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. 2020 May 27;11:2652. doi: 10.1038/s41467-020-16488-y

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© The Author(s) 2020

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 — a Live imaging of Ndc80^f/f (control) and Ndc80^f/f Zp3-Cre (Ndc80-deleted) oocytes expressing EGFP-Map4 (microtubules, green) and H2B-mCherry (chromosomes, magenta). Spindle shapes were reconstructed in 3D. Asterisks show chromosomes that fell off the spindle. Anaphase I was defined based on characteristic chromosome and spindle dynamics (see Methods). Three independent experiments were performed. See also Supplementary Movie 1, 2, and 3. b Establishment of spindle bipolarity requires Ndc80 in MI. Spindle shapes in 3D were categorized based on the aspect ratio, surface irregularity, and stability (see Methods). Spindles that maintained a bipolar state for 1 h or longer with no collapse prior to anaphase I onset were categorized as stable bipolar spindles. c Ndc80 is required for preventing aneuploidy in eggs. The number of chromosomes were counted at metaphase II (n = 28, 28 oocytes from three independent experiments). Bars represent mean. See also Supplementary Movie 4. d The Ndc80 complex and Ndc80ΔN/Nuf2ΔN construct. In the Ndc80 complex, Ndc80 and Nuf2 directly bind to microtubules via the head domains at their N-termini, and to Spc24 and Spc25 at their C-termini. Ndc80 contains a loop domain in the central region, which recruits microtubule-binding proteins. Ndc80ΔN (a.a. 461–642) and Nuf2ΔN (a.a. 276–463) contain neither the head nor loop domain but retain the Spc24- and Spc25-binding domains. e Ndc80ΔN/Nuf2ΔN rescues spindle defects. Ndc80^f/f Zp3-Cre oocytes coexpressing Ndc80ΔN and Nuf2ΔN (Ndc80ΔN/Nuf2ΔN) and those expressing full-length Ndc80 (Ndc80-WT) were monitored for spindle formation. EGFP-Map4 (microtubules, green) and H2B-mCherry (chromosomes, magenta) signals at metaphase I (5.5 h after NEBD) are shown. Spindle shapes were reconstructed in 3D at metaphase I (5.5 h after NEBD) and categorized based on the aspect ratio and surface irregularity (n = 23, 23, 21 oocytes from three independent experiments). Mean +/− SD are presented. **p = 0.0026; ***p = 0.0002; by two-tailed unpaired Student’s t-test. See also Supplementary Movie 5. Scale bars, 10 μm.