Fig. 3

Activation of DNA damage response pathways in iPS cell lines in response to TdR. a MIFF1, b MIFF3, and c MIFF4 iPS cell lines show a reduced CHK1 activation in western blots, comparable with what is observed in the Shef5N normal hES cell line (b) and in contrast to the strong activation observed in HCT116 cells (b). In all iPS cell lines, there is a reduced γH2AX phosphorylation compared with that observed in HCT116 treated with the CHK1 inhibitor Gö6976 (b), indicating that DNA damage is not enhanced in these cell lines in response to replication inhibitor TdR, despite the absence of a clear CHK1 activation. In addition, RPA is not hyperphosphorylated in any iPS cell lines, suggesting that ssDNA formation is suppressed. In contrast, HCT116 cells treated with the CHK1 inhibitor Gö6976 (b) show a marked hyperphosphorylation of RPA. d MIFF1 and f MIFF4 activate ATM by phosphorylation of Ser1981 after TdR treatment. This is accompanied by the phosphorylation of P53 at S15 in MIFF1 (d), MIFF3 (e), and MIFF4 (f). CHK1 checkpoint kinase 1, DMSO dimethyl sulfoxide, γH2AX γ histone 2AX, MIFF mRNA-induced foreskin fibroblast, RPA replication protein A, TdR thymidine