Fig. 7

Functional investigation of the candidate target gene EGLN3. a Tumor-exclusive EGLN3-derived peptides detected by HLA ligandomics in patient cohort 1. The number of positive tumors and the HLA restriction of the respective peptides are given. b Cellular metabolites regulated by EGLN3 knockdown (EGLN3) in the 786-O kidney carcinoma cell line (ctr. siRNA 1, cells transfected with the non-targeting siRNA pool 1; UT, untreated cells). Metabolites were identified by untargeted metabolomics analysis. c Legend for the graphs in the figure. Untreated, untreated cells; ctr. siRNA 1/2, cells transfected with two different non-targeting siRNA pools; EGLN3, EGLN3 knockdown cells. d Percentage of cells in S-phase. The A498 and 786-O cell lines were used in the experiments. The effect of EGLN3 knockdown was non-significant (p ≥ 0.05). e Percentage of apoptotic A498 and 786-O cells. The asterisks mark significant effects with p < 0.05. f Profiles of extracellular acidification rate (ECAR) in A498 and 786-O cells treated with glucose, oligomycin, and 2-DG (Glycolytic Stress Test, Agilent Technologies). The x-axis shows the measurement cycle. g Effect on glycolysis in A498 and 786-O cells. The asterisks mark significant differences (p < 0.05), whereas ns indicates non-significant differences. h Profiles of oxygen consumption in A498 and 786-O cells treated with oligomycin, FCCP, and rotenone/antimycin A (Mito Stress Test, Agilent Technologies). i Effects on ATP production, basal respiration, and maximal respiration. The asterisks mark significant differences (p < 0.05), whereas ns indicates non-significant differences. j Brightfield images of spheroids formed by EGLN3 knockdown and control cells of the Caki1 and A498 cell lines. k Cell viability of Caki1, A498, and 786-O cells assessed by the WST-1 proliferation reagent