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. 2013 Jun 24;288(32):23348–23357. doi: 10.1074/jbc.M113.465302

FIGURE 5.

FIGURE 5.

Che-1 regulates Chk1 centrosomal localization. A, total cellular extracts from HCT116 cells, untreated or treated with 1 μm Doxorubicin for 2 h, were immunoprecipitated (I.P.) with anti-Chk1 monoclonal antibody and analyzed by Western blot using the indicated antibodies. Neg. control, negative control. B, HCT116 cells transiently transfected with GFP siRNA (siControl) or Che-1 siRNA (siChe-1-1) were immunostained with the indicated antibodies. Bar diagrams show the percentages of cells with centrosomal colocalization of γ-tubulin and Chk1. C, isolated centrosomes or whole cell lysates from HCT116 cells transiently transfected as in B were analyzed by Western blot with the indicated antibodies. D, HCT116 cells transiently transfected with shRNA GFP (shControl) or shRNA Chk1 (shChk1) were immunostained with the indicated antibodies. Bar diagrams show the percentages of cells with centrosomal colocalization of γ-tubulin and Che-1. E, isolated centrosomes or whole cell lysates from HCT116 cells transiently transfected as in D were analyzed by Western blot using the indicated antibodies. F, isolated centrosomes or whole cell lysates from HCT116 cells transiently transfected as in B were analyzed by Western blot with the indicated antibodies. Nuclei were visualized by staining with 1 μg/ml Hoechst dye 33258.The scale bar represents 5 μm. At least 3 × 100 cells were counted per data bar. Results are given as mean ± S.D.