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. 2018 Nov 8;9:4695. doi: 10.1038/s41467-018-07163-4

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© The Author(s) 2018

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. 3 — vSrc-transformed cells extrude in early mitosis. a A schematic model of cell cycle regulation. Highlighted in red are the molecules whose roles in cell extrusion were tested in this work. b The effect of Wee1 expression on vSrc-driven extrusion. Embryos were injected with the following constructs: dUAS:EGFP-vSrc and dUAS:EGFP-vSrc;Wee1. Data represent the number of tall and extruded cells divided by the total number of GFP-positive cells. Data are mean ± s.d. of three independent experiments (total number of embryos: n_Src = 21; n_Src,Wee1 = 27). *P < 0.05 (Student’s t-test). c The effect of constitutively active Cdc25 on vSrc-driven extrusion. Embryos were injected with the following constructs: dUAS:EGFP-vSrc and dUAS:EGFP-vSrc;CA-Cdc25. Data are mean ± s.d. of three independent experiments (total number of embryos: n_Src = 18; n_Src,Cdc25 = 21). *P < 0.05 (Student’s t-test). d Constitutively active Cdk1 rescues Wee1 inhibition of vSrc-driven extrusion. Embryos were coinjected with the following constructs: dUAS:EGFP-vSrc alongside either dKrt18:myr-Cherry, dKrt18:Cherry-Wee1 or dKrt18:Cherry-Wee1;CA-Cdk1. Data are mean ± s.d. of three independent experiments (total number of embryos: n_Src = 30; n = 29; n_{Src,Wee1,Cdk1} = 33). *P < 0.05 (Student’s t-test). e Time-lapse imaging of H2B-GFP in mitosis (top panel) and extrusion (bottom panel). Embryos were injected with the dUAS:myr-Cherry-vSrc;H2B-GFP construct. Movies were taken over 4 h and frames were extracted from a representative movie. t = 0 indicates either the beginning of mitosis (chromatin condensation) or the moment of extrusion. Scale bars, 10 µm. f Quantification of the chromatin volume (defined as the volume that H2B-GFP signal occupies in space) measured using the surface function of Imaris. The blue line follows chromatin volume change over time in a dividing EVL cell expressing myr-Cherry (averaged data from 3 cells), the green line—in a dividing vSrc cell (averaged data from 4 cells), the red line—in a extruding vSrc cell (averaged data from five cells). Error bars represent s.d.