Fig 1: S100A13 modulates NF-?B activity and SASP expression during cell senescence. (A) HCT116 cells were transfected with the control vector, S100A13 wild type or mutant type, and treated with Dox (100 nM) for 3 days. Then cell lysates were subjected to western blot analysis using the indicated antibodies. (B) ER:Ras IMR90 cells stably transduced with PHBLV, S100A13 wild type or mutant type were given 4-OHT for 6 days. Cell lysates were then subjected to Western blot analysis for the indicated proteins. (C) HCT116 cells were transfected with control (siNC) or two independent siRNAs against S100A13, and treated with Dox (100 nM) for 4 days. Then cell lysates were subjected to western blot analysis using the indicated antibodies. (D) ER:Ras IMR90 cells stably transduced with control (siNC) or two independent shRNAs against S100A13, were given 4-OHT for 6 days. Cell lysates were then subjected to Western blot analysis for the indicated proteins. (E and F) HCT116 cells were transfected with vector, S100A13 wild type or mutant type, or transfected with control (siNC) or two independent siRNAs against S100A13, and treated with Dox (100 nM) for 4 days. Then mRNA levels of some SASP genes were analyzed by real-time qPCR (n=3). (G) HCT116 cells were transfected with wild type S100A13, and treated with Dox (100 nM) for 3 days. Control IgG (0.6 ug/ml) or neutralizing antibody IL-1a (0.6 ug/ml) were added for the last 2 days. Then cell surface-bound IL-1a were analyzed by FACS (n=3). (H) HCT116 cells were transfected with siRNA#2 against S100A13, and treated with Dox (100 nM) for 3 days. Solvent or recombinant IL-1a protein (300 ng/ml) were added for the last 2 days. Then cell surface-bound IL-1a were analyzed by FACS (n=3). Three independent experiments were performed. Error bars represent means ± SD (n = 3) *P < 0.05, **P < 0.01, ***P < 0.005, ****P < 0.001 in (E–H).
Fig 2: S100A13 modulates multiple SASP and cellular senescence regulators. (A–D) HCT116 cells were transfected with the control, S100A13 wild type or mutant type (A and C), or transfected with the control (siNC) or two independent siRNAs against S100A13 (B and D), and then treated with Dox (100 nM) for 3 or 4 days. Then cell lysates were subjected to western blot analysis for the indicated proteins. (E) HCT116 cells were transfected with wild type S100A13, and treated with Dox (100 nM) for 3 days. Control IgG (0.6 ug/ml) or neutralizing antibody IL-1a (0.6 ug/ml) were added for the last 2 days. Then the indicated proteins were analyzed by Western blot. (F) HCT116 cells were transfected with siRNA#2 against S100A13, and treated with Dox (100 nM) for 3 days. Solvent or recombinant IL-1a protein (300 ng/ml) were added for the last 2 days. Then the indicated proteins were analyzed by Western blot. (G and H) ER:Ras IMR90 cells were given 4-OHT for toal 6 days in the presence of the indicated doses of Amlexanox (G); or TTM (H); then the indicated proteins were analyzed by Western blot. Three independent experiments were performed.
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