MiR-204 Shifts the Epithelial to Mesenchymal Transition in Concert with the Transcription Factors RUNX2, ETS1, and cMYB in Prostate Cancer Cell Line Model

Krassimira Todorova,Kristiyan Kanev,Diana Zasheva,Soren Hayrabedyan

doi:10.1155/2014/840906

Abstract

Epithelial to mesenchymal transition is an essential step in advanced cancer development. Many master transcription factors shift their expression to drive this process, while noncoding RNAs families like miR-200 are found to restrict it. In this study we investigated how the tumor suppressor miR-204 and several transcription factors modulate main markers of mesenchymal transformation like E- and N-cadherin, SLUG, VEGF, and SOX-9 in prostate cancer cell line model (LNCaP, PC3, VCaP, and NCI-H660). We found that SLUG, E-cadherin, and N-cadherin are differentially modulated by miR-204, using miR-204 specific mimics and inhibitors and siRNA gene silencing (RUNX2, ETS-1, and cMYB). The genome perturbation associated TMPRSS2-ERG fusion coincided with shift from tumor-suppressor to tumor-promoting activity of this miRNA. The ability of miR-204 to suppress cancer cell viability and migration was lost in the fusion harboring cell lines. We found differential E-cadherin splicing corroborating to miR-204 modulatory effects. RUNX2, ETS1, and cMYB are involved in the regulation of E-cadherin, N-cadherin, and VEGFA expression. RUNX2 knockdown results in SOX9 downregulation, while ETS1 and cMYB silencing result in SOX9 upregulation in VCaP cells. Their expression was found to be also methylation dependent. Our study provides means for understanding cancer heterogeneity in regard to adapted therapeutic approaches development.

Highlights

Prostate cancer is one of the most common malignancies and the second leading cause of death from cancer in men
We investigated how miR-204 and the master transcription factors (TF) (SLUG, SOX9, RUNX2, ETS1, and cMYB) are implicated in epithelial to mesenchymal transition (EMT) (Ecadherin, N-cadherin) and in the expression of angiogenesis marker VEGFA, using cell lines model LNCaP [21], PC3 [22], VCaP [23], and NCI-H660 [24] of Androgen receptor (AR) sensitive and refractive prostate cancer cell lines, being either TMPRSS2-ERG fusion harboring or fusion-free
LNCaP, androgen receptor (AR) responsive (AR+), p53 enabled; PC3 (bone metastasis derived, p53 null and AR-unresponsive (AR−)); VCaP; and NCI-H660 prostate cancer cell lines were purchased from the ATCC (VA, US)

Summary

Introduction

Prostate cancer is one of the most common malignancies and the second leading cause of death from cancer in men. Cellular dedifferentiation and epithelial to mesenchymal transition (EMT), by contrast, are a hallmark of malignant transformation and metastatic disease In this process the reexpression of conserved developmental programs plays a key role [2]. ERG disrupts AR signalling by inhibiting AR expression, binding to and inhibiting AR activity at gene-specific loci, and inducing repressive epigenetic programs via direct activation of the EZH2 The latter causes an epigenetic silencing of developmental regulators and tumor suppressor genes, subverting cancer cells to a stem-cell-like epigenetic state [4]. During this process, part of which is EMT, a transformation of epithelial cells into the invasive and proliferating mesenchymal cells occurs. The main event that follows is the transcriptional shift, involving ETS1, SLUG (SNAI2), and other transcription factors (TF) that suppress the epithelial markers (e.g., E-cadherin and βcatenin) and activate mesenchymal ones

Methods

Results

Conclusion