Modulation of NF-kappa-B Signalling

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Abstract

The NF-κB transcription factor p65/RelA controls hundreds of genes involved in inflammation, immunity, cell proliferation and survival. Cells treated with the inflammatory cytokine TNF-α show repeated nuclear-cytoplasmic translocations of p65 maintained by multiple negative feedback loops. Application of pulses of TNF-α at various set intervals can entrain the translocation frequency and cause different patterns of NF-κB-dependent gene transcription. Translocation frequency can also be altered by biological and environmental factors such as temperature and diclofenac treatment. These observations have led to the hypothesis that the dynamics of NF-κB play a role in the transcriptional responses dependent on particular cellular states. NF-κB can physically interact with members of other molecular networks including glucocorticoid signalling, the circadian clock and the cell cycle. In a previous study, two tumour cell lines showed differential p65 dynamics when treated with TNF-α at different cell cycle stages which concomitantly also altered cell cycle length. Evidence suggested that physical interactions between p65 and the cell cycle regulators E2F1 and E2F4 could, at least in part, be responsible for these effects. This thesis describes studies designed to investigate if similar responses exist in more normal cells. Although transcript levels of NF-κB target genes were suppressed in MEF cells treated with TNF-α during S phase relative to G1/S, NF-κB translocations appeared unaffected. Knowledge of the interactions with other molecular networks is central to understanding the context-dependent role of NF-κB, and the complexity of these dynamic interactions requires a Systems Biology approach. Within this study, a model of the interaction between NF-κB and the cell cycle was tested for its ability to recapitulate biological data from HeLa cells. Also, the effect of cell cycle timing of TNF-α treatment on NF-κB dynamics in HeLa cells was examined using ImageStream flow cytometry. It is suggested that a Systems Biology approach could be used to extend a study, initial results of which are presented here, of the interaction between NF-κB and glucocorticoid signalling. Understanding the interactions between NF-κB signalling and other networks should have clinical benefits, for example for cancer treatment dependent on circadian timing.

Keyword(s)

Cell cycle; Inflammation; Microscopy; Modelling; NF-kappa-B; Signalling

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