Fig. 1: IL-17 induces cellular copper uptake through induction of STEAP4.

a Western blots analysis for copper trafficking-associated proteins from lysates of mouse colon epithelial organoids treated with different inflammatory cytokines for 8 h. Three independent experiments were done and representative blots were shown. Untr untreated. b IL-17 primed or unprimed mouse colon organoids were stained for STEAP4 (St4), E-cadherin (E-cad) and Ctr1 and integrin α. Scale bar, 100 µm. c SOD enzyme activity recovery from TTM inhibition in wild-type (WT) mouse organoids with or without 12 h of IL-17 priming. d SOD enzyme activity recovery in Steap4 wild-type (WT) and knockout (KO) organoids. e, f Fluorometric detection of intracellular copper in primary mouse colon organoids (e) or Ls174t cells (f). CF4 copper Fluor-4 probe, Ctrl CF4 control probe. Scale bar,100 µm. n = 5 biologically independent cell cultures. g Fluorometric detection of intracellular copper in Steap4 WT and Ctr1 knockdown cells. Cells were treated with or without IL-17 for 12 h and copper were supplemented into the medium for 6 h in all the groups. Scale bar,100 µm. n = 5 biologically independent cell cultures. h Atomic absorption spectroscopy measurement of copper content in Steap4 WT and KO Ls174t cells after copper addition. Cells were primed with or without IL-17 for 12 h and then 10 µM Cu(II) were supplemented into the medium of the indicated group for 6 h. n = 5 biologically independent cell cultures. i Copper content in cytosolic and mitochondrial fraction in Steap4 WT and KO cells, respectively. Cells were treated the same as described in panel (h). n = 5 biologically independent cell cultures. Results were normalized to cell numbers. All data show mean ± SEM. Two-tailed Student’s t test was performed for panel e (**P = 0.0057; ***P = 0.0004; ****P < 0.0001), f (***P = 0.0004; ****P < 0.0001), g (*P = 0.015; **P = 0.0027; ***P = 0.0004), h (*P = 0.0383; **P = 0.0022; ***P = 0.0009), i (***P = 0.0003; ****P < 0.0001; n.s., not significant). Data represent three independent experiments with similar results.