Condensin-depleted ES cells arrest at metaphase and accumulate large, misshapen interphase nuclei. (A) Fractions of mitotic cells in each stage of mitosis in mock- or Smc2-depleted ES cells. Live cells expressing H2A-GFP were counted and scored for each phase of mitosis. Means and standard deviations of triplicate samples are shown. (B) Representative time-lapse live cell imaging of mock- or Smc2-depleted H2A-GFP ES cells starting in metaphase and imaged at the times (in minutes) indicated in each box. ES cells grow in clusters and rotate within the cluster, resulting in different views of the metaphase plate during the time lapse. (C) Fields of H2A-GFP–expressing ES cells either mock or Smc2 depleted for 4 d. White arrowheads denote multipolar mitoses. (D) ES cells either mock or Smc2 depleted for 4 d were fixed, cytospun onto slides, and DAPI stained, and 1-µm z sections were captured. Deconvolved z sections at the midpoint of each stack are shown. (E) Box plot of the nuclear volumes (nuc. vol.) in thousands of femtoliters (fL × 103) of 10–20 ES cell nuclei for each KD indicated. The top and bottom edges of each box represent the 75th and 25th percentiles, respectively, whereas the bold line represents the median of each group. The asterisk indicates a statistically significant difference between the nuclear volumes of mock and Smc2 KD cells (P < 2 × 10−5; Student’s t test). Bars: (B) 2.5 µm; (C) 30 µm; (D) 10 µm.
