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Fig. 1. Cenp-F is required for cell division. HeLa cells were transfected with siRNA duplexes then analysed 48 hours later by immunoblotting and immunofluorescence. (A) Immunoblot showing effective repression of Cenp-F, with tubulin acting as a loading control. (B) Quantitation of at least 100 mitotic cells by immunofluorescence scoring the levels of kinetochore-bound Cenp-F as either high, medium or low. (C) Images of mitotic cells showing that Cenp-F and Bub1 colocalize at kinetochores in control cells, but that Cenp-F is absent from kinetochores in Cenp-F RNAi cells; bar, 5 µm. (D) Images of mitotic cells stained to detect Cenp-F and Aurora A showing a tripolar spindle in the Cenp-F-deficient cell; bar, 5 µm. (E) Quantitation of mitotic cells with two or more spindle poles. In the bar graphs, values represent the mean±s.e.m. of three independent experiments in which at least 100 mitotic cells were counted.
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