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Evidence that TNF-TNFR1-TRADD-TRAF2-RIP-TAK1-IKK pathway mediates constitutive NF-κB activation and proliferation in human head and neck squamous cell carcinoma

Oncogene volume 26, pages 1385–1397 (2007)Cite this article

Abstract

Constitutively activated nuclear factor-κB (NF-κB) has been associated with a variety of aggressive tumor types, including head and neck squamous cell carcinoma (HNSCC); however, the mechanism of its activation is not fully understood. Therefore, we investigated the molecular pathway that mediates constitutive activation of NF-κB in a series of HNSCC cell lines. We confirmed that NF-κB was constitutively active in all HNSCC cell lines (FaDu, LICR-LON-HN5 and SCC4) examined as indicated by DNA binding, immunocytochemical localization of p65, by NF-κB-dependent reporter gene expression and its inhibition by dominant-negative (DN)-inhibitory subunit of NF-κB (IκBα), the natural inhibitor of NF-κB. Constitutive NF-κB activation in HNSCC was found to be due to constitutive activation of IκBα kinase (IKK); and this correlated with constitutive expression of phosphorylated forms of IκBα and p65 proteins. All HNSCC showed the expression of p50, p52, p100 and receptor-interacting protein; all linked with NF-κB activation. The expression of constitutively active NF-κB in HNSCC is mediated through the tumor necrosis factor (TNF) signaling pathway, as NF-κB reporter activity was inhibited by DN-TNF receptor-associated death domain (TRADD), DN-TNF receptor-associated factor (TRAF)2, DN-receptor-interacting protein (RIP), DN-transforming growth factor-β-activated kinase 1 (TAK1), DN-κ-Ras, DN-AKT and DN-IKK but not by DN-TRAF5 or DN-TRAF6. Constitutive NF-κB activation was also associated with the autocrine expression of TNF, TNF receptors and receptor-activator of NF-κB and its ligand in HNSCC cells but not interleukin (IL)-1β. All HNSCC cell lines expressed IL-6, a NF-κB-regulated gene product. Furthermore, treatment of HNSCC cells with anti-TNF antibody downregulated constitutively active NF-κB, and this was associated with inhibition of IL-6 expression and cell proliferation. Our results clearly demonstrate that constitutive activation of NF-κB is mediated through the TRADD-TRAF2-RIP-TAK1-IKK pathway, making TNF a novel target in the treatment of head and neck cancer.

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Abbreviations

ALLN:

N-acetyl-leu-leu-norleucinal

IκB:

inhibitory subunit of NF-κB

IKK:

IκBα kinase

JNK:

c-Jun N terminal kinase

MAPK:

mitogen-activated protein kinase

MEKK:

mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase

NF-κB:

nuclear factor-κB

PBS:

phosphate-buffered saline

TRAF:

TNF receptor-associated factor

RANK:

receptor activator of NF-κB

RANKL:

receptor activator of NF-κB ligand

TNF:

tumor necrosis factor

TRADD:

TNF receptor-associated death domain

RIP:

receptor-interacting protein

TAK1:

transforming growth factor-β-activated kinase 1

TAB1:

TAK1-binding protein

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Acknowledgements

We thank Ann Sutton for her critical review of this manuscript. Supported in part by the Clayton Foundation for Research (BBA), Department of Defense US Army Breast Cancer Research Program grant BC010610 (BBA), grant P01 CA91844 from the National Institutes of Health (NIH) on lung chemoprevention (BBA) and a P50 Head and Neck Specialized Programs of Research Excellence grant from the NIH (BBA).

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Author notes

  1. M M Chaturvedi: On leave from the Department of Zoology, University of Delhi, Delhi 110007, India

Authors and Affiliations

  1. Department of Experimental Therapeutics, The University of Texas MD Anderson Cancer Center, Houston, TX, USA

    D G Jackson-Bernitsas, H Ichikawa, Y Takada, B G Darnay, M M Chaturvedi & B B Aggarwal

  2. Department of Head and Neck Surgery, The University of Texas MD Anderson Cancer Center, Houston, TX, USA

    J N Myers

  3. Department of Molecular and Cellular Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX, USA

    X L Lin

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  1. D G Jackson-Bernitsas
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  2. H Ichikawa
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  8. B B Aggarwal
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Correspondence to B B Aggarwal.

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Jackson-Bernitsas, D., Ichikawa, H., Takada, Y. et al. Evidence that TNF-TNFR1-TRADD-TRAF2-RIP-TAK1-IKK pathway mediates constitutive NF-κB activation and proliferation in human head and neck squamous cell carcinoma. Oncogene 26, 1385–1397 (2007). https://doi.org/10.1038/sj.onc.1209945

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