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. Author manuscript; available in PMC: 2018 Mar 1.
Published in final edited form as: Nat Cell Biol. 2017 Jul 31;19(9):1061–1070. doi: 10.1038/ncb3586

Figure 5. Engagement of cGAS is a common feature of multiple senescence triggers.

Figure 5

(a) MEFs with the indicated genotypes were exposed to 12 Gy ionizing irradiation or stimulated with Palbociclib. Expression levels of Cxcl10 or protein levels of IL-6 were quantified by RT-qPCR or ELISA, respectively. (b) WI-38 cells (left panel) were exposed to 12 Gy ionizing irradiation or WI-38 ER:RAS cells (right panel) were treated with 4-OHT (500 nM) for 7 days. At day 7 (irradiated cells) or at day 3 (WI-38 ER:RAS cells) were transfected with a non-targeting control siRNA (si Control) or with siRNAs against cGAS (si cGAS #1, si cGAS #2). Expression of depicted genes was determined by RT-qPCR. (c) Expression of lamin B1 in individual WT MEFs exposed to irradiation (left panel) or treated with Palbociclib (right panel) for 7 days was determined by confocal fluorescence microscopy (CTCF = Corrected Total Cell Fluorescence). (d) Expression of lamin B1 in individual WI-38 cells exposed to irradiation (left panel) or WI-38 ER:RAS cells treated with 4-OHT was assessed by confocal fluorescence microscopy (CTCF = Corrected Total Cell Fluorescence). (e) cGAS KO MEFs carrying a Dox-inducible lentiviral vector containing a FLAG-tagged murine cGAS construct were irradiated (12 Gy) or treated with Palbociclib. 2 days post treatment cells were cultured in the presence of Dox overnight and then stained for FLAG (green) and nuclei (DAPI, grey). One representative experiment out of 2 experiments is shown (e) or mean and s.d. of n=51 individual control MEFs compared with n=48 individual irradiated MEFs (c, left); n=50 individual control MEFs compared with n=58 individual Palbociclib treated MEFs (c, right); n=177 individual control WI-38 cells compared with n=158 individual irradiated WI-38 cells (d, left); n=151 individual control WI-38 cells compared with n=87 individual 4-OHT treated WI-38 cells (d, right) are shown. Data shown in (a) and (b) are from n=2 independent experiments with the column representing the mean. P values were calculated by unpaired t-test (c, d) (**** P < 0.0001, ns = not significant). Scale bar: 20 µm. Source data are available in Supplementary Table 1.