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. 2017 Jan 10;3:16049. doi: 10.1038/celldisc.2016.49

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Copyright © 2017 The Author(s)

This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/

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Aurora-A promotes the association of Aurora-B with Haspin and Plk1. (a) Flag-tagged kinase deficient (KD)-Aurora-B was expressed and immunoprecipitated (IP) in 293T cells. glutathione S-transferase (GST)-fused Haspin-N proteins were preincubated with human recombinant Aurora-A (rhAurA) with or without ATP and were mixed with KD-Aurora-B on FLAG-Beads. Pull-down assay was conducted with FLAG beads, and Haspin-N was detected with GST antibody. (b) A statistical analysis of three experiments is shown for a. (c) Flag-tagged Aurora-B was co-expressed with green fluorescent protein (GFP)-Haspin-N, Plk1 or INCENP. Two hours before harvest, cells were treated with 0.5 μM nocodazole, and Aurora-A inhibitor or dimethyl sulfoxide (DMSO) was added as indicated. Co-IP was performed with FLAG beads. (d) The IP-binding ratio (GFP: FLAG) for precipitates in c is quantified. Three or four independent experiments were performed. Values represent mean±s.e.m. **P<0.01; ***P<0.001; NS: not significant (Student’s t-test). (e) A model is shown to illustrate the function of Aurora-A in promoting Haspin-H3T3-ph-CPC (chromosomal passenger complex) feedback loop in early mitosis. See the Discussion section for a detailed description. EGFP, enhanced GFP; SAC, spindle assembly checkpoint; WB, western blotting.