Fig. 4: USP32 increases BAG3 stability and deubiquitinates BAG3. | Oncogenesis

Fig. 4: USP32 increases BAG3 stability and deubiquitinates BAG3.

From: USP32 facilitates non-small cell lung cancer progression via deubiquitinating BAG3 and activating RAF-MEK-ERK signaling pathway

Fig. 4

A, B After transfection of overexpression plasmid Myc-USP32 or small interfering siUSP32 into H1299 and A549 cells for 48 h, afterwards they were treated with 50 μg/mL cycloheximide (CHX), and the cells were collected at the indicated times, and then subjected to Western blotting analysis of BAG3 protein levels. C–F NSCLC cells after overexpression and interference with USP32 were treated with or without MG132 for 6 h, respectively, and BAG3 levels were determined by immunoblotting. G In the presence of Myc-vector or Myc-USP32 plasmid, HA-labeled ubiquitin and Flag-BAG3 were co-transfected with H1299 cells for 48 h, after which the cells were harvested by adding MG132 treatment for 6 h, and cell lysates were immunoprecipitated experimentally with anti-Flag antibody, and ubiquitination was detected by HA antibody. H H1299 cells were co-transfected with Myc- USP32/Flag- BAG3/ HA-Ubiquitin-K48/HA-Ubiquitin-K63 plasmids and treated with MG132 for 6 h. Cell lysates were immunoprecipitated with anti-Flag antibody, and ubiquitylation was detected by HA antibody. *p < 0.05; **p < 0.01; ***p < 0.001.

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