Figure 5. Depletion of Cdc25C protein prevents LMW-E–mediated centrosome amplification and chromosome missegregation.

A, pTRE-LMW-E MCF-7 cells were transfected with control siRNA or Cdc25C-specific siRNA for 72 h, and Cdc25C and cyclin E expression were analyzed by western blotting (left panels). Cyclin E immunofluorescence (IF) (right panels) was also used to examine cyclin LMW-E levels in the presence or absence of pTRE-LMW-E induction. Cdc25C IF was used to examine Cdc25C levels in the siRNA control or siCdc25C pTRE-LMW-E induced cells. B, pTRE-LMW-E cells were treated with control or Cdc25C-specific siRNA for 72 h, treated with or without LMW- E induction, and costained for β–tubulin (green) and γ–tubulin (red), and DAPI (blue) to analyze the mitotic defects. C, Cells treated as in B were costained with pericentrin (green), γ–tubulin (red), or DAPI (blue) to analyze the number of centrosomes. D, Left panel, centrosomes were counted in cells (n= 500) stained with γ–tubulin and pericentrin for each condition in three independent experiments. *P < 0.01. Right panel, mitotic cells (n= 100) were analyzed for abnormalities such as micronulei and chromosome missegregation for each condition in three independent experiments. *P < 0.01.