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Fig. 3. Aberrant mitosis and apoptosis are consequences of FACE1 knockdown. (A) RNAi mediated knockdown of FACE1 led to mitotic arrest of HeLa SS6 cells within 24 hours of transfection. Arrested cells (top) displayed abnormal mitotic spindle architecture and disturbed chromosome congression. Control cells (bottom) transfected with GL2 siRNA showed normal mitosis. Cells were stained with tubulin antibody and with Hoechst dye. (B) TUNEL assay on FACE1-silenced (top) and control cells (bottom) 72 hours after siRNA transfection shows that the mitotically arrested cells have entered apoptosis. (C) Quantification of mitotic arrest after FACE1 silencing. The fraction of cells showing mitotic arrest increased with time after transfection, reaching a maximum of 35% of the total cell population at 48 hours. Cells transfected with either GL2 or with lamin A siRNA did not arrest in mitosis. (D) Quantification of apoptosis after FACE1 silencing. The percentage of apoptotic cells reached a maximum of 25% 72 hours after transfection and then decreased. Apoptosis was observed only in FACE1-silenced cells, but not in control (GL2 siRNA) or in lamin A siRNA-transfected cells. (E) Cell growth measured with the CellScreen apparatus was reduced dramatically after silencing with FACE1 siRNA. Bar, 15 µm.





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