Figure 5.
CENP-F functions to limit dynein stripping of the corona. (a) Left: Immunofluorescence microscopy images from the dynein-stripping assay for CENP-E in CENP-F-Mut1 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against CENP-E and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore CENP-E intensity relative to CENP-C in the dynein-stripping assay. (b) Left: Immunofluorescence microscopy images of the dynein-stripping assay for CENP-E in CENP-F-Mut2 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against CENP-E and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore CENP-E intensity relative to CENP-C in the dynein-stripping assay. (c) Left: Immunofluorescence microscopy images of the dynein-stripping assay for Mad2 in CENP-F-Mut1 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against Mad2 and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore Mad2 intensity relative to CENP-C in the dynein-stripping assay. (d) Left: Immunofluorescence microscopy images of the dynein-stripping assay for Mad2 in CENP-F-Mut2 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against Mad2 and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore Mad2 intensity relative to CENP-C in the dynein-stripping assay. (e) Left: Immunofluorescence microscopy images of the dynein-stripping assay for Zwlich in CENP-F-Mut1 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against Zwlich and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore Zwlich intensity relative to CENP-C in the dynein-stripping assay. (f) Left: Immunofluorescence microscopy images of the dynein-stripping assay for Zwlich in CENP-F-Mut2 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against Zwlich and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore Zwlich intensity relative to CENP-C in the dynein-stripping assay. Boxes depict the median and first and third quartiles, and whiskers represent Q1 and Q3 ± 1.5× interquartile range. ****, P < 0.0001.

CENP-F functions to limit dynein stripping of the corona. (a) Left: Immunofluorescence microscopy images from the dynein-stripping assay for CENP-E in CENP-F-Mut1 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against CENP-E and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore CENP-E intensity relative to CENP-C in the dynein-stripping assay. (b) Left: Immunofluorescence microscopy images of the dynein-stripping assay for CENP-E in CENP-F-Mut2 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against CENP-E and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore CENP-E intensity relative to CENP-C in the dynein-stripping assay. (c) Left: Immunofluorescence microscopy images of the dynein-stripping assay for Mad2 in CENP-F-Mut1 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against Mad2 and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore Mad2 intensity relative to CENP-C in the dynein-stripping assay. (d) Left: Immunofluorescence microscopy images of the dynein-stripping assay for Mad2 in CENP-F-Mut2 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against Mad2 and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore Mad2 intensity relative to CENP-C in the dynein-stripping assay. (e) Left: Immunofluorescence microscopy images of the dynein-stripping assay for Zwlich in CENP-F-Mut1 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against Zwlich and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore Zwlich intensity relative to CENP-C in the dynein-stripping assay. (f) Left: Immunofluorescence microscopy images of the dynein-stripping assay for Zwlich in CENP-F-Mut2 cells. Cells were treated with control or DHC siRNA, arrested in nocodazole or monastrol, and stained with DAPI and antibodies against Zwlich and CENP-C. Scale bar, 5 µm. Right: Quantification of the kinetochore Zwlich intensity relative to CENP-C in the dynein-stripping assay. Boxes depict the median and first and third quartiles, and whiskers represent Q1 and Q3 ± 1.5× interquartile range. ****, P < 0.0001.

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