Computational analysis of dynamics within the Nuclear Factor-kappaB signalling system

The Nuclear Factor kappaB (NF-κB) proteins are a very important family of transcription factors. The signalling system of the NF-κB transcription factors drives cellular inflammation and immune responses, playing a central role in cell proliferation and apoptosis. NF-κB regulates the transcription of over 300 target genes including several negative feedback genes, such as the Inhibitory kappaBs (IκBs) and zinc-finger protein A20. Disruption to NF-κB signalling has been implicated in many autoimmune diseases and cancers. The ever growing list of diseases in which NF-κB is deregulated has made this one of the most intensely studied eukaryotic transcription factors. In response to continuous TNFα stimulation the system exhibits nuclear-cytoplasmic oscillations in the localisation of NF-κB with a robust period. Pulsatile stimulation causes nuclear translocations of NF-κB that are entrained to the pulse frequency. The altered frequency of these oscillations results in a different pattern of NF-κB dependent gene expression. This suggests the hypothesis that altering the frequency of NF-κB oscillations could provide a novel means to control the output of the system, allowing the exploitation of this system as a therapeutic target. The complex nonlinear dynamics in the NF-κB system make it an ideal candidate for a systems level analysis. Throughout this thesis a Systems Biology approach has been used to elucidate, both computationally and experimentally, how the period of NF-κB oscillations can be altered. This involved the development of a computational framework to characterise core network topology and parameter sensitivities of a recent deterministic model of the NF-κB system. Within this framework a model reduction algorithm has been described that has general applications to many other biochemical models. Bifurcation theory has been employed to characterise the system’s sensitivities. Computational analyses provided a basis to relate new experimental insights to our existing understanding of the system. Ectopic expression of the negative feedbacks IκBα and A20 showed different effects on the NF-κB oscillatory period. Computational analysis demonstrated that the manipulation of the delay in their transcriptional initiation could markedly change their influence on the oscillations. Single cell imaging also revealed a portion of A20 localised in perinuclear compartments. Two novel perturbations to the NF-κB dynamics are also considered: the effect after treatment with the non-steroidal anti-inflammatory drug (NSAID) diclofenac, and the effect of altered temperature. Theoretical analyses of these data lead to a number of hypotheses about how such perturbations influence the system. Ultimately, this thesis presents a number of new insights into modulating the period of NF-κB oscillations and computational tools to aid in the analysis of biochemical networks