Interaction of NMNAT2 with SIRT3 promotes mitotic entry of A549. (A) Interaction of NMNAT2 with SIRT3 did not affect DNA synthesis. A549 cells were transfected with plasmids and synchronized at the G1/S transition as described in Materials and methods. Cells were pulse-labeled with BrdU (50 μ M) for 30 min at indicated time-points after release from the second thymidine block. BrdU positive cells were detected by immunostaining and scored manually. More than 500 cells were counted in each of the three experiments. (B) Interaction of NMNAT2 with SIRT3 promoted mitotic entry. Cell cycle progression of > 1,000 cells was recorded by time-lapse videomicroscopy. The number of mitotic cells was scored by examination of individual cells. Values are means of three experiments. All data are presented as the mean ± SEM.