![Codepletion of Cep57 and Cep57L1 causes the increase of centrosome number during interphase.(A) Depletion of Cep57 induced precocious centriole disengagement in mitosis, but not in interphase. HeLa cells were treated with siControl or siCep57 and immunostained with antibodies against CP110 (green) and Cep192 (red). White arrows indicate the distance between the disengaged centrioles. (B) Histograms represent the frequency of cells in interphase and mitosis with the indicated phenotypes observed in A. Values are mean percentages ± SD from three independent experiments (n = 50 for interphase and n = 30 for mitosis for each experiment). (C) Codepletion of Cep57 and Cep57L1 (Cep57/Cep57L1) induced precocious centriole disengagement and increase of centrosome number in interphase. HeLa cells were treated with siControl or siCep57/Cep57L1 and immunostained with antibodies against CP110 (green) and Cep192 (red). White arrows indicate the distance between the disengaged centrioles. Note that because the centrosomal linker does not function well in HeLa cells, the disengaged centrioles were sometimes well separated in Cep57/Cep57L1-depleted cells. (D–F) Histograms represent the frequency of the interphase cells with more than two Cep192 foci (D), centriole disengagement (E), or more than four centrioles (F), respectively. The quantification was performed on all interphase cells at random. Values are the mean percentages ± SD from three independent experiments (n = 50 for each experiment). (G) Precocious centriole disengagement in interphase induced by Cep57/Cep57L1 codepletion was rescued by exogenous expression of Cep57 or Cep57L1. HeLa cells were treated with siControl or siCep57/Cep57L1, followed by the transfection with FLAG empty (control), FLAG-Cep57 (RNAi resistant [RNAi-R]), or FLAG-Cep57L1 (RNAi-R). The cells were immunostained with antibodies against FLAG (green), CENP-F (red), and Cep192 (cyan). (H) Histograms represent the frequency of cells in the G2 phase with more than two Cep192 foci in each condition observed in G. Values are percentages from three independent experiments (n = 30 for each experiment). All scale bars, 5 µm in the low-magnification view, 1 µm in the inset. A two-tailed, unpaired Student’s t test was used in B to obtain the P value. Dunnett’s multiple comparisons test was used in D, E, F, and H to obtain P values. ***, P < 0.001; *, P < 0.05.](https://cdn.rupress.org/rup/content_public/journal/jcb/220/3/10.1083_jcb.202005153/4/jcb_202005153_fig1.png?Expires=1772861706&Signature=L8ZIUlRCc0uMG2bChSV6ZeX11rwnMfgle~FJlufUEGQUKBADsQj3d6uVu0GuJ2gBBy4K6ZLn9XbXHZjAAyGtAf54OuJRw-r2PvxjDa-c80JSEfF-hMAEwMwuxumhANbRSWHOyWTicg2E5FC4blCUaYUDHKREXBq2-vQtDwJn-zk526qlumNnONyQesEuPOTfxXv9LhJKrn7AX~9CYT3RvwHiNSib7iQp9ctjqO8o9Pcaa5nWZIuUfmICfT-~dNOddvNfetIL23RJjMp3n4dmu9FxGksxa38KkRFexljQBqoaRiF9ke7~FPB2tdutlduMlO9P4oX2K~l-PrRClHlULw__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Codepletion of Cep57 and Cep57L1 causes the increase of centrosome number during interphase. (A) Depletion of Cep57 induced precocious centriole disengagement in mitosis, but not in interphase. HeLa cells were treated with siControl or siCep57 and immunostained with antibodies against CP110 (green) and Cep192 (red). White arrows indicate the distance between the disengaged centrioles. (B) Histograms represent the frequency of cells in interphase and mitosis with the indicated phenotypes observed in A. Values are mean percentages ± SD from three independent experiments (n = 50 for interphase and n = 30 for mitosis for each experiment). (C) Codepletion of Cep57 and Cep57L1 (Cep57/Cep57L1) induced precocious centriole disengagement and increase of centrosome number in interphase. HeLa cells were treated with siControl or siCep57/Cep57L1 and immunostained with antibodies against CP110 (green) and Cep192 (red). White arrows indicate the distance between the disengaged centrioles. Note that because the centrosomal linker does not function well in HeLa cells, the disengaged centrioles were sometimes well separated in Cep57/Cep57L1-depleted cells. (D–F) Histograms represent the frequency of the interphase cells with more than two Cep192 foci (D), centriole disengagement (E), or more than four centrioles (F), respectively. The quantification was performed on all interphase cells at random. Values are the mean percentages ± SD from three independent experiments (n = 50 for each experiment). (G) Precocious centriole disengagement in interphase induced by Cep57/Cep57L1 codepletion was rescued by exogenous expression of Cep57 or Cep57L1. HeLa cells were treated with siControl or siCep57/Cep57L1, followed by the transfection with FLAG empty (control), FLAG-Cep57 (RNAi resistant [RNAi-R]), or FLAG-Cep57L1 (RNAi-R). The cells were immunostained with antibodies against FLAG (green), CENP-F (red), and Cep192 (cyan). (H) Histograms represent the frequency of cells in the G2 phase with more than two Cep192 foci in each condition observed in G. Values are percentages from three independent experiments (n = 30 for each experiment). All scale bars, 5 µm in the low-magnification view, 1 µm in the inset. A two-tailed, unpaired Student’s t test was used in B to obtain the P value. Dunnett’s multiple comparisons test was used in D, E, F, and H to obtain P values. ***, P < 0.001; *, P < 0.05.
