Fig. 1

RNA sequencing analysis of glyoxalase 1 (GLO1)-depleted MDA-MB-231 cells highlights a pro-metastatic signature related to methylglyoxal (MG) stress. a GLO1 protein level in shNT, shGLO1#1 and #2 MDA-MB-231 cells. β-actin protein is used as loading control. Western blot is representative of three independent experiments. b Different steps of the high-throughput transcriptome analysis of GLO1-depleted MDA-MB-231 cells. c Correlation analysis of messenger RNA (Mrna) expression changes measured upon GLO1 depletion in shGLO1 (#1 and #2) clones when compared to shNT clone. Orange and red dots represent genes that are significantly differentially expressed (q < 0.05 and log2 fold change (FC) > 1). Red dots represent genes of the pro-metastatic signature. d Heatmap representing gene expression levels in the three replicates of all three conditions (shNT, shGLO1#1 and shGLO1#2 MDA-MB-231 cells) for genes significantly differentially expressed for both shGLO1 clones. Color scale corresponds to the expression Z-score across all samples calculated for each gene. e Significantly differentially expressed genes were analyzed for their gene ontology using ToppFun Suite software. Graph represents the percentage of genes from the input in the 12 most significantly affected biological processes for both shGLO1#1 and shGLO1#2 cells. Biological processes underlined in red are involved in the metastatic cascade. f All the genes listed in the biological processes related to metastasis were defined as the pro-metastatic gene signature of GLO1-depleted MDA-MB-231 cells. Down-regulated and up-regulated genes of the pro-metastatic gene signature are represented in a heatmap. Color scale corresponds to Z-score. g Tenascin C, Lumican and CD24 mRNA levels were assessed by qRT-PCR in GLO1-depleted MDA-MB-231 cells. h Tenascin C protein expression was detected in both cell-free deposited protein extracts and total cellular proteins of shGLO1 cells. i Cell surface CD24 protein level was assessed by flow cytometry. j TGFBI protein expression in GLO1-depleted MDA-MB-231 cells was assessed by western blot. All immunoblot data were quantified by densitometric analysis and normalized for ponceau red or β-actin. Numbers represent fold increase relative to the condition shown with bold number. Western blots are representative of two independent experiments. Data were analyzed using one-way analysis of variance followed by Dunnett post-hoc test and shown as mean values ± SEM of three independent experiments. *p < 0.05, **p < 0.01 and ***p < 0.001