title: Analysis of the TGFβ-induced program in primary airway epithelial cells shows essential role of NF-κB/RelA signaling network in type II epithelial mesenchymal transition.
abstract: BACKGROUND
  The airway epithelial cell plays a central role in coordinating the pulmonary response to injury and inflammation. Here, transforming growth factor-β (TGFβ) 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.
  
  
  RESULTS
  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-κB/RelA. Surprisingly, we also observed 527 of the EMT DEGs were also regulated by the TNF-NF-κB/RelA pathway. This Type II EMT program was compared to Type III EMT in TGFβ 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 TGFβ-induced the NF-κB/RelA pathway by observing a 2-fold change in NF-κB/RelA nuclear translocation. A small molecule IKK inhibitor blocked TGFβ-induced core transcription factor (SNAIL1, ZEB1 and Twist1) and mesenchymal gene (FN1 and VIM) expression.
  
  
  CONCLUSIONS
  These data indicate that NF-κB/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.
authors.affiliation: Department of Internal Medicine, University of Texas Medical Branch (UTMB), Galveston, TX, USA. bitian@utmb.edu.|Sealy Center for Molecular Medicine, UTMB, Galveston, TX, USA. bitian@utmb.edu.
authors.forename: Bing
authors.initials: B
authors.lastname: Tian
authors.affiliation: Institute for Translational Sciences, UTMB, Galveston, TX, USA. xuli@utmb.edu.|Department of Biochemistry and Molecular Biology, UTMB, Galveston, TX, USA. xuli@utmb.edu.
authors.forename: Xueling
authors.initials: X
authors.lastname: Li
authors.affiliation: Department of Internal Medicine, University of Texas Medical Branch (UTMB), Galveston, TX, USA. mrkalita@utmb.edu.
authors.forename: Mridul
authors.initials: M
authors.lastname: Kalita
authors.affiliation: Sealy Center for Molecular Medicine, UTMB, Galveston, TX, USA. sgwiden@utmb.edu.
authors.forename: Steven G
authors.initials: SG
authors.lastname: Widen
authors.affiliation: Department of Internal Medicine, University of Texas Medical Branch (UTMB), Galveston, TX, USA. junyang@utmb.edu.
authors.forename: Jun
authors.initials: J
authors.lastname: Yang
authors.affiliation: Department of Internal Medicine, University of Texas Medical Branch (UTMB), Galveston, TX, USA. subhavna@utmb.edu.|Sealy Center for Molecular Medicine, UTMB, Galveston, TX, USA. subhavna@utmb.edu.|Institute for Translational Sciences, UTMB, Galveston, TX, USA. subhavna@utmb.edu.
authors.forename: Suresh K
authors.initials: SK
authors.lastname: Bhavnani
authors.affiliation: Institute for Translational Sciences, UTMB, Galveston, TX, USA. brdang@utmb.edu.
authors.forename: Bryant
authors.initials: B
authors.lastname: Dang
authors.affiliation: Sealy Center for Molecular Medicine, UTMB, Galveston, TX, USA. askudlic@utmb.edu.|Institute for Translational Sciences, UTMB, Galveston, TX, USA. askudlic@utmb.edu.|Department of Biochemistry and Molecular Biology, UTMB, Galveston, TX, USA. askudlic@utmb.edu.
authors.forename: Andrzej
authors.initials: A
authors.lastname: Kudlicki
authors.affiliation: Sealy Center for Molecular Medicine, UTMB, Galveston, TX, USA. masinha@utmb.edu.|Department of Biochemistry and Molecular Biology, UTMB, Galveston, TX, USA. masinha@utmb.edu.|Bioinformatics Program, UTMB, Galveston, TX, USA. masinha@utmb.edu.
authors.forename: Mala
authors.initials: M
authors.lastname: Sinha
authors.affiliation: Sealy Center for Molecular Medicine, UTMB, Galveston, TX, USA. fakong@utmb.edu.|Department of Biochemistry and Molecular Biology, UTMB, Galveston, TX, USA. fakong@utmb.edu.|Bioinformatics Program, UTMB, Galveston, TX, USA. fakong@utmb.edu.
authors.forename: Fanping
authors.initials: F
authors.lastname: Kong
authors.affiliation: Sealy Center for Molecular Medicine, UTMB, Galveston, TX, USA. tgwood@utmb.edu.|Institute for Translational Sciences, UTMB, Galveston, TX, USA. tgwood@utmb.edu.|Department of Biochemistry and Molecular Biology, UTMB, Galveston, TX, USA. tgwood@utmb.edu.
authors.forename: Thomas G
authors.initials: TG
authors.lastname: Wood
authors.affiliation: Sealy Center for Molecular Medicine, UTMB, Galveston, TX, USA. baluxon@utmb.edu.|Institute for Translational Sciences, UTMB, Galveston, TX, USA. baluxon@utmb.edu.|Department of Biochemistry and Molecular Biology, UTMB, Galveston, TX, USA. baluxon@utmb.edu.|Bioinformatics Program, UTMB, Galveston, TX, USA. baluxon@utmb.edu.
authors.forename: Bruce A
authors.initials: BA
authors.lastname: Luxon
authors.affiliation: Department of Internal Medicine, University of Texas Medical Branch (UTMB), Galveston, TX, USA. arbrasie@utmb.edu.|Sealy Center for Molecular Medicine, UTMB, Galveston, TX, USA. arbrasie@utmb.edu.|Institute for Translational Sciences, UTMB, Galveston, TX, USA. arbrasie@utmb.edu.
authors.forename: Allan R
authors.initials: AR
authors.lastname: Brasier
mesh_terms: D004847:Epithelial Cells / Q000378:metabolism; D058750:Epithelial-Mesenchymal Transition / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics* / Q000473:pathology; D016328:NF-kappa B / Q000235:genetics; D015398:Signal Transduction / Q000235:genetics; D013234:Stem Cells / Q000378:metabolism; D051996:Transcription Factor RelA / Q000235:genetics* / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors / Q000235:genetics*
chemical_list: D016328:NF-kappa B; C498143:RELA protein, human; D051996:Transcription Factor RelA; D016212:Transforming Growth Factor beta