Single-cell time-lapse imaging of the dynamic control of NF-κB signalling

The transcription factor NF-κB (nuclear factor κB) regulates critical cellular processes including the inflammatory response, apoptosis and the cell cycle. Over the past 20 years many of the components of the NF-κB signalling pathway have been elucidated along with their functions. Recent research in this field has focused on the dynamic regulation and network control of this system. With key roles in so many important cellular processes, it is critical that NF-κB signalling is tightly regulated. Recently, single-cell imaging and mathematical modelling have identified that the timing of cellular responses may play an important role in the regulation of this pathway. p65/RelA (RelA) has been shown to translocate between the nucleus and cytoplasm with varying oscillatory patterns in different cell lines leading to differences in transcriptional outputs from NF-κB-regulated genes. Variations in the timing or persistence of these movements may control the maintenance and differential expression of NF-κB-regulated genes. Introduction to the NF-κB (nuclear factor κB) network The NF-κB signalling pathway has been shown to be active in virtually all cell types and to regulate a number of essential cellular functions. These include inflammation, apoptosis and the cell cycle. NF-κB is critical in the initiation of inflammation [1] (e.g. by induction of pro-inflammatory cytokine transcription), the perpetuation of inflammation (through positive feedback loops) and termination of the inflammatory response (often by inducing apoptosis) [2]. NF-κB proteins include RelA/p65, RelB, c-Rel (the product of the cellular homologue of the avian-reticuloendotheliosis-virus transforming gene), p100/p52 and p105/p50. The active transcription factor is a homodimer or heterodimer, although some complexes (e.g. p50–p50) have repressor functions. IκB (inhibitory κB) proteins hold NF-κB transcription factors in the cytoplasm by masking the nuclear localization sequence. Upon activation, the IκB proteins are phosphorylated, leading to their subsequent ubiquitination and degradation allowing NF-κB to translocate to the cell nucleus and activate gene transcription [3]. Different stimuli may activate either canonical (RelA) or non-canonical (RelB) pathways, which differ by way of alternative dimeric combinations of the family members (Figure 1). Dynamic intracellular localization of NF-κB Initially, NF-κB activation was regarded as a simple switch where NF-κB nuclear localization was terminated by NFκB-directed synthesis of IκB, thus preventing further