Fig. 6. SIRT7 is involved in resumption of rDNA transcription at the exit from mitosis. (A) BrUTP incorporation in living cells was detected using anti-BrdU antibodies in interphase HeLa cells in the absence (a-b) or presence of AMD (0.1 µg/ml, c-d, or 4 µg/ml, e-f, for 2 hours) or sirtinol (100 µM for 6 hours, g-h). (B) Knockdown of SIRT1 and SIRT7 was analyzed by 10% SDS-PAGE and immunoblotting using anti-SIRT1 and anti-SIRT7-N-terminal antibodies. (C) SIRT1 and BrUTP incorporation were detected using anti-SIRT1 (b,e) and anti-BrdU (c,f) antibodies, respectively, in control (a-c) or SIRT1-depleted (d-f) HeLa cells. (D) SIRT7 and BrUTP incorporation were detected using anti-SIRT7 (b,e) and anti-BrdU (c,f) antibodies, respectively, in control (a-c) or SIRT7-depleted (d-f) HeLa cells. (E) SIRT7 and rDNA transcription sites were immunodetected using anti-SIRT7-C-terminal (b,f,j) and anti-BrdU (c,g,k) antibodies in prometaphase mitotic HeLa cells, in the absence (a-d) or presence (e-h) of roscovitine (150 µM for 1 hour). Mitotic HeLa cells were also treated with 100 µM sirtinol for 2 hours before addition of 150 µM roscovitine for 1 hour (i-l). Red and green labelings were merged (d,h,l). Bars, 10 µm.