NF-κB regulates mesenchymal transition for the induction of non-small cell lung cancer initiating cells.
The epithelial-to-mesenchymal transition (EMT) is a de-differentiation process that has been implicated in metastasis and the generation of cancer initiating cells (CICs) in solid tumors. To examine EMT in non-small cell lung cancer (NSCLC), we utilized a three dimensional (3D) cell culture system in which cells were co-stimulated with tumor necrosis factor alpha (TNF) and transforming growth factor beta (TGFβ). NSCLC spheroid cultures display elevated expression of EMT master-switch transcription factors, TWIST1, SNAI1/Snail1, SNAI2/Slug and ZEB2/Sip1, and are highly invasive. Mesenchymal NSCLC cultures show CIC characteristics, displaying elevated expression of transcription factors KLF4, SOX2, POU5F1/Oct4, MYCN, and KIT. As a result, these putative CIC display a cancer "stem-like" phenotype by forming lung metastases under limiting cell dilution. The pleiotropic transcription factor, NF-κB, has been implicated in EMT and metastasis. Thus, we set out to develop a NSCLC model to further characterize the role of NF-κB activation in the development of CICs. Here, we demonstrate that induction of EMT in 3D cultures results in constitutive NF-κB activity. Furthermore, inhibition of NF-κB resulted in the loss of TWIST1, SNAI2, and ZEB2 induction, and a failure of cells to invade and metastasize. Our work indicates that NF-κB is required for NSCLC metastasis, in part, by transcriptionally upregulating master-switch transcription factors required for EMT.
Department of Biochemistry & Molecular Genetics, University of Virginia, Charlottesville, Virginia, USA.
Manish
M
Kumar
David F
DF
Allison
Natalya N
NN
Baranova
J Jacob
JJ
Wamsley
Adam J
AJ
Katz
Stefan
S
Bekiranov
David R
DR
Jones
Marty W
MW
Mayo
D000818:Animals; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D018929:Cell Culture Techniques; D045744:Cell Line, Tumor; D002471:Cell Transformation, Neoplastic / Q000235:genetics / Q000378:metabolism*; D004195:Disease Models, Animal; D058750:Epithelial-Mesenchymal Transition / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D000090062:Kruppel-Like Factor 4; D008175:Lung Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D009362:Neoplasm Metastasis; D014411:Neoplastic Stem Cells / Q000378:metabolism; D010641:Phenotype; D018874:Spheroids, Cellular; D014407:Tumor Cells, Cultured
C000715467:KLF4 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D016328:NF-kappa B