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. 2017 Jul 28;8:150. doi: 10.1038/s41467-017-00209-z

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — High Aurora-B activity promotes KT attachment to MT-walls. a Representative images show high Aurora-B activity on lateral kinetochores. Monastrol treated cells were immunostained with antibodies against Tubulin, Aurora-BpThr232 and total Aurora-B (AurB) (left panel) or CREST antisera and antibodies against Tubulin and either HEC1pSer55 (middle panel) or HEC1pSer44 (right panel). Cropped images show lateral-kinetochores. Scale: 5 μm in uncropped images; 1 μm in cropped images. b Graphs show higher average signal intensities of HEC1pSer55 (left) and HEC1pSer44 (right) in lateral compared to end-on kinetochores as assessed from at least nine randomly chosen kinetochores from cells in a. CREST signal intensities are used as internal controls. c Experimental regime: Cells transfected with plasmid vectors encoding Mis12-INCENP-GFP were exposed to Monastrol and MG132 with either ZM447439 or DMSO (solvent control), prior to immunostaining. d Images of cells expressing Mis12-INCENP-GFP treated as in c and immunostained with antibodies against Tubulin, SKAP and GFP. White arrowheads in cropped images show ‘Lateral’ kinetochore lacking SKAP (upper panel) and ‘End-on’ kinetochore enriched with SKAP (lower panel). Scale: 5 μm in uncropped and 2 μm in cropped images. Boxed areas in a and d correspond to cropped images. e Graph shows percentage of lateral, end-on and detached kinetochores in Mis12-INCENP-GFP expressing cells treated as in c. Each circle represents value from one cell. Black horizontal bar marks average values from three independent experimental repeats. ‘*’ indicates statistically significant difference on the basis of P-values obtained using unpaired Student’s t-test