The Cancer Genome Atlas (TCGA) network study of 12 cancer types (PanCancer 12) revealed frequent mutation of TP53, and amplification and expression of related TP63 isoform ΔNp63 in squamous cancers. Further, aberrant expression of inflammatory genes and TP53/p63/p73 targets were detected in the PanCancer 12 project, reminiscent of gene programs comodulated by cREL/ΔNp63/TAp73 transcription factors we uncovered in head and neck squamous cell carcinomas (HNSCCs). However, how inflammatory gene signatures and cREL/p63/p73 targets are comodulated genome wide is unclear. Here, we examined how the inflammatory factor tumor necrosis factor-α (TNF-α) broadly modulates redistribution of cREL with ΔNp63α/TAp73 complexes and signatures genome wide in the HNSCC model UM-SCC46 using chromatin immunoprecipitation sequencing (ChIP-seq). TNF-α enhanced genome-wide co-occupancy of cREL with ΔNp63α on TP53/p63 sites, while unexpectedly promoting redistribution of TAp73 from TP53 to activator protein-1 (AP-1) sites. cREL, ΔNp63α and TAp73 binding and oligomerization on NF-κB-, TP53- or AP-1-specific sequences were independently validated by ChIP-qPCR (quantitative PCR), oligonucleotide-binding assays and analytical ultracentrifugation. Function of the binding activity was confirmed using TP53-, AP-1- and NF-κB-specific REs or p21, SERPINE1 and IL-6 promoter luciferase reporter activities. Concurrently, TNF-α regulated a broad gene network with cobinding activities for cREL, ΔNp63α and TAp73 observed upon array profiling and reverse transcription–PCR. Overlapping target gene signatures were observed in squamous cancer subsets and in inflamed skin of transgenic mice overexpressing ΔNp63α. Furthermore, multiple target genes identified in this study were linked to TP63 and TP73 activity and increased gene expression in large squamous cancer samples from PanCancer 12 TCGA by CircleMap. PARADIGM inferred pathway analysis revealed the network connection of TP63 and NF-κB complexes through an AP-1 hub, further supporting our findings. Thus, inflammatory cytokine TNF-α mediates genome-wide redistribution of the cREL/p63/p73, and AP-1 interactome, to diminish TAp73 tumor suppressor function and reciprocally activate NF-κB and AP-1 gene programs implicated in malignancy.
Access optionsAccess options
Subscribe to Journal
Get full journal access for 1 year
only $51.94 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Rent or Buy article
Get time limited or full article access on ReadCube.
All prices are NET prices.
Cancer Genome Atlas N. Comprehensive genomic characterization of head and neck squamous cell carcinomas. Nature 2015; 517: 576–582.
Hoadley KA, Yau C, Wolf DM, Cherniack AD, Tamborero D, Ng S et al. Multiplatform analysis of 12 cancer types reveals molecular classification within and across tissues of origin. Cell 2014; 158: 929–944.
Soussi T, Wiman KG . Shaping genetic alterations in human cancer: the p53 mutation paradigm. Cancer Cell 2007; 12: 303–312.
Collavin L, Lunardi A, Del Sal G . p53-family proteins and their regulators: hubs and spokes in tumor suppression. Cell Death Differ 2010; 17: 901–911.
Leslie M . Brothers in arms against cancer. Science 2011; 331: 1551–1552.
Lu H, Yang X, Duggal P, Allen CT, Yan B, Cohen J et al. TNF-alpha promotes c-REL/DeltaNp63alpha interaction and TAp73 dissociation from key genes that mediate growth arrest and apoptosis in head and neck cancer. Cancer Res 2011; 71: 6867–6877.
Marcel V, Dichtel-Danjoy ML, Sagne C, Hafsi H, Ma D, Ortiz-Cuaran S et al. Biological functions of p53 isoforms through evolution: lessons from animal and cellular models. Cell Death Differ 2011; 18: 1815–1824.
Yang X, Lu H, Yan B, Romano RA, Bian Y, Friedman J et al. DeltaNp63 versatilely regulates a broad NF-kappaB gene program and promotes squamous epithelial proliferation, migration, and inflammation. Cancer Res 2011; 71: 3688–3700.
Deyoung MP, Ellisen LW . P63 and p73 in human cancer: defining the network. Oncogene 2007; 26: 5169–5183.
Perez CA, Pietenpol JA . Transcriptional programs regulated by p63 in normal epithelium and tumors. Cell Cycle 2007; 6: 246–254.
Seiwert TY, Zuo Z, Keck MK, Khattri A, Pedamallu CS, Stricker T et al. Integrative and comparative genomic analysis of HPV-positive and HPV-negative head and neck squamous cell carcinomas. Clin Cancer Res 2015; 21: 632–641.
Bunting K, Rao S, Hardy K, Woltring D, Denyer GS, Wang J et al. Genome-wide analysis of gene expression in T cells to identify targets of the NF-kappa B transcription factor c-Rel. J Immunol 2007; 178: 7097–7109.
Kastan MB, Zhan Q, el-Deiry WS, Carrier F, Jacks T, Walsh WV et al. A mammalian cell cycle checkpoint pathway utilizing p53 and GADD45 is defective in ataxia-telangiectasia. Cell 1992; 71: 587–597.
Dong G, Chen Z, Li ZY, Yeh NT, Bancroft CC, Van Waes C . Hepatocyte growth factor/scatter factor-induced activation of MEK and PI3K signal pathways contributes to expression of proangiogenic cytokines interleukin-8 and vascular endothelial growth factor in head and neck squamous cell carcinoma. Cancer Res 2001; 61: 5911–5918.
Dong G, Lee TL, Yeh NT, Geoghegan J, Van Waes C, Chen Z . Metastatic squamous cell carcinoma cells that overexpress c-Met exhibit enhanced angiogenesis factor expression, scattering and metastasis in response to hepatocyte growth factor. Oncogene 2004; 23: 6199–6208.
Andreasen PA . PAI-1—a potential therapeutic target in cancer. Curr Drug Targets 2007; 8: 1030–1041.
Boccaccio C, Sabatino G, Medico E, Girolami F, Follenzi A, Reato G et al. The MET oncogene drives a genetic programme linking cancer to haemostasis. Nature 2005; 434: 396–400.
Cromer A, Carles A, Millon R, Ganguli G, Chalmel F, Lemaire F et al. Identification of genes associated with tumorigenesis and metastatic potential of hypopharyngeal cancer by microarray analysis. Oncogene 2004; 23: 2484–2498.
O'Donnell RK, Kupferman M, Wei SJ, Singhal S, Weber R, O'Malley B et al. Gene expression signature predicts lymphatic metastasis in squamous cell carcinoma of the oral cavity. Oncogene 2005; 24: 1244–1251.
Rickman DS, Millon R, De Reynies A, Thomas E, Wasylyk C, Muller D et al. Prediction of future metastasis and molecular characterization of head and neck squamous-cell carcinoma based on transcriptome and genome analysis by microarrays. Oncogene 2008; 27: 6607–6622.
Romano RA, Ortt K, Birkaya B, Smalley K, Sinha S . An active role of the DeltaN isoform of p63 in regulating basal keratin genes K5 and K14 and directing epidermal cell fate. PLoS One 2009; 4: e5623.
Romano RA, Sinha S . Dynamic life of a skin keratinocyte: an intimate tryst with the master regulator p63. Indian J Exp Biol 2011; 49: 721–731.
Ng S, Collisson EA, Sokolov A, Goldstein T, Gonzalez-Perez A, Lopez-Bigas N et al. PARADIGM-SHIFT predicts the function of mutations in multiple cancers using pathway impact analysis. Bioinformatics 2012; 28: i640–i646.
Vaske CJ, Benz SC, Sanborn JZ, Earl D, Szeto C, Zhu J et al. Inference of patient-specific pathway activities from multi-dimensional cancer genomics data using PARADIGM. Bioinformatics 2010; 26: i237–i245.
Sethi G, Shanmugam MK, Ramachandran L, Kumar AP, Tergaonkar V . Multifaceted link between cancer and inflammation. Biosci Rep 2012; 32: 1–15.
Wong CK, Vaske CJ, Ng S, Sanborn JZ, Benz SC, Haussler D et al. The UCSC Interaction Browser: multidimensional data views in pathway context. Nucleic Acids Res 2013; 41: W218–W224.
Du J, Romano RA, Si H, Mattox A, Bian Y, Yang X et al. Epidermal overexpression of transgenic deltaNp63 promotes type 2 immune and myeloid inflammatory responses and hyperplasia via NF-kappaB activation. J Pathol 2014; 232: 356–368.
King KE, Ponnamperuma RM, Allen C, Lu H, Duggal P, Chen Z et al. The p53 homologue DeltaNp63alpha interacts with the nuclear factor-kappaB pathway to modulate epithelial cell growth. Cancer Res 2008; 68: 5122–5131.
Chatterjee A, Chang X, Sen T, Ravi R, Bedi A, Sidransky D . Regulation of p53 family member isoform DeltaNp63alpha by the nuclear factor-kappaB targeting kinase IkappaB kinase beta. Cancer Res 2010; 70: 1419–1429.
Rocco JW, Leong CO, Kuperwasser N, DeYoung MP, Ellisen LW . P63 mediates survival in squamous cell carcinoma by suppression of p73-dependent apoptosis. Cancer Cell 2006; 9: 45–56.
Leong CO, Vidnovic N, DeYoung MP, Sgroi D, Ellisen LW . The p63/p73 network mediates chemosensitivity to cisplatin in a biologically defined subset of primary breast cancers. J Clin Invest 2007; 117: 1370–1380.
Natan E, Joerger AC . Structure and kinetic stability of the p63 tetramerization domain. J Mol Biol 2012; 415: 503–513.
Belyi VA, Levine AJ . One billion years of p53/p63/p73 evolution. Proc Natl Acad Sci USA 2009; 106: 17609–17610.
Koeppel M, van Heeringen SJ, Kramer D, Smeenk L, Janssen-Megens E, Hartmann M et al. Crosstalk between c-Jun and TAp73alpha/beta contributes to the apoptosis-survival balance. Nucleic Acids Res 2011; 39: 6069–6085.
Rosenbluth JM, Mays DJ, Jiang A, Shyr Y, Pietenpol JA . Differential regulation of the p73 cistrome by mammalian target of rapamycin reveals transcriptional programs of mesenchymal differentiation and tumorigenesis. Proc Natl Acad Sci USA 2010; 108: 2076–2081.
Riley T, Sontag E, Chen P, Levine A . Transcriptional control of human p53-regulated genes. Nat Rev Mol Cell Biol 2008; 9: 402–412.
Smeenk L, van Heeringen SJ, Koeppel M, van Driel MA, Bartels SJ, Akkers RC et al. Characterization of genome-wide p53-binding sites upon stress response. Nucleic Acids Res 2008; 36: 3639–3654.
Yang A, Zhu Z, Kettenbach A, Kapranov P, McKeon F, Gingeras TR et al. Genome-wide mapping indicates that p73 and p63 co-occupy target sites and have similar dna-binding profiles in vivo. PLoS One 2010; 5: e11572.
Ortt K, Sinha S . Derivation of the consensus DNA-binding sequence for p63 reveals unique requirements that are distinct from p53. FEBS Lett 2006; 580: 4544–4550.
Koshiba S, Ichimiya S, Nagashima T, Tonooka A, Kubo T, Kikuchi T et al. Tonsillar crypt epithelium of palmoplantar pustulosis secretes interleukin-6 to support B-cell development via p63/p73 transcription factors. J Pathol 2008; 214: 75–84.
el-Deiry WS, Tokino T, Velculescu VE, Levy DB, Parsons R, Trent JM et al. WAF1, a potential mediator of p53 tumor suppression. Cell 1993; 75: 817–825.
Vayssade M, Haddada H, Faridoni-Laurens L, Tourpin S, Valent A, Benard J et al. P73 functionally replaces p53 in adriamycin-treated, p53-deficient breast cancer cells. Int J Cancer 2005; 116: 860–869.
Vikhanskaya F, Toh WH, Dulloo I, Wu Q, Boominathan L, Ng HH et al. P73 supports cellular growth through c-Jun-dependent AP-1 transactivation. Nat Cell Biol 2007; 9: 698–705.
Toh WH, Siddique MM, Boominathan L, Lin KW, Sabapathy K . c-Jun regulates the stability and activity of the p53 homologue, p73. J Biol Chem 2004; 279: 44713–44722.
Bancroft CC, Chen Z, Dong G, Sunwoo JB, Yeh N, Park C et al. Coexpression of proangiogenic factors IL-8 and VEGF by human head and neck squamous cell carcinoma involves coactivation by MEK-MAPK and IKK-NF-kappaB signal pathways. Clin Cancer Res 2001; 7: 435–442.
Shaulian E, Karin M . AP-1 as a regulator of cell life and death. Nat Cell Biol 2002; 4: E131–E136.
Ondrey FG, Dong G, Sunwoo J, Chen Z, Wolf JS, Crowl-Bancroft CV et al. Constitutive activation of transcription factors NF-(kappa)B, AP-1, and NF-IL6 in human head and neck squamous cell carcinoma cell lines that express pro-inflammatory and pro-angiogenic cytokines. Mol Carcinogen 1999; 26: 119–129.
Nottingham LK, Yan CH, Yang X, Si H, Coupar J, Bian Y et al. Aberrant IKKalpha and IKKbeta cooperatively activate NF-kappaB and induce EGFR/AP1 signaling to promote survival and migration of head and neck cancer. Oncogene 2014; 33: 1135–1147.
Barenco M, Brewer D, Papouli E, Tomescu D, Callard R, Stark J et al. Dissection of a complex transcriptional response using genome-wide transcriptional modelling. Mol Syst Biol 2009; 5: 327.
Bid HK, Roberts RD, Cam M, Audino A, Kurmasheva RT, Lin J et al. DeltaNp63 promotes pediatric neuroblastoma and osteosarcoma by regulating tumor angiogenesis. Cancer Res 2014; 74: 320–329.
Zhang Y, Liu T, Meyer CA, Eeckhoute J, Johnson DS, Bernstein BE et al. Model-based analysis of ChIP-Seq (MACS). Genome Biol 2008; 9: R137.
Jiang H, Wang F, Dyer NP, Wong WH . CisGenome Browser: a flexible tool for genomic data visualization. Bioinformatics 2010; 26: 1781–1782.
Shin H, Liu T, Manrai AK, Liu XS . CEAS: cis-regulatory element annotation system. Bioinformatics 2009; 25: 2605–2606.
Lawrence CE, Altschul SF, Boguski MS, Liu JS, Neuwald AF, Wootton JC . Detecting subtle sequence signals: a Gibbs sampling strategy for multiple alignment. Science 1993; 262: 208–214.
Machanick P, Bailey TL . MEME-ChIP: motif analysis of large DNA datasets. Bioinformatics 2011; 27: 1696–1697.
We thank Dr Fan Yang (NCI/NIH), Dr Kairong Cui (NHLBI/NIH), Dr Bingmei Zhu (NIDDK/NIH), Jeffery Burnett (NIDCD/NIH), Jamie Coupar (NIDCD/NIH), Guanmei Liang (Thomas Jefferson High School for Science and Technology, Alexandria, VA, USA) and Eric Nicolson (Ithaca High School, Ithaca, New York, NY, USA) for their technical assistance and suggestions. The authors express appreciation to Drs James W Rocco and Leif W Ellisen (Harvard University) for providing ΔNp63 and TAp63 expression vectors, Dr Thomas Gilmore (Boston University) for cRel expression plasmids, Professor Gerry Melino (University of Leicester) for TAp73α expression plasmids, Dr J Silvio Gutkind (NIDCR/NIH) for IL-6 promoter reporter plasmids, Dr Gourisankar Ghosh (UCSD) for the cRel expression plasmid, and Drs Michal Karin (University of California, San Diego), Cheng-Ming Chiang (University of Texas, Southwestern) and Xuan Liu (University of California, Riverside) for critique of and helpful suggestions for the manuscript. HL, HS, XY, AM, MJ, YB, CVW and ZC are supported by intramural projects ZIA-DC-000073, ZIA-DC-000074 and RAR and SS are supported by a grant from NIH (R01AR049238). JMS acknowledges support from NCI (R01-CA180778 and U24-CA143858), Stand Up to Cancer, Prostate Cancer Foundation and the Movember Foundation.
The authors declare no conflict of interest.
Supplementary Information accompanies this paper on the Oncogene website
About this article
Lymphotoxin-β receptor-NIK signaling induces alternative RELB/NF-κB2 activation to promote metastatic gene expression and cell migration in head and neck cancer
Molecular Carcinogenesis (2019)
LL202 ameliorates colitis against oxidative stress of macrophage by activation of the Nrf2/HO‐1 pathway
Journal of Cellular Physiology (2019)
Molecular Carcinogenesis (2019)
Cell Death & Disease (2019)