-catenin Moreover, CDK8 specific siRNA inhibited the proliferation of colon cancer cells, promoted their apoptosis and arrested these cells in the G0/G1 phase. Interference of CDK8 might be an effective strategy through β-catenin regulation of colon cancer.
Highlights
To investigate the expression of cyclin-dependent kinase 8 (CDK8) and b-catenin in colon cancer and evaluate the role of CDK8 in the proliferation, apoptosis and cell cycle progression of colon cancer cells, especially in HCT116 cell line
Effect of CDK8-small interfering RNA (siRNA) transfection on CDK8 and b-catenin expression in HCT116 cells Six hours after CDK8-siRNA transfection, the transfection efficiency was detected by FACS
The results showed that mRNA expression of CDK8 and b-catenin was markedly lower in the cateninCyclin-dependent kinase (CDK)-siRNA group compared with the other two groups (P < 0.01) (Figure 2)
Summary
To investigate the expression of cyclin-dependent kinase 8 (CDK8) and b-catenin in colon cancer and evaluate the role of CDK8 in the proliferation, apoptosis and cell cycle progression of colon cancer cells, especially in HCT116 cell line. It has been well known that mutations of key genes in the Wnt/b-catenin signaling pathway play an important role in the occurrence and development of colon cancer [1,2]. The mediator complex can bind to RNA polymerase II, which participates in eukaryotic gene transcription such as the transcription of the b-catenin signaling pathway. CDK8 plays an important regulatory role in cell cycle control and cell growth at the transcription level and it is proposed to be a proto-oncogene in human colon cancer [8,9,10]. We verified the mRNA and protein expression levels of CDK8 and b-catenin in colon cancer tissues
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