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NFκB and HIF display synergistic behaviour during hypoxic inflammation

Cellular and Molecular Life Sciences volume 69, pages 1319–1329 (2012)Cite this article

Abstract

The oxygen-sensitive transcription factor hypoxia inducible factor (HIF) is a key regulator of gene expression during adaptation to hypoxia. Crucially, inflamed tissue often displays regions of prominent hypoxia. Recent studies have shown HIF signalling is intricately linked to that of the pro-inflammatory transcription factor nuclear factor kappa B (NFκB) during hypoxic inflammation. We describe the relative temporal contributions of each to hypoxia-induced inflammatory gene expression and investigate the level of crosstalk between the two pathways using a novel Gaussia princeps luciferase (Gluc) reporter system. Under the control of an active promoter, Gluc is expressed and secreted into the cell culture media, where it can be sampled and measured over time. Thus, Gluc constructs under the control of either HIF or NFκB were used to resolve their temporal transcriptional dynamics in response to hypoxia and to cytokine stimuli, respectively. We also investigated the interactions between HIF and NFκB activities using a construct containing the sequence from the promoter of the inflammatory gene cyclooxygenase 2 (COX-2), which includes functionally active binding sites for both HIF and NFκB. Finally, based on our experimental data, we constructed a mathematical model of the binding affinities of HIF and NFκB to their respective response elements to analyse transcriptional crosstalk. Taken together, these data reveal distinct temporal HIF and NFκB transcriptional activities in response to hypoxic inflammation. Furthermore, we demonstrate synergistic activity between these two transcription factors on the regulation of the COX-2 promoter, implicating a co-ordinated role for both HIF and NFκB in the expression of COX-2 in hypoxic inflammation.

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Abbreviations

COX-2:

Cyclooxygenase2

DMOG:

Dimethyl-oxaloylglycine

DMSO:

Dimethyl-sulfoxide

Gluc:

Gaussia Luciferase

HIF:

Hypoxia inducible factor

HRE:

Hypoxia response element

NFκB:

Nuclear factor kappa B

NRE:

NFκB response element

pGluc:

Plasmid encoding Gluc

PHD:

Prolyl hydroxylase

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Acknowledgments

We thank Drs. Catrióna Johnston and Jens Rauch (Systems Biology Ireland) for discussion. This work was supported by Science Foundation Ireland (Grant No. 06/CE/B1129).

Conflict of interest

None.

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Authors and Affiliations

  1. Conway Institute, University College Dublin, Dublin, 4, Ireland

    Ulrike Bruning, Susan F. Fitzpatrick & Cormac T. Taylor

  2. Systems Biology Ireland, University College Dublin, Dublin, 4, Ireland

    Susan F. Fitzpatrick, Till Frank, Marc Birtwistle, Cormac T. Taylor & Alex Cheong

  3. School of Physics, University College Dublin, Dublin, 4, Ireland

    Till Frank

Authors
  1. Ulrike Bruning
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  2. Susan F. Fitzpatrick
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  3. Till Frank
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  4. Marc Birtwistle
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  6. Alex Cheong
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Corresponding author

Correspondence to Alex Cheong.

Additional information

U. Bruning, S. F. Fitzpatrick, C. T. Taylor and A. Cheong contributed equally to this work.

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Bruning, U., Fitzpatrick, S.F., Frank, T. et al. NFκB and HIF display synergistic behaviour during hypoxic inflammation. Cell. Mol. Life Sci. 69, 1319–1329 (2012). https://doi.org/10.1007/s00018-011-0876-2

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  • DOI: https://doi.org/10.1007/s00018-011-0876-2

Keywords

  • NFκB
  • Hypoxia inducible factor
  • Inflammation
  • Transcription
  • Crosstalk
  • Mathematical modelling