Supplementary Figure 2: Dynamic canonical NOTCH1 signalling is responsible for reciprocal regulation of TGF-β ligands and proinflammatory cytokines during senescence.
From: NOTCH1 mediates a switch between two distinct secretomes during senescence
(a) ER:HRASG12V IMR90 cells were examined in a time series analysis of TGF-β ligand expression by qRT-PCR (upper) and JAG1 expression by immunoblotting (lower) after RAS induction by 4OHT; values are mean ± s.e.m.; the experiment was performed four times for day 0 through 6 and three times for day 8 with similar results. (b) ER:HRASG12V IMR90 cells with and without 3 days treatment with 4OHT and DAPT at the indicated concentrations were analysed by qRT-PCR; One-way ANOVA with Dunnett’s multiple comparison test; values are mean ± s.e.m.; n = 6 biologically independent experiments for HES1, IL1A and IL8, n = 5 biologically independent experiments for TGFB1. ∗P ≤ 0.05, ∗∗P ≤ 0.01, ∗∗∗P ≤ 0.001. (c) Time-dependent behaviour of TRE-N1ICD-FLAG IMR90 cells after induction of N1ICD expression by addition of doxycycline: immunoblot demonstrating expression of indicated proteins. (d) TRE-N1ICD IMR90 cells with and without canonical NOTCH-inhibitor dnMAML1 were analysed for colony forming capacity. (e) TRE-N1ICD IMR90 cells treated with or without doxycycline for 7 days were then either cultured with continued doxycycline treatment or in regular media for a further 10 days before analysis of colony forming capacity. Statistics source data for a and b are provided in Supplementary Table 2.
