INCENP is required for proper targeting of Survivin to the centromeres andthe anaphase spindle during mitosis

Three lines of investigation have suggested that interactions between Survi vin and the chromosomal passenger proteins INCENP and Aurora-B kinase may b e important for mitotic progression, first, interference with the function of Survivin/BIR1, INCENP, or Aurora-B kinase leads to similar defects in mi tosis and cytokinesis [1-7] (see [8] for review). Second, INCENP and Aurora -B exist in a complex in Xenopus eggs [9] and in mammalian cultured cells [ 7], Third, interference with Survivin or INCENP function causes Aurora-B ki nase to be mislocalized in mitosis in both C. elegans and vertebrates [5, 7 , 9], Here, we provide evidence that Survivin, Aurora-B, and INCENP interac t physically and functionally, Direct visualization of Survivin-GFP in mito tic cells reveals that it localizes identically to INCENP and Aurora-B, Sur vivin binds directly to both Aurora-B and INCENP in yeast two-hybrid and in vitro pull-down assays. The in vitro interaction between Survivin and Auro ra-B is extraordinarily stable in that it resists 3 M NaCl. Finally, Surviv in and INCENP interact functionally in vivo; in cells in which INCENP local ization is disrupted, Survivin adheres to the chromosomes and no longer con centrates at the centromeres or transfers to the anaphase spindle midzone, Our data provide the first biochemical evidence that Survivin can interact directly with members of the chromosomal passenger complex.