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. 2018 Apr 18;9:1541. doi: 10.1038/s41467-018-04000-6

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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. 5 — Constitutive expression of the Ras-MAPK pathway causes overgrowths due to permanent activity of the JNK pathway after irradiation. Images of wing discs of genotypes: a control tubGal80^ts/+;hh-Gal4/+(n = 13), b UAS-Ras^V12/tubGal80^ts;hh-Gal4/+(n = 9) and c tubGal80^ts/UAS-Ras^V12;hhGal4/UAS-puc^2A (n = 11) 72 h after irradiation. Non-irradiated wing discs of genotypes: d control tubGal80^ts/+;hh-Gal4/+(n = 9) and e UAS-Ras^V12/tubGal80^ts;hh-Gal4/+ (n = 10). Anterior compartments are stained with Ci (green). JNK activity is monitored by Mmp1 (red). Staining with TOPRO facilitates delimiting the discs. Note in b the large overgrowth of the posterior compartment that contains UAS-Ras^V12, associated with ectopic permanent JNK activity. Much of the overgrowth is prevented (c) when JNK function is compromised by overexpressing puc. There is no Mmp1activity (red) in non-irradiated discs (d, e). f Quantification of the posterior compartment area (in percentage) over the total disc area in the genotypes: non-irradiated tubGal80^ts/+;hh-Gal4/+control (column: control No X-rays, n = 12), tubGal80^ts/UAS-Ras^V12;hh-Gal4/+(column: Ras^V12 No X-rays, n = 6), and irradiated tubGal80^ts/+;hh-Gal4/+control (column: control X-rays, n = 8), tubGal80^ts/UAS-Ras^V12;hh-Gal4/+(column: Ras^V12 X-rays, n = 9) and tubGal80^ts/UAS-Ras^V12;hhGal4/UAS-puc^2A (column: Ras^V12 puc X-rays, n = 34). There is a statistically significant size increase of the posterior compartment after irradiation when Ras^V12 is expressed, compared with non-irradiated discs and with irradiated controls (***p-value < 0.0004, Mann–Whitney U-test). However, if JNK activity is suppressed by overexpressing puc, the size of the posterior compartment significantly decreases (***p-value < 0.0001, Mann–Whitney U-test). Columns in the graph represent mean ± s.d. Images of wing discs of genotypes: g STAT-GFP/tubGal80^ts;UAS-Ras^V12/hh-Gal4 (n = 20) and h UAS-GFP,tubGal80^ts/UAS-Ras^V12;hh-Gal4/+(n = 9) 72 h after irradiation; g shows the ectopic expression of STATGFP (green) only in the posterior compartment (delimited by the lack of Ci in red) where Ras^V12 is expressed; h shows the ectopic expression of Wg (red) only in the posterior compartment (green) where Ras^V12 is active. Scale bars are 100 μm