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. 2020 Jul 3;11:3317. doi: 10.1038/s41467-020-17196-3

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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. 5 — a Scheme representing the three functions of the actin cytoskeleton during oriented divisions of epiblast cells⁵. In metaphase, epiblast cells form a polarized F-actin cap aligned with respect to the A/V axis. The formation of this F-actin cap depends on the small GTPase RhoA. The transmembrane protein Antxr2a binds to actin, and it is therefore recruited to and enriched at the actin cap to form a cap itself. Both caps colocalize. Antxr2a interacts then with an active form of RhoA to activate Diaphanous, which connects the mitotic spindle with the caps, rotating the spindle to align it with the caps and therefore with the embryonic axis. As a result, divisions of epiblast cells are oriented along the A/V embryonic axis. b, e Representative confocal images from a time-lapse movie of epiblast cells at metaphase (META) and anaphase (ANA) expressing the F-actin biosensor (Lifeact-mRFP) (b) or Antxr2a-eYFP (e) in control embryos (left panels) or sptlc1 morphants (right panels) from at least ten independent experiments. Yellow dashed line highlights F-actin or Antxr2a cap. Green dashed line, division plane. Animal pole, up. The embryonic axis is represented by a bi-directional arrow on the right. Scale bars: 10 µm. c Quantification of cortical F-actin accumulation at the cap of epiblast cells in control (n = 19 cells; green) and sptlc1 morphants (n = 15 cells; red) expressing Lifeact-mRFP. d, g Polar graphs showing the frequency distribution of the angle between the F-actin cap (d) or the Antxr2a cap (g) and the plane of the A/V embryonic axis in control (left graph) and sptcl1 morphants (right graph). n (number of cells analyzed) over N (number of embryos). In (d, g), χ² test was used (see Supplementary Table 1). f Antxr2a accumulation at the cap in epiblast cells expressing WT-Antxr2a-eYFP in control (n = 24 cells; green) or in sptlc1 morphants (n = 15 cells; red) and expressing AAC-zAntxr2a-eYFP in control morphants (n = 19 cells; blue). Data in (c, f) are represented as the mean value +/− SEM and an unpaired, two-tailed student’s t-test was used. DP (Division plane). Source data are provided as a Source Data file.