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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. 2 — Constitutive Aurora-B activity at the outer-kinetochore allows lateral attachments but disrupts end-on conversion. a Single-plane time-lapse images of Z-stacks show the fate of a lateral KT (red) attached to MT (green). Time-lapse images of HeLa cells expressing mKate2-Tubulin and either Mis12-INCENP-GFP or Mis12-INCENP (TAA)-GFP mutant were acquired in the presence of Monastrol. Yellow arrows mark shrinking lateral K-fibre and white arrows show the synchronous movement of the KT with MT-end. White arrowheads mark the KT tracked. For three final time-points (left panels) ‘intensified’ images are included to highlight the K-fibre. Scale bar: 2 μm. b Graphs show the percentage of lateral (left) or end-on (right) kinetochores that transitioned into other attachment states (D-detached, L-lateral and E-end-on) in time-lapse movies as in a. c Graph shows the difference between ΔMT (change in MT-end position) and ΔKT (change in KT position) through time, used to define synchronous or asynchronous movements. Values less than 0.25 μm (marked by dashed line), sustained for at least 30 s, are indicated as synchronous (green) movement between the KT and MT-end. All other values are indicated as asynchronous movements of the KT and MT-end. d Graph shows the time spent by lateral kinetochores on MT-walls, before changing into another attachment state, in cells expressing either Mis12-INCENP-GFP or Mis12-INCENP (TAA)-GFP. Red and black bars mark mean-time and SD values, respectively, across kinetochores in three experiments. Error bars in b and c are SEM values across experiments b or across kinetochores c. ‘*’ and ‘#’ indicate statistically significant and insignificant differences, respectively, on the basis of P-values obtained across three experimental repeats using a Proportion test b or unpaired Student’s t-test d. e Cartoon shows how Aurora-B at the outer-kinetochore disrupts the end-on conversion process by detaching lateral kinetochores prior to end-on conversion. Aurora-B at the outer-kinetochore does not interfere with the capture or maintenance of kinetochores on microtubule walls. Reducing Aurora-B activity allows the formation of mature end-on attachments