GSE61220
Analysis Of The TGFb-Induced Program In Primary Airway Epithelial Cells Shows Essential Role Of NF-kB/RelA Signaling Network In Type II Epithelial Mesenchymal Transition
The airway epithelial cell plays a central role in coordinating pulmonary response to injury and inflammation.  Here, transforming growth factor-b (TGFb) activates gene expression programs to induce stem cell-like properties, inhibit expression of differentiated epithelial adhesion proteins and express mesenchymal contractile proteins.  This process is known as epithelial mesenchymal transition (EMT); although much is known about the role of EMT in cellular metastasis in an oncogene-transformed cell, less is known about Type II EMT, that occurring in normal epithelial cells.  In this study, we applied next generation sequencing (RNA-seq) in primary human airway epithelial cells to understand the gene program controlling Type II EMT and how cytokine-induced inflammation modifies it. Generalized linear modeling was performed on a two-factor RNA-seq experiment of 6 treatments of telomerase immortalized human small airway epithelial cells (3 replicates).  Using a stringent cut-off, we identified 3,478 differentially expressed genes (DEGs) in response to EMT.  Unbiased transcription factor enrichment analysis identified three clusters of EMT regulators, one including SMADs/TP63 and another NF-kB/RelA.  Surprisingly, we also observed 527 of the EMT DEGs were also regulated by the TNF-NF-kB/RelA pathway.  This Type II EMT program was compared to Type III EMT in TGFb stimulated A549 alveolar lung cancer cells, revealing significant functional differences. Moreover, we observe that Type II EMT modifies the outcome of the TNF program, reducing IFN signaling and enhancing integrin signaling.  We confirmed experimentally that TGFb-induced the NF-kB/RelA pathway by observing a 2-fold change in NF-kB/RelA nuclear translocation.  A small molecule IKK inhibitor blocked TGFb-induced core transcription factor (SNAIL1, ZEB1 and Twist1) and mesenchymal gene (FN1 and VIM) expression.  These data indicate that NF-kB/RelA controls a SMAD-independent gene network whose regulation is required for initiation of Type II EMT.  Type II EMT dramatically affects the induction and kinetics of TNF-dependent gene networks. Overall design: A human small airway epithelial cell line was treated with TGF-Beta to induce the epithelial to mesenchymal transition. TGF-Beta treated and untreated cells were further treated with TNF-alpha for 1 and 12 hours. Three replicates for each treatment and untreated controls were performed for a total of 18 samples.