Fig. 6: PP2A-B56 is needed to maintain stable kinetochore-microtubules attachment at metaphase.

A Schematic illustrating known and potential contributions of PP2A-B56 in stabilizing kinetochore-microtubule attachments during mitosis. B Evaluating the effects on chromosome alignment in HeLa-FRT cells with/without directSLiMs^LIE9/AAA expression and treated with vehicle or rapamycin. Top panel: protocol used to visualise chromosome alignment in fixed samples (see Methods for details). Bottom panel: graph showing mean frequencies of chromosome misalignments ( ± SEM) of 2 experiments, 100 cells quantified per condition per experiment. Individual data points of the bar charts are provided in Source Data file. C Evaluating the timings of chromosome misalignments during a metaphase arrest, in HeLa HistH1H4C-TagGFP2 cells expressing the directSLiMs^{LIE1/LIE9/AAA} and treated with vehicle or rapamycin. Top panel shows protocol and bottom panel shows timings of chromosome misalignments from 2 experiments, 50 cells quantified per condition per experiment. D Example images of the misalignment timings shown in C. E Kinetochore intensity of NDC80-pS55 in nocodazole-arrested HeLa-FRT cells with directSLiMs^{LIE1/LIE9/AAA} treated with vehicle or rapamycin. 60 cells quantified from 4 repeats. F Evaluating the timings of chromosome misalignments during a metaphase arrest, in HeLa HistH1H4C-TagGFP2 cells expressing the directSLiM^LIE1 and treated with/without rapamycin and the AurBi ZM-447439, as indicated. Top panel shows protocol and bottom panel shows timings of chromosome misalignments from 2 experiments, 50 cells quantified per condition per experiment. G Example images of the misalignment timings shown in F. H Fixed chromosome alignment assays. Top panel shows protocol and bottom panel mean mean frequencies of chromosome misalignments ( ± SEM) of 2 experiments, 100 cells quantified per condition per experiment. Individual data points of the bar charts are provided in Source Data file. I Kinetochore intensities of SKAP from cells treated as in H. SKAP is a marker of mature end-kinetochore microtubules attachments²². 20 cells quantified from 2 repeats. Source data are provided as a Source Data file. Data information: Violin plots show the distributions of misalignment timings between cells. For each violin plot, each dot represents an individual cell, the horizontal line represents the median and the vertical one the 95% CI of the median, which can be used for statistical comparison of different conditions (see Methods). Scale bar: 10 μm.