Fig. 4: USP7 stabilizes KDM4A proteins via deubiquitination and blockade of the proteasome-mediated degradation pathway. | Cell Death & Disease

Fig. 4: USP7 stabilizes KDM4A proteins via deubiquitination and blockade of the proteasome-mediated degradation pathway.

From: USP7 promotes chemotherapy resistance and DNA damage response through stabilizing and deubiquitinating KDM4A in bladder cancer

Fig. 4: USP7 stabilizes KDM4A proteins via deubiquitination and blockade of the proteasome-mediated degradation pathway.

A T24 and 5637 cells were infected with lentiviral control versus USP7-specific shRNA (sh#1) expression vectors. Upon 24 h treatment with MG132 (20 μM) versus CQ (20 μM), total proteins were normalized for WB assay. B T24 cell were infected with lentiviral control versus USP7-specific shRNA (sh#1) expression vectors. Next, cells were treated with MG132 (20 μM) for 12 h and cell lysates were subjected to IP with KDM4A antibody for WB assay. C similarly to (B), T24 cell were infected with lentiviral vectors (empty vector control, HA-USP7-WT, versus HA-USP7-mutant). Upon 12 h treatment with MG132 (20 μM), the cell lysates were subjected to IP with the KDM4A antibody for WB assay. D T24 cell were pre-treated with DMSO versus USP7 inhibit (P5091, 30uM) for 24 h, and then treated with MG132 (20 μM) for additional 12 h. The cell lysates were subjected to IP with the KDM4A antibody for WB assay. E T24 cell were infected with lentiviral control versus USP7-specific shRNA (sh#1) expression vectors. Cells were next treated for indicted time course with cycloheximide (CHX, 5 μM) and total proteins were normalized for WB assay. F Similarly to (E), T24 cells were pre-treated with DMSO versus USP7 inhibitor (P5091, 30uM) for 24 h. Cells were next treated for indicted time course with cycloheximide (CHX, 5 μM) and total proteins were normalized for WB assay. G T24 cells were transfected with control (empty vector) versus HA-USP7-WT or HA-USP7-mutant expression vector. Cells were next treated for indicted time course with cycloheximide (CHX, 5 μM) and total proteins were normalized for WB assay. H 12 pairs of bladder cancer tissues (T) and normal bladder mucosal epithelium (N) were freshly collected. Total proteins were normalized for WB analysis. I Representative images of IHC staining for KDM4A and USP7 proteins in 37 cases of bladder cancer versus 10 cases of normal bladder tissues. J, K Statistics analysis of USP7 IHC staining in 37 cases of bladder cancer versus 10 cases of normal bladder tissues (J) and its correlation with KDM4A expression levels in cancer cases (R = 0.512, p = 0.0017, Spearman).

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