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. Author manuscript; available in PMC: 2011 Oct 25.
Published in final edited form as: Nat Cell Biol. 2007 Oct 14;9(11):1253–1262. doi: 10.1038/ncb1645

Figure 5.

Figure 5

SIRT1 desumoylation by UV radiation and hydrogen peroxide contributes to stress-induced apoptosis. (a) H1299 cells expressing HA–SIRT1 and His–Sumo-1 were treated with 0.5 mM hydrogen peroxide for 6 h or irradiated with 0.3 J UV. Sumoylated SIRT1 was isolated by Ni-NTA agarose and detected by immunoblotting with antibodies against HA. Cell extracts were immunoblotted with antibodies against HA and β-actin. (b) Cells were treated as in a and the level of endogenous SIRT1 sumoylation was determined by reciprocal coimmunoprecipitations. (c) Cells were cotransfected with HA–p53 and Flag–SIRT1 and treated with UV radiation or hydrogen peroxide. HA immunoprecipitates were immunoblotted with antibodies against p53 or p53 acetylated at Lys 382. Cellular extracts were immunoblotted with antibodies against Flag. (d) H1299 cells stably transfected with either wild-type Flag–SIRT1 or Flag–SIRT1K743R were treated with UV radiation or hydrogen peroxide. The percentage of apoptotic cells was determined by monitoring DNA content by flow cytometry. Each data point is the average of triplicate samples (n = 3) analysed in parallel. The error bars represent s.d. Uncropped images of the scans are shown in the Supplementary Information, Fig. S1.