Figure 7: Expression of non-phosphorylatable EB2 leads to lagging chromosomes and aneuploidy.

(a) MCF10A cells were infected to generate a polyclonal population expressing FLAG-tagged EB2-WT and EB2-7A. Cell extracts were immunoblotted with the indicated antibodies. (b,c) Quantification of the proportions of EB2-WT- and EB2-7A-expressing MCF10A cells exhibiting lagging chromosomes. Representative images of normal and lagging chromosomes in b. Arrows indicate lagging chromosomes. Data are means± s.d. from three independent experiments (≥200 cells per experiment). *P<0.001; NS, not significant (unpaired t-test). (d,e) Proportion of cells with karyotypes deviating from the modal chromosome number was determined after 30 generations in the indicated cells. Individual chromosome numbers from chromosome spreads were determined in e. Data are means± s.d. from three independent experiments (≥50 cells per experiment). *P<0.001; NS, not significant (unpaired t-test). (f) Fluorescence in situ hybridization (FISH) analysis with probes for centromeric DNA of chromosomes 3 (red), 7 (green) and 17 (blue) was performed six generations after the indicated lentiviral infection. (g) Quantification of the proportion of cells exhibiting centromere signals deviating from the modal in f. Data are means± s.d. from three independent experiments (≥350 cells for each condition per experiment). **P<0.01; NS, not significant (unpaired t-test) compared with control. Scale bars, 5 μm.