Fig. 6. Nocodazole-induced centrosome splitting depends on protein kinase activity. (A,B) U2OS cells either treated for 1 hour with nocodazole, either alone or in combination with staurosporine (500 µM) or 6-DMAP (2.5 mM), before centrosome splitting was analyzed. Alternatively, MT disassembly was induced by incubating cells for 1 hour on ice. As assessed by immunofluorescent staining with antibodies against 
-tubulin, all treatments caused complete disassembly of MTs (not shown). (A) Histogram indicating the percentages of cells with split centrosomes; (B) 
-tubulin stained centrosomes, illustrating the suppression of nocodazole-induced centrosome splitting by staurosporine. Bar, 10 µm. (C,D) U2OS cells were transfected for 24 hours with catalytically inactive mutants of the kinases indicated, and 1 hour prior to fixation they were treated with nocodazole. For control, untransfected cells were treated with nocodazole alone or with nocodazole plus staurosporine. Then, all samples were scored for centrosome splitting (C). (D) -tubulin stained centrosomes, illustrating the suppression of nocodazole-induced centrosome splitting by the catalytically inactive Nek2 K37R mutant. Bar, 10 µm.
