XB-IMG-147432
Xenbase Image ID: 147432
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FIGURE 1:. MgcRacGAP localizes to cell–cell junctions, and Mgc MO, MO + MgcS386E, and MO + R384A exhibit cytokinesis defects. (A) Protein diagram of Mgc showing location of Arg-384 and Ser-386 within the GAP domain, as well as mutants used in this study. (B) Internal and apical plane en face views and side views of endogenous Mgc localization in fixed embryos injected with either mChe-membrane alone (top) or MO + mChe-membrane (bottom) and then stained with an anti-Mgc antibody. Red arrowheads indicate midbodies; red arrow indicates the contractile ring in control embryo; yellow arrow indicates attempted division site in MO embryo; yellow arrowheads indicate apical localization of endogenous Mgc at cell–cell junctions. Scale bars, 20 μm. (C) Normalized intensity of endogenous Mgc at the contractile ring in control or MO embryos during early ingression measured from embryos that were fixed and stained with an anti-Mgc antibody. Control (number of cells, embryos, and independent experiments, respectively [n] = 10, 7, 4), MO (n = 11, 7, 4). (D) Peak intensity determined from line scans measuring the normalized Mgc intensity across bicellular junctions. Control (n = 20, 5, 4), MO (n = 18, 6, 3). (E) Images of fixed embryos injected with MO or MO + MgcWT or GAP-domain mutants. mChe-membrane marker (pseudocolored green) was used as an injection marker, and embryos were stained with DAPI (pseudocolored red). Scale bars, 40 μm. (F) Quantification of the percentage of cells that display multinucleation. The p values from Student's t tests were calculated from comparisons of MO cells with two nuclei or with three or more nuclei pairwise to the respective nuclei group for each indicated construct. The same analysis was conducted for pairwise comparison of MO + WT with two nuclei or with three or more nuclei to each indicated construct. Control (n = 266, 9, 3), MO (n = 184, 9, 6), MO + WT (n = 275, 9, 6), MO + S386A (n = 247, 8, 5), MO + S386E (n = 275, 7, 5), and MO + R384A (n = 202, 5, 4). (G) Quantification of cell width graphed as a box-and-whiskers plot. Control (n = 333, 6, 3), MO (n = 120, 9, 3), MO + WT (n = 208, 8, 3), MO + S386A (n = 264, 9, 3), MO+S386E (n = 207, 7, 3), and MO+R384A (n = 219, 10, 3). All error bars shown represent SEM unless otherwise noted. Image published in: Breznau EB et al. (2015) © 2015 Breznau et al. This image is reproduced with permission of the journal and the copyright holder. This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike license Larger Image Printer Friendly View |