Ectopic expression of TWIST1 upregulates the stemness marker OCT4 in the esophageal squamous cell carcinoma cell line KYSE30

Mohammad Hossein Izadpanah,Yasaman Fahim,Mohammad Mahdi Forghanifard,Mohammad Reza Abbaszadegan

doi:10.1186/s11658-017-0065-x

Abstract

BackgroundThe transcription factor TWIST1 plays an important role in the epithelial–mesenchymal transition (EMT) process and in the migration, invasion and metastasis of cancer cells. OCT4, which is a homeobox transcription factor, has an important role in the self-renewal potential of cancer cells. Our aim here is to elucidate impact of ectopic expression of TWIST1 on OCT4 gene expression in esophageal squamous cell carcinoma (ESCC).MethodsThe ESCC line was KYSE30. GP293T cells were transfected with purf-IRES-GFP and pGP plasmids to produce recombinant viral particles. A semi-confluent KYSE30 culture was transduced with the prepared retroviral particles. mRNA extraction and cDNA synthesis were performed from normal KYSE30 cells and those ectopically expressing TWIST1. Expressional analysis of TWIST1 and OCT4 were performed with relative comparative real-time PCR.ResultsEctopic expression of TWIST1 in KYSE30 cells was related to its significant overexpression: nearly nine-fold higher in GFP-hTWIST1 KYSE-30 cells than in control GFP cells. This induced expression of TWIST1 caused significant upregulation of OCT4 in GFP-hTWIST1 KYSE-30 cells: nearly eight-fold higher. In silico analysis predicted the correlation of TWIST1 and OCT4 through ETS2.ConclusionsOverexpressed TWIST1 can be correlated with upregulation of the cancer stem cell marker OCT4 and the protein may play critical regulatory role in OCT4 gene expression. Since OCT4 is involved in the self-renewal process, the results may suggest a new linkage between TWIST1 and OCT4 in the cell biology of ESCC, highlighting the probable role of TWIST1 in inducing self-renewal.

Highlights

Esophageal cancer (EC) is the eighth most common cancer and the sixth leading cause of cancer-related death worldwide [1]
This level of TWIST1 overexpression caused a significant increase in Octamer-binding transcription factor 4 (OCT4) gene expression: nearly eight-fold in Green fluorescent protein (GFP)-hTWIST1 KYSE30 cells compared to the control (Fig. 2a)
TWIST1 and OCT4 may be linked through the ETS protooncogene 2 (ETS2) gene Having checked the probable interactions of TWIST1 and OCT4, we found that ETS2 can act as an intermediate communicator for these markers (Fig. 3a)

Summary

Introduction

Esophageal cancer (EC) is the eighth most common cancer and the sixth leading cause of cancer-related death worldwide [1]. Esophageal squamous cell carcinoma (ESCC) is the sixth most common cancer among men and ninth among women worldwide. TWIST1 belongs to a class of transcription regulators with a basic helix-loop-helix (bHLH) DNA-binding domain. It identifies a hexanucleotide consensus sequence called E-box (CANNTG) in the promoter region of target genes [4]. The transcription factor TWIST1 plays an important role in the epithelial–mesenchymal transition (EMT) process and in the migration, invasion and metastasis of cancer cells. OCT4, which is a homeobox transcription factor, has an important role in the self-renewal potential of cancer cells. Our aim here is to elucidate impact of ectopic expression of TWIST1 on OCT4 gene expression in esophageal squamous cell carcinoma (ESCC)

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