Fig. 6

NF-κB adaptation to repeated temperature exposure. a Schematic representation of the thermotolerance: accumulation of inducible HSPs prevents IKK denaturation to repeated HS exposure. b Schematic representation of the repeated HS treatment protocol: cells exposed to two 1 h HS at indicated interval and subjected to cytokine stimulation. c Model simulation of TNFα-induced NF-κB responses following different HS protocols. Shown are sample 100 trajectories of single cell (and average responses based on 1000 simulated cells, in number of molecules, in black) treated in normal temperature 37 °C (left, in green), 4 h after a single 1 h 43 °C HS exposure (middle, in red) and immediately after second 1 h 43 °C HS exposure (right, in burgundy). d Model simulation of IL1β-induced NF-κB responses following different HS protocols, as in c. e Nuclear NF-κB trajectories in MCF7 cells stably expressing p65-EGFP in response to TNFα stimulation. Cells are either treated in normal conditions (left, data from Fig. 1), 4 h after a single 1 h 43 °C exposure (middle, data from Fig. 1) or immediately after second 1 h 43 °C HS exposure (right). Top: individual single cell nuclear NF-κB trajectories (n = 50, 50 and 40 per condition, respectively, in aribirary fluorescene units) depicted with coloured lines; black lines represented the population averages. Bottom: heat maps of trajectories normalized across all conditions in e and f (represented on a 0–3 scale). Cells monitored for up to 10 h from the beginning of cytokine stimulation. f Characteristics of TNFα-induced responses from e. From the left: distribution of area under the curve (AUC), first peak amplitude, and time to first response. Individual cell data are depicted with circles (with mean ± SD per condition). Kruskal-Wallis one-way ANOVA with Dunn’s multiple comparisons test was used to assess differences between groups (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001, ns – not significant). g Nuclear NF-κB trajectories in MCF7 cells stably expressing p65-EGFP following treatment with IL1β, represented as in e. p65-EGFP trajectories of cells cultured in normal conditions or exposed to IL1β after 4 h recovery from 1 h 43 °C are taken from Fig. 2. h Characteristics of IL1β-induced responses from g, data are represented as in f