Fig. 5.

Sirt2 deacetylates RRM2 in a cell-cycle-dependent manner. a H1299 cells were synchronized at the G1/S boundary by double-thymidine block and then released into normal culture medium. RRM1, RRM2, Sirt2, KAT7, and cyclin A were analyzed by western blot at various time points. As asynchronous. b, c H1299 cells were transfected with Ctrl or Sirt2 siRNA or treated with Sirt2 inhibitor AGK2, followed by synchronization, cell cycling, and analysis of RRM2 acetylation as above. d, e H1299 cells were treated with Sirt2 siRNA or Sirt2 inhibitor (AGK2), followed by analysis of RNA activity. The error bars indicate ± s.d. of three separate experiments. *P <0.05, by two-tailed t test. f H1299 cells were transfected with Ctrl or Sirt2 siRNA or treated with Sirt2 inhibitor AGK2, followed by measurement of dNTP levels. The error bars indicate ± s.d. of three separate experiments. *P<0.05, by two-tailed t test. g, h H1299 cells were transfected with Sirt2 siRNA or treated with Sirt2 inhibitor AGK2, followed by analysis of cell proliferation and cell-cycle distribution using BrdU/7-AAD staining or colony-formation assay. The error bars indicate ± s.d. of three separate experiments. ***P <0.001, by two-tailed t test