CTCF Expression and Dynamic Motif Accessibility Modulates Epithelial-Mesenchymal Gene Expression.

Abstract

Simple SummaryEpithelial–mesenchymal transition (EMT) facilitates cell migration, invasion, and, consequently, metastasis, which ultimately contributes to breast-cancer-related fatalities. In this study, we define the DNA accessibility dynamics that permit EMT and its reversal, MET. We demonstrate the progressive repression of E-cadherin, beginning with the loss of the membrane-bound fraction, and followed by the loss of CDH1 reporter expression. We identify that EMT is characterized by a global increase in accessible chromatin—nearly doubling the number of accessible regions. Furthermore, we find that regions exhibiting chromatin alterations are enriched in binding motifs for CTCF. Additionally, our data suggest that CTCF repression slows the loss of epithelial gene expression while accelerating the gain of mesenchymal gene expression, facilitating a state of partial EMT.Epithelial–mesenchymal transition (EMT) and its reversal, mesenchymal–epithelial transition (MET) drive tissue reorganization critical for early development. In carcinomas, processing through EMT, MET, or partial states promotes migration, invasion, dormancy, and metastatic colonization. As a reversible process, EMT is inherently regulated at epigenetic and epigenomic levels. To understand the epigenomic nature of reversible EMT and its partial states, we characterized chromatin accessibility dynamics, transcriptomic output, protein expression, and cellular phenotypes during stepwise reversible EMT. We find that the chromatin insulating protein machinery, including CTCF, is suppressed and re-expressed, coincident with broad alterations in chromatin accessibility, during EMT/MET, and is lower in triple-negative breast cancer cell lines with EMT features. Through an analysis of chromatin accessibility using ATAC-seq, we identify that early phases of EMT are characterized by enrichment for AP-1 family member binding motifs, but also by a diminished enrichment for CTCF binding motifs. Through a loss-of-function analysis, we demonstrate that the suppression of CTCF alters cellular plasticity, strengthening the epithelial phenotype via the upregulation of epithelial markers E-cadherin/CDH1 and downregulation of N-cadherin/CDH2. Conversely, the upregulation of CTCF leads to the upregulation of EMT gene expression and an increase in mesenchymal traits. These findings are indicative of a role of CTCF in regulating epithelial–mesenchymal plasticity and gene expression.