Fig. 3: Ambra1 deletion induces an invasive phenotype in BrafV600E/+;Pten−/− mice. | Nature Communications

Fig. 3: Ambra1 deletion induces an invasive phenotype in BrafV600E/+;Pten−/− mice.

From: Loss of Ambra1 promotes melanoma growth and invasion

Fig. 3

a Upregulated processes (p < 0.01) and b GSEA (NES, normalized enrichment score; FDR, false discovery rate) in BPA−/− (n = 3) vs. BPA+/+ (n = 3) tumors. c Representative images and d histopathological evaluation of Picrosirius Red staining in BPA+/+ (n = 6) and BPA−/− (n = 6) tumors. e Differential expression of selected ECM-related genes in tumors (n = 3 each group, FC > 2, padj < 0.05.) f Gelatin degradation in AMBRA1-silenced SK-Mel-5 cells. F-Actin and nuclei were counterstained using Phalloidin and Hoechst. Images are representative. Each dot represents the average count per field ± SD (n = 3; **p = 0.0022; ****p < 0.0001, one-way ANOVA). g Wound closure area in AMBRA1-silenced SK-Mel-5. Quantification is shown as percentage ± SD vs. T0 at indicated times. White and yellow lines outline wound edge at T0 and at the times indicated. Images are representative (n = 4; **p = 0.0021 6 h; *p = 0.026 9 h; ****p < 0.0001 12 h, 18 h and 24 h, siAMBRA1#1 vs. siScr. n = 3; °p = 0.0234 6 h; °°°°p < 0.0001 9 h, 12 h, 18 h and 24 h, siAMBRA1#2 vs. siScr, two-way ANOVA). h Count of migrating cells in AMBRA1-silenced SK-Mel-5. Data are expressed as mean ± SD, images representative (n = 3; *p = 0.0289 siAMBRA1#1 vs. siScr, *p = 0.0454 siAMBRA1#2 vs. siScr, one-way ANOVA). WB in f–h indicate silencing efficiency. i RT-qPCR analyses of Cdh1, Cdh2, Zeb-1, Snai1, Vim, Fn1 and Ctnnb1 in BPA−/− (n = 4–8) and BPA+/+ (n = 4–8). Data normalized to L34 are expressed as fold-change (*p = 0.0158 Chd1; *p = 0.0417 Chd2; **p = 0.0096 Zeb-1; ***p = 0.0006 Snai1; *p = 0.0239 Vim; *p = 0.0458 Fn1, two-tailed unpaired t-test, BPA−/− vs. BPA+/+). j WB analyses of Cdh1, Cdh2, ß-Catenin, Vimentin and Snai1 in bulk tumors (n = 3 each group). k RT-qPCR analyses of CDH2, FN1, VIM and SNAI1 in AMBRA1-silenced SK-Mel-5. Data normalized to L34 are expressed as fold-change ± SD (n = 4; **p = 0.0045 CDH2; **p = 0.0022 FN1; *p = 0.0353 VIM; ***p = 0.0005 SNAI1, two-tailed unpaired t-test, siAMBRA1 vs. siScr, dashed line). l Representative (n = 4) WB analyses of CDH1, CDH2, ß-Catenin, VIM and SNAI1 in AMBRA1-silenced SK-Mel-5. Box-and-whisker plots in f, i represent minimum-to-maximum values. Top and bottom whiskers denote upper (Q3) and lower (Q1) quartiles, respectively. Boxes refer to interquartile ranges. Medians are denoted as horizontal line in the middle of the boxes.

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