(1 mM) for 8 days or for 2 days followed by inhibitor wash-out and continued incubation in media without 23dMB-PP1 for an additional 6 days. At day 8, plates were washed with PBS, fixed for 5 minutes with 96% methanol and stained with 0.1% crystal violet in water for 30 minutes. Supporting Information Figure S1 A: Quantification of flow cytometric analysis of fraction of mitotic cells treated with nocodazole for 18 hours. Cells were transiently transfected with indicated LAP-Mps1 constructs. D664A is an enzymatically inactive mutant. B: Box-and-whisker diagrams of time spent in mitosis of UTR-Mps1as cell treated with taxol in combination with increasing concentrations of different bulky PP1 inhibitors. ND; not determined. (Molecular structures displayed in right panel.) Found at: doi:10.1371/journal.pone.0010251.s001 (0.27 MB TIF) Figure S2 A: pT676-Mps1 and Mps1 immunoblot of UTR- Mps1WT, UTR-Mps1as and UTR-Mps1M602G cells treated with thymidine (thy) for 20 hours or with taxol and MG132 in combination with indicated inhibitors for 1 hour. 1NM, 1NM- PP1; 3MB, 3MB-PP1; SP, SP600125. B: Quantification of flow cytometric analysis of fraction of mitotic UTRM10, UTR- Mps1WT, UTR-Mps1as and UTR-Mps1M602G cells treated with taxol in combination with indicated inhibitors for 17 hours. C: Chromosome alignment in UTRM10, UTR-Mps1WT, UTR- Mps1as and UTR-Mps1M602G cells treated with MG132 for 50 minutes in combination with the indicated inhibitors. Graph displays percentages of cells with misaligned chromosomes. Found at: doi:10.1371/journal.pone.0010251.s002 (0.84 MB TIF) Figure S3 A–C: Time-lapse analysis by DIC microscopy of HCT- and UTR-derived cells treated with (A) taxol (HCT-derived cells) or STLC (UTR-derived cells) in combination with DMSO or 23dMB-PP1, or nocodazole (B, C) in combination with DMSO or 23dMB-PP1. Graphs in A display the time from NEB to the first References 1. Ruchaud S, Carmena M, Earnshaw WC (2007) Chromosomal passengers: conducting cell division. Nat Rev Mol Cell Biol 8(10): 798–812. 2. Musacchio A, Salmon ED (2007) The spindle-assembly checkpoint in space and time. Nat Rev Mol Cell Biol 8(5): 379–393. 3. Kops GJPL (2009) Dividing the goods: co-ordination of chromosome biorientation and mitotic checkpoint signalling by mitotic kinases. Biochem Soc Trans 37(Pt 5): 971–975. 4. Winey M, Goetsch L, Baum P, Byers B (1991) MPS1 and MPS2: novel yeast genes defining distinct steps of spindle pole body duplication. The Journal of Cell Biology 114(4): 745–754. 5. 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