Suppression of Zeb2 gene expression by a small interference RNA (siRNA) and evaluation of its effect on biological characteristics of HL-60 human leukemia cells

Abstract

ZEB2 or SMAD interacting protein1 (SIP1) is a transcription factor of the Zinc fingers family that plays a crucial role in the epithelial-mesenchymal transition (EMT). EMT is a normal process, but its malignant malignant status initiates varieties of cancers. Although, various studies have shown that suppression of ZEB2 expression causes EMT inhibition, but ZEB2 pro-oncogenic activities to induce EMT and anti-apoptotic effects are largely uncharacterized. Herein, we seek to further investigate the regulatory role of ZEB2 in pro-oncogenic processes during EMT in acute promyelocytic leukemia (HL-60) cell line. Methods: HL-60 cells were transfected with the optimum concentration of specific ZEB2 siRNA. Relative expression levels of ZEB2, P53, SNAIL1, microRNA-200c, and microRNA-34a mRNA were measured using qRT-PCR. MTT assay was used to determine the cytotoxic effects of ZEB2 suppression and Flow cytometric assay using annexin V-FITC/propidium iodide (PI) staining was performed to evaluate the apoptosis rate of the cells. Results: It was investigated that ZEB2 siRNA effectively inhibited ZEB2 expression and had cytotoxic effects on HL-60 cells, which were in accordance with flow cytometric data. Knockdown of the ZEB2 gene caused a significant up-regulation in P53 gene expression and also increased expression of tumor-suppressing microRNAs including miR200c and miR34a. Interestingly, ZEB2 knockdown caused a significant down-regulation in the SNAIL1 gene expression.