Figure 8
From: ER stress affects processing of MHC class I-associated peptides
Stability of cytosolic HEL and ER-retained HEL during ER stress. (A) Rate of synthesis of cytosolic HEL and ER-retained HEL. EL4/HEL-ER-SIINFEKL and EL4/HEL-Cyto-SIINFEKL cells were incubated in control conditions (4.5 mg/ml) or deprived of glucose for 17 hours and pulse-labeled with [35S]methionine/[35S]cysteine for 1 hour. Cell extracts were lysed and subjected to immunoprecipitation with anti-HEL antibody. Immunoprecipitated proteins were separated by SDS-PAGE and analyzed by fluorography. One representative experiment out of two is shown. (B) Stability of cytosolic HEL and ER-retained HEL. EL4/HEL-ER-SIINFEKL and EL4/HEL-Cyto-SIINFEKL cell lines were incubated in control conditions (4.5 mg/ml) or deprived of glucose for 18 hours. Then, 100 μg/ml of cycloheximide were added to inhibit protein synthesis and cell lysates taken at different times were immunoblotted against HEL or β-actin (used as loading control). One representative immunoblot out of three is shown. (C) Graph represents relative intensities of HEL (means and SD) from three independent experiments. *P < 0.05 when comparing control vs. no glucose conditions.