Phosphorylation of INCENP at Ser894 is required for telophase progression. (A) HeLa-S3 clones inducibly expressing 6Myc-INCENP-WT, -S894A, -S894D, or vector alone were fixed 48 h after addition of doxycycline. Cells were immunostained with anti-Myc antibody and counterstained with Hoechst 33258. For Myc-positive cells, cell number and nuclear number were counted by microscopy. Data represent the average of ∼100 cells from three independent experiments. (B) Percentage of mitotic cells (including cells in prometaphase, metaphase, anaphase, and telophase) in HeLa-S3 clones inducibly expressing 6Myc-INCENP-WT, -S894A, -S894D, or vector alone (blue bars). Cells were counted as in A. (C) HeLa-S3 clones inducibly expressing 6Myc-INCENP-WT, -S894A, -S894D, or vector alone were fixed 11 h after release from a thymidine block, and Myc-positive mitotic cells were counted by microscopy. Data represent the average of ∼50 mitotic cells from three independent experiments.