Figure 5.
SIRT1 is recruited to DNA breaks and is required for efficient DSB repair. (A) SIRT1 expression in parental U2OS-DRGFP cells, three independent shSIRT1 expressing cell lines (SIRT1-KD) and shRFP expressing controls. (B) ChIP analysis of SIRT1 and Nbs1 binding to a DSB in shRFP (control) and SIRT1 KD cells from (A). (C) DSB ChIP analysis of Rad51 recruitment using Ds-red-transfected (−) or I-SceI transfected cells (+) from (B) 24h after transfection. (D) SIRT1 knock-down results in reduced DSB repair as measured by GFP rexpression (see Figure S7A). Control and SIRT1-KD cell lines from (A) were transfected as in (C) and analyzed by FACS. (E) Loss of SIRT1 causes increased genomic instability upon oxidative stress. Untreated or H2O2 treated shSIRT1- or control shRNA-expressing ES cells were subjected to Q-FISH analysis. Telomeres are shown in red. Arrows indicate a chromatid break (left) and a fused centromere (right). The fraction of aberrant metaphases is shown. Data are represented as mean +/− SEM.