CLASP2 phosphorylation on S1234 is required for normal chromosome alignment and segregation. (A and B) Selected frames of videos from HeLa cells stably expressing mCherry-H2B Histone (red) and transiently transfected with GFP-CLASP2 wild type (green; A) or GFP-CLASP2 S1234A mutant (green; B). Images show representative frames of the videos to draw attention to chromosome alignment, segregation, and cytokinesis. A’ and B’ panels show H2B channel alone. Large arrowheads indicate misaligned and lagging chromosomes. Small arrowheads indicate sister chromatid pairs. Time is shown in hours:minutes. Bar, 10 µm. (C) Quantification of chromosome alignment defects in cells expressing GFP-CLASP2 wild-type (n = 18) or S1234A mutant (n = 34). Bars represent the percentage of cells with 0, 1, 2, or more misaligned chromosomes at anaphase onset. The results represent a pool from at least three independent experiments.