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. 2018 Dec 3;9:5129. doi: 10.1038/s41467-018-07638-4

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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. 6 — The disruption of Islr downregulates the Dvl2-mediated canonical Wnt signaling pathway in myoblasts. a Immunofluorescence staining for MyHC in control shRNA (shCtrl) and Islr shRNA stable (shIslr) C2C12 cells. N = 3 cell cultures in each group. Scale bar = 50 μm. The percentage of MyHC⁺ cells is shown on the right. b Immunofluorescence analysis of cell morphologies in shCtrl and shIslr C2C12 cells by staining live cells with DiI. N = 3 cell cultures in each group. Scale bar = 25 μm. c Heatmap of the changes in selected gene expression levels in shCtrl and shIslr C2C12 cells at G2 and D3 by RNA-seq. d Expression analysis of Wnt target genes and myogenic genes in shCtrl and shIslr C2C12 cells at 3 d in differentiation medium using qRT-PCR. e Western blot analysis of active β-catenin protein levels in nuclear lysates extracted from shCtrl and shIslr C2C12 cells after 7 d in differentiation medium. f Luciferase activity of TOP/FOP in shCtrl and shIslr C2C12 cells after 3 d in differentiation medium. g Western blot analysis of Islr and Dvl2 protein levels in shCtrl and shIslr C2C12 cells after 2 d in growth medium. h Western blot analysis of Islr and Dvl2 protein levels in shCtrl and shIslr C2C12 cells after 7 d in differentiation medium. i Western blot analysis of Dvl2 protein levels in primary myoblasts of control and Islr cKO mice after 3 d in differentiation medium. Error bars represent the means ± s.d. ^*P < 0.05, ^**P < 0.01, ^***P < 0.001; Student’s t test