Figure 3.

Disrupting the INCENP–HP1 interaction delocalizes HP1 from mitotic centromeres and increases chromosome missegregation. A, lysates of nocodazole-arrested mitotic HeLa cells with or without stable expression of the indicated exogenous INCENP-GFP proteins were immunoblotted. B and C, HeLa cells and the indicated stable cell lines transfected with control or INCENP siRNA were treated with nocodazole for 3 h. Mitotic cells collected by shake-off were spun on slides and stained with DAPI and antibodies for GFP, HP1α, and CENP-C (B). The mitotic cell lysates were immunoblotted (C). D and E, asynchronous HeLa cells and the indicated stable cell lines transfected with control or INCENP siRNA were stained with DAPI and antibodies for Aurora B, CENP-C, or ACA. Example cells in prometaphase (D) and interphase (E) are shown. F and G, asynchronous HeLa cells and the indicated stable cell lines were fixed and stained with ACA and DAPI. The percentage of cells with lagging chromosomes was determined in 100 anaphase cells (F). Example images are shown (G). Scale bars, 10 μm. See also Fig. S2.