BAG3 and BAG6 differentially affect the dynamics of stress granules by targeting distinct subsets of defective polypeptides released from ribosomes

摘要

BAG6 colocalizes with DRiPs and participate in their clearance. a HeLa cells were treated with OP-puro (25 μM) for 1 h or with OP-puro (25 μM) and MG132 (20 μM) for 3 h. Cells were fixed and then subjected to immunofluorescence using specific antibodies for BAG3 or BAG6. OP-puro-labeled nascent chains were visualized by click chemistry with Alexa594-Azide (DRiPs). Scale bars = 10 μm. b HeLa cells were lipofected for 72 h with siRNA non-targeting control or against BAG6. Then, cells were incubated with puromycin 10 μg/ml for 45 min, followed by recovery in drug-free medium for 13 h. Proteins were extracted from the treated cells and subjected to SDS-PAGE, followed by immunoblotting using the indicated antibodies. Quantitation of the percentage of puromycilated peptide chains is reported. n = 3, +/− SEM, p = 0.002