Defective T-Segment Transit may Activate the Mad2-Dependent Checkpoint.

<p>Cell cycle analyses showing that Top2^L475A,L480P activates Mad2-dependent but Rad53-independent checkpoint signaling. <b>a</b>, Cartoon describing catalytic defect in Top2^L475A,L480P which has a reduced rate of T-segment transport due to inefficient G-segment cleavage and DNA-gate opening. <b>b–d</b>, Population Assays: Histogram plots show average G2/M duration; see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003832#s3" target="_blank">Material and Methods</a> for statistical analysis. Western blots show each Top2 mutant relative to Tub1 loading control at G1 and G2. <i>a</i> values are significantly different to <i>b</i> values in the histogram plots. Strains with the same letter are not significantly different. <b>e–f</b>, Single-cell assays: <i>e</i>, plots of average spindle length versus time for single cells aligned on the x-axis at the time of SPB separation (<i>i.e.</i> at time point 12 min). Error bars show standard deviation of lengths. <i>f</i>, Histogram plots of average time interval between SPB separation and the initiation of spindle elongation in anaphase B (+/− s.e.).