Supplementary Figure 3.

Mechanistic aspects of iron-related dysregulation of protein secretion. (A) Interleukin 6 (IL6) mRNA (P = NS, paired t test, N = 3 per group). (B and C) Oxidative stress (*P < .05, both N = 3 per group). (B) Heme-oxygenase (HO-1) mRNA (P = .01, paired t test). (C) Glutathione peroxidase 1 (GPX1) mRNA (P = .049, paired t test). (D–I) Endoplasmic reticulum stress (all N = 3 per group). (D) Unspliced X-box binding protein (XBP1) mRNA (P = NS, paired t test). (E) Spliced XBP1 mRNA (P = NS, paired t test). (F) Immunoglobulin binding protein (BiP) mRNA (P = NS, paired t test). (G) Endoplasmic reticulum degradation-enhancing α-mannidose-like protein (EDEM) mRNA (P = NS, paired t test). (H) Activating transcription factor 4 (ATF4) mRNA (P = NS, paired t test). (I) CCAAT/enhancer-binding protein homologous protein (CHOP) mRNA (P = NS, paired t test). (J–M) Enrichment with signal peptide and exosome proteins. (J) Proportion of proteins down-regulated significantly by iron with signal peptide vs no signal peptide. (K) Proportion of proteins not down-regulated significantly by iron with signal peptide vs no signal peptide. (L) Proportion of proteins down-regulated significantly by iron with exosome secretion vs no exosome secretion. (M) Proportion of proteins not down-regulated significantly by iron with exosome secretion vs no exosome secretion. Of the 61 proteins down-regulated significantly, 62% (38 of 61) had signal peptide, whereas of the remaining proteins only 47% (129 of 277) had signal peptide. The 1-tailed Fisher exact test showed significant enrichment with signal peptide (P = .02) among the group down-regulated significantly. In contrast, there was no significant enrichment of the exosomal pathway (P = .51, 1-tailed Fisher exact test), because 15% (9 of 61) of the proteins down-regulated significantly and 14% (39 of 277) of the remaining secretome proteins had been reported previously in the high-confidence proteins from the EVpedia database.