Fig 1: Effects of MG132 treatment on GRP-78, PERK, ATF4, ATF6 and CHOP expression during hyperoxia exposure. a Western blot analysis of the AECII that were exposed to hyperoxia for 72 h, 30 g of total protein per well were loaded and subjected to Western blot analysis with corresponding antibodies shown on the left. The part of original data were presented in Additional file 1: Fig. S6. B–F Quantification of signals on the A, the results showed an increase in GRP78, PERK, ATF4, ATF6 and CHOP protein expression in hyperoxia group in comparison with normoxia group in 72 h. Values are presented as mean ± SD; ** P < 0.05 versus control
Fig 2: Effects of the protein expression of GRP-78, PERK, ATF4, ATF6 and CHOP in AECII after hyperoxia exposure. A Western blot analysis of the AECII that were exposed to 95% oxygen (hyperoxia) or not (normoxia) for 24, 48 and 72 h, 30 g of total protein per well were loaded and subjected to Western blot analysis with corresponding antibodies shown on the left. The part of original data were presented in Additional file 1: Fig. S3. B–F Quantification of signals on the Fig. 3A, the results showed an increase in GRP78, PERK, ATF4, ATF6 and CHOP protein expression in hyperoxia group in comparison with normoxia group in 24, 48, and 72 h. Values are presented as mean ± SD; ** P < 0.05 versus normoxia group
Fig 3: Isolation and identification of senescence-associated extracellular vesicles. EVs collected from ultracentrifugation were tested by (A) western blot. Cell lysates and EVs were tested with TSG101 antibody (1:2,000), GRP78 antibody (1:2,000), and ß-actin (1:2,000). (B) Nanosight analysis of EVs size and concentration. A histogram showing the relationship between particle size distribution and concentration. The X-axis indicates diameter of particles, and the Y-axis represents concentration of particles. TSG101, tumor susceptibility gene 101; GRP78, glucose-regulated protein 78; EVs, extracellular vehicles; CTRL, control.
Fig 4: ER stress and impaired protein homeostasis in vcpex3/ex3 embryos. (A) Loss of Vcp leads to an accumulation of ubiquitinated proteins in vcpex3/ex3 embryos compared to WT controls (N = 4). (B,C) Significantly increased Hspa5 protein levels suggest ER stress in vcpex3/ex3 embryos (N = 4; p = 0.0286; two-tailed t-test). Error bars indicate s.d.; * p < 0.05. (B,D) Total RPS6 protein levels are unchanged in vcpex3/ex3 mutant embryos (N = 4; p = 0.0286; two-tailed t-test), whereas phosphoRPS6 protein levels. Error bars indicate s.d.; ns, not significant. (B,E) are significantly increased, suggesting an activation of mTORC1 signaling (N = 4; p = 0.0286; two-tailed t-test). ß-Actin was used as a loading control. Error bars indicate s.d.; * p < 0.05. ns: not significant.
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