Figure 3.

Rapamycin suppresses NF-κB transcriptional activity. (a) Transcripts for SASP factors secreted by DMSO- or rapamycin (Rapa)-treated senescent (ionizing radiation; Sen (IR)) HCA2 cells were quantified by qPCR 8 days after ionizing radiation exposure. Cells were incubated in serum-free media without rapamycin for the last 24 h. For the heatmap, the average signal from non-senescent (NS), senescent (ionizing radiation; Sen (IR)) DMSO-and rapamycin-treated HCA2 cells was used as the baseline (for numbers, see Supplementary Table 1). Colour intensities represent log₂-fold changes from the baseline. Signals higher than the baseline are shown in yellow; signals lower than the baseline are shown in blue. (b) Transcripts for SASP factors from DMSO- or rapamycin-treated senescent (ionizing radiation; Sen (IR)) HCA2 cells were quantified by qPCR 2 days after ionizing radiation exposure. For the heatmap, the average signal from non-senescent, senescent DMSO-and rapamycin-treated cells was used as the baseline (for numbers, see Supplementary Table 2). Colour intensities represent log₂-fold change from the baseline. Signals higher than the baseline are shown in yellow; signals lower than the baseline are shown in blue. (c) Cell extracts were prepared from non-senescent (NS) and senescent (ionizing radiation; Sen (IR)) HCA2 cells expressing an NF-κB–luciferase reporter construct. Cells were treated with DMSO (control) or rapamycin for 7 days, and analysed for luciferase activity as described previously^11,15. Non-senescent luciferase activity was set at 1. For a and b, shown are the average of three independent experiments. For c, shown is one representative of two independent experiments, each with triplicate samples. For raw data, see Supplementary Table 4.