Effects Of Twist1 On Human Myeloid Leukemia

The Twist gene was originally identified as being required for mesoderm induction in Drosophila in 1987, a highly conserved transcription factor that belongs to the family of basic helix-loop-helix (bHLH) proteins. The numerous studies have reported the upregulation of Twistlin a variety of human cancers. Functional studies have indicated that Twist 1 may play a major role in tumour promotion and progression, by inhibiting differentiation, anti-apoptosis, favouring cell survival, and/or inducing an epithelial-mesenchymal-like transition. As an oncogene, Twist 1 has been a known candidate cancer gene therapeutic molecule, but the effects on leukemia remain elusive. In this study, we try to reveal the effect of overexpression of Twistl gene in K562 and U937 cell lines on myeloid leukemia.To explore the function of Twistl on myeloid leukemia, we constructed Twistl eukaryotic expression vector and transduced into K562 and U937 cells by using lentivirus infection method. A Real-time quantitive RT-PCR (RQ-RT-PCR) method and western blot were applied to assay the Twist gene expression levels. Then we assessed the effects of Twist gene in K562 and U937 on cell proliferation, colony-forming ability, cell cycle, apoptosis, and MDR by using growth curve analysis, colony forming assay, flow cytometry, Annexin—Ⅴ/PI and MTT assay. Results show that transfection with Twist gene significantly increased tumor cell survival and invasion, caused cell cycle increase in S stage (p<0.05) and induced the decrease of drug sensitivity.Mechanism investigation with gene-microarray showed that the expression of some cell cycle associated genes increased, such as CDKN1C,CDC14B,ANAPC5, and others decreased, such as CCNE2 and TFDP1 in K562 cells transfected with Twistl shRNA, as compared with K562 cells transfected with empty plasmid. K562 cells transfected Twistl shRNA were possibly arrested in G1 phase, and fiercely repressing the transformation to S phase. Also downregulation of Twistl represses the growth of tumor cells in human leukemia cellline K562. All these show that Twistl is necessary for tumor cells survival.Data from gene-microarray also showed the greatly increase of the expression FOXO3 and HIPK2 tumor suppressors which can facilitate the apoptosis of tumor cells effectively. Meanwhile autophagy associated genes like ATG12 decreased in K562 cells transfected with Twistl shRNA, which can promote drugs resistance and inhibit apoptosis of human cancers. In addition, the genes associated with invasion also changed to some extent, involving EMT, stem cell character and etc.Taken together, these data shows that Twistl take the important part in leukemogenesis and myeloid leukemia progression.