Fig. 5: Comparison of ER stress induction methods. | Nature Communications

Fig. 5: Comparison of ER stress induction methods.

From: Simultaneous proteome localization and turnover analysis reveals spatiotemporal features of protein homeostasis disruptions

Fig. 5

a Bar charts showing activation of known ER stress markers upon tunicamycin (Tunic.) treatment for 8 h and 16 h. X-axis: ER stress markers; y-axis: expression ratio (n = 6 normal AC16; n = 3 tunicamycin). *: P < 0.01; **: P < 0.001; ***: P < 0.0001; multiple-testing corrected (FDR adjusted) P (two-sided). Error bars: expression ratios ± s.d. b Histograms of the determined log10 protein turnover rates in control and tunicamycin-treated cells (n = 3). c Boxplot showing the log2 turnover rate ratios in tunicamycin over normal AC16 cells for proteins that are localized to the ER (T) or not (F); Golgi apparatus (GA), or the lysosome. P values: two-tailed t-test. Center line: median; box limits: interquartile range; whiskers: 1.5x interquartile range; n = 286, 2234, 62, 2458, 139, 2381 proteins over 3 biological independent samples. A Bonferroni corrected threshold of 0.05/13 is considered significant. d Gene set enrichment analysis (GSEA) of turnover rate ratios in tunicamycin treatment. Color: multiple-testing corrected (FDR adjusted) P in GSEA (two-sided) x-axis: normalized enrichment score (NES). Size: proteins in the gene set. e Example of best fit curves in the first-order kinetic model at the protein level between normal (gray), tunicamycin (blue) and thapsigargin (red) treated AC16 cells showing known ER stress markers with elevated turnover in both ER stress inducers (HSPA5, HSP90B1, and PDIA4) as well as stress response proteins with elevated turnover only in tunicamycin (PDIA3, DNAJB11, NIBAN1). Point: best-fit k; line: first-order kinetic curve of k; bands: fitting s.e. f Alluvial plot showing the migration of ER, GA, and peroxisome/endosome proteins toward the lysosome (left). On the right, the ultracentrifugation fraction profiles of the differentially localized proteins RRBP1, FKBP11, GANAB, MANF, IKBIP, and P3H1 are shown that are targeted toward the lysosome in both tunicamycin and thapsigargin treatment (BANDLE differential localization probability > 0.95). Numbers in boxes are the BANDLE allocation probability in each condition (n = 3).

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