Fig. 4

RPS15 overexpression promotes the translation of core E2F and p38 MAPK pathway proteins. a Schematic illustrating the ribosome profiling of KYSE30 cells (Con) and RPS15-overexpressing KYSE30 cells (RPS15). b GO analysis (upper) and KEGG-pathway analysis (lower) of transcripts preferentially bound by ribosomes in the RPS15 group. The most enriched ribosomal/translational GO gene sets and associated FDR values are shown, which were mainly enriched in translation and MAPK signaling pathways. c Heat map of the log2 fold-change of the RPS15 group relative to that of the control group for ribosome protein genes for mRNA expression, ribosome-protected fragments (RPF), and translation efficiency (TE). d Heat map of the log2 fold-change of the RPS15 group relative to that of the control for each gene of the E2F pathway and p38 MAPK pathway target-gene set for mRNA expression, RPF, and TE. Genes were categorized according to their GO biological process functions in the relative order of the TE fold-change. e Scatter plot of the translational efficiency of individual RP genes (right panel) and the hallmark p38 MAPK-target gene set transcripts (left panel). The x-axis represents the log2 fold-change in RNA-seq and the y-axis represents the log2 fold-change in ribosome profiling. f Expression of the hallmark p38 MAPK-target gene set in RPS15-overexpressing KYSE30 cells (left) and KYSE450 cells (right) determined by Western blot. g Western blot analysis results of expression of the hallmark p38 MAPK-target gene set in KYSE150 cells (left) and KYSE510 cells (right) with or without RPS15 knockout. h, i Statistical analyses of Boyden Chamber migration and invasion assays for KYSE30 cells (h) and KYSE450 cells (i) with stably transfected control vector or RPS15-overexpressing vector with or without SB203580 treated for 24 h. The statistical analysis results are shown. j Growth curves measured using Incucyte live-cell analyses of KYSE30 cells stably transfected with control vector (blue) or RPS15-overexpressing vector (red) and treated with SB203580 or SB203580 combined DDP for 72 h. k Growth curves measured using Incucyte live-cell analyses of KYSE450 cells stably transfected with control vector (blue) or RPS15-overexpressing vector (red) and treated with SB203580 or SB203580 combined DDP for 72 h. Data were analyzed using unpaired t-tests. *P < 0.05, **P < 0.01, and ***P < 0.001. l Western blot analysis to detect expression of the hallmark MAPK-target gene set in KYSE30 cells (upper) and KYSE450 cells (lower) stably transfected with control vector or RPS15-overexpressing vector and treated with SB203580 or DDP for 2 days. Con control, Veh vehicle