A systematic approach to decipher crosstalk in the p53 signaling pathway using single cell dynamics

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Abstract
The transcription factors NF-κB and p53 are key regulators in the genotoxic stress response and are critical for tumor development. Although there is ample evidence for interactions between both networks, a comprehensive understanding of the crosstalk is lacking. Here, we developed a systematic approach to identify potential interactions between the pathways. We perturbed NF-κB signaling by inhibiting IKK2, a critical regulator of NF-κB activity, and monitored the altered response of p53 to genotoxic stress using single cell time lapse microscopy. Fitting subpopulation-specific computational p53 models to this time-resolved single cell data allowed to reproduce in a quantitative manner signaling dynamics and cellular heterogeneity for the unperturbed and perturbed conditions. The approach enabled us to untangle the integrated effects of IKK/ NF-κB perturbation on p53 dynamics and thereby derive potential interactions between both networks. Intriguingly, we find that a simultaneous perturbation of multiple processes is necessary to explain the observed changes in the p53 response. Specifically, we show interference with the activation and degradation of p53 as well as the degradation of Mdm2. Our results highlight the importance of the crosstalk and its potential implications in p53-dependent cellular functions. Cells can respond to external and internal inputs by transducing information to the nucleus where transcription factors initiate corresponding cellular responses. Cellular signaling is mediated by several pathways; molecular networks that can interact with each other, which alters signal processing and modulates cellular responses. As deregulated signaling can lead to the development of tumors it is important to understand not only how signaling pathways function but also the contribution of their interaction on the signaling dynamics. Here, we analyzed the interplay of the IKK/ NF-κB and p53 pathway, which are both critical for the cellular response to DNA damage and have been implicated in tumor development. To systematically identify interaction points between both pathways, we perturbed IKK/ NF-κB signaling and tracked the changes in the response of p53 to DNA damage. Using computational methods, we show that several reactions in the p53 pathway are simultaneously affected by NF-κB signaling and that this combined action is necessary to explain altered behaviour of the p53 pathway. Hence, our results provide new insights into the interplay between the NF-κB and p53 pathway and help to gain a more comprehensive understanding of the crosstalk.
Funding Information
  • Deutsche Krebshilfe (111645)
  • Deutsche Forschungsgemeinschaft (Graduiertenkolleg 1772)
  • Bundesministerium für Forschung, Technologie und Raumfahrt (0316047A)
  • Helmholtz-Gemeinschaft (iMed Initiative)

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