Fig. 2.

Plk3 is expressed in G₁. (a and b) MCF10A cells were synchronized by 72 h of serum starvation. (a) Serum and hormones then were added to the cells, and the cells analyzed at 4-h intervals by FACS. (b) Total cellular protein was analyzed from duplicate plates by Western blot. (a) Under these conditions, G₂/M occurred between 24–32 h. (b) The level of Plk3 protein peaked in G₁ near 8 h, whereas the level of Plk1 peaked as expected during G₂/M. Levels of the G₁ cyclins, cyclin D1 and cyclin E1, were analyzed for comparison. Nucleolin and Erk2 loading controls are included. (c and d) HeLa cells were synchronized by double thymidine block. Cells were analyzed at 2-h intervals by FACS (c), and total cellular protein was analyzed from duplicate plates by Western blot (d). (c) Double thymidine block synchronizes cells in early S phase with slightly more than 2N DNA content (0 h). M phase cyclin B1 expression peaked at 10 h after release and again at 24 h. The level of Plk3 protein peaked in G₁ at 14–16 h (c and d), concurrent with the peak expression of the G₁ phase cyclin D1. Erk2 loading control is also shown.