Figure 6
Downregulation of CDK1 leads to multiple abnormalities in kinetochore, centrosome and centriole numbers in hESC. (A) Specific antibody to kinetochores, CREST (red), was used for immunofluorescence staining of hESC treated with control siRNA (a, c) and CDK1 siRNAs (b, d) at 2 days post transfection. DNA was counterstained with DAPI (blue) in all images. Examples of hESC with supernumericial kinetochores at interphase nuclei observed in control siRNA (a) and CDK1 siRNA-treated cells (b). (B) Immunostaining of centrosomes with a specific antibody, Pericentrin (red). Examples of hESC with normal (b, d, f) and supernumerary centrosomes (a, c, g, h) at 2 days post transfection with control and CDK1 siRNAs, respectively. Note abnormal nucleus blebbing on the CDK1 siRNA group shown by long arrows (e). Centrosomes are shown by short white arrows. (C) Centrin2-specific antibody (red) for visualisation by immunofluorescence of centrioles at 2 days post hESC transfection with control siRNA (a, b, c) and CDK1 RNAi (d–k). Normal (a: two centrioles; pointed by arrow) and abnormal centriole number (b) was observed in the control siRNA-transfected group in interphase nucleuses (b) and during mitosis (c). White arrows point to centrioles. Knockdown of CDK1 in hESC causes abnormalities in centriole number in interphase nuclei (d), nuclei blebbing with abnormal centriole number (e, f) and abnormal mitoses (g–k) with wrong distribution and number of centrioles (g–k). A–C: Scale bar=5 μm. Images are representative of at least three independent experiments
