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. Author manuscript; available in PMC: 2016 Jan 1.
Published in final edited form as: Nat Cell Biol. 2014 Dec 15;17(1):31–43. doi: 10.1038/ncb3076

Figure 6. Retention of Cep68 or PCNT in late mitosis prevents the removal of Cep215.

Figure 6

a) Expression of non-degradable Cep68(S332A) results in the retention of Cep215 on centrosomes in late mitosis. Cells expressing Cep68 or Cep68(S332A) were fixed and stained with antibodies recognizing Cep68 (red) and anti-Cep215 (green) antibodies. Cells in interphase and metaphase expressing HA-tagged Cep68 constructs were analyzed by immunofluorescence. The localization of Cep215 in telophase in cells expressing FLAG-tagged Cep68 constructs was analyzed by three-dimensional structured-illumination microscopy (3D-SIM). Scale bars represent 1 μm. The graph below shows the relative intensity of Cep215 quantified in metaphase and telophase cells expressing HA-tagged Cep68 or Cep68(S332A). The bars represent the mean ± standard deviation (S.D.). Cep68 metaphase: n=30 centrosomes; Cep68(S332A) metaphase: n=26 centrosomes; Cep68 telophase: n=30 centrosomes; Cep68(S332A) telophase: n=32 centrosomes. ****P < 0.0001.

b) Expression of non-cleavable PCNT(R2231A) in late mitosis prevents Cep215 removal. HeLa cells were transiently transfected with FLAG-PCNT or FLAG-PCNT(R2231A). Cells were fixed and stained with antibodies to FLAG (red) and Cep215 (green). The localization of Cep215 in telophase cells expressing FLAG-tagged PCNT constructs was analyzed by 3D-SIM. The areas in the white boxes are shown at higher magnification directly above the corresponding image. Scale bars represent 1 μm. This experiment was reproduced three times.