DUB

HEK-293 cells expressing Strep-CLASPIN were first arrested in S-phase with HU or in mitosis with nocodazole. Cycloheximide was then added to prevent further protein synthesis in the presence or absence of the USP9X inhibitor WP1130 and samples were harvested at the indicated times. A sample from the same cell line uninduced (U) was loaded as control.

Ubiquitination inhibitor PYR-41 and deubiquitination inhibitor PR-619 pretreated macrophages with or without Teuvincenone F (25 μM) for 2 h, following stimulated with LPS (100 ng/ml) for 30 min, and then subjected to immunoprecipitation with anti-NEMO antibody followed by immunoblot analysis with specific antibodies. Similar results were obtained from three independent experiments.

(e) MM.1S cells were treated with RRx-001 (1.25 μM), P5091 (3 μM) or RRx-001 plus P5091 for 12 h; protein extracts were analyzed for total DNMT activity using the EpiQuik DNMT activity kit (mean±s.d.; P<0.001; n=3). (f) MM.1S cells were treated with RRx-001 (1.25 μM), P5091 (3 μM) or RRx-001 plus P5091 for 12 h; protein lysates were then subjected to immunoblot analysis using antibodies specific against p21, HDM2, p53 or GAPDH.

IU1 treatment increased levels of peIF2a in both WT and S63del L. Sciatic nerves were treated ex vivo with 200 lM IU1 for 16 hr and the levels of p-eIF2a and eIF2a were analyzed by western blot. A Student’s t-test was performed between treated and untreated conditions of the same genotype

Representative results of Western blot showed the effects of LDN (10.0 μM) treatments on the expressions of LC3-II and LC3-I from Normoxia and OGD groups of cPKCc+/+ and cPKCc/ cortical neurons (n = 6 per group).

A USP14 inhibitor directly inhibits OSCC cell proliferation and triggers apoptosis. (A-D) OSCC cells were treated with indicated doses of b-AP15 for 24 h. (A) Cell proliferation was monitored by CCK8 assay. b-AP15 dramatically decreased cancer cells viability in a dose-dependent manner (p < 0.01). All values represented means ± SD of three independent experiments and each was performed in triplicate. (B, C) Flow cytometry analysis indicated that b-AP15 triggered significant apoptosis of OSCC cells (p < 0.01). Data were obtained in more than three independent experiments. (D) Apoptosis-related proteins were examined by western blot analysis. Inhibition of USP14 with b-AP15 induced a massive increase of ubiquitinated proteins, which then triggered apoptosis of cancer cells, activating caspase 3 to induce cleavage of caspase 3 and PARP.

HEK-293 cells expressing Strep-CLASPIN were first arrested in S-phase with HU or in mitosis with nocodazole. Cycloheximide was then added to prevent further protein synthesis in the presence or absence of the USP9X inhibitor WP1130 and samples were harvested at the indicated times. A sample from the same cell line uninduced (U) was loaded as control.

Ubiquitination inhibitor PYR-41 and deubiquitination inhibitor PR-619 pretreated macrophages with or without Teuvincenone F (25 μM) for 2 h, following stimulated with LPS (100 ng/ml) for 30 min, and then subjected to immunoprecipitation with anti-NEMO antibody followed by immunoblot analysis with specific antibodies. Similar results were obtained from three independent experiments.

(e) MM.1S cells were treated with RRx-001 (1.25 μM), P5091 (3 μM) or RRx-001 plus P5091 for 12 h; protein extracts were analyzed for total DNMT activity using the EpiQuik DNMT activity kit (mean±s.d.; P<0.001; n=3). (f) MM.1S cells were treated with RRx-001 (1.25 μM), P5091 (3 μM) or RRx-001 plus P5091 for 12 h; protein lysates were then subjected to immunoblot analysis using antibodies specific against p21, HDM2, p53 or GAPDH.

IU1 treatment increased levels of peIF2a in both WT and S63del L. Sciatic nerves were treated ex vivo with 200 lM IU1 for 16 hr and the levels of p-eIF2a and eIF2a were analyzed by western blot. A Student’s t-test was performed between treated and untreated conditions of the same genotype

Representative results of Western blot showed the effects of LDN (10.0 μM) treatments on the expressions of LC3-II and LC3-I from Normoxia and OGD groups of cPKCc+/+ and cPKCc/ cortical neurons (n = 6 per group).

A USP14 inhibitor directly inhibits OSCC cell proliferation and triggers apoptosis. (A-D) OSCC cells were treated with indicated doses of b-AP15 for 24 h. (A) Cell proliferation was monitored by CCK8 assay. b-AP15 dramatically decreased cancer cells viability in a dose-dependent manner (p < 0.01). All values represented means ± SD of three independent experiments and each was performed in triplicate. (B, C) Flow cytometry analysis indicated that b-AP15 triggered significant apoptosis of OSCC cells (p < 0.01). Data were obtained in more than three independent experiments. (D) Apoptosis-related proteins were examined by western blot analysis. Inhibition of USP14 with b-AP15 induced a massive increase of ubiquitinated proteins, which then triggered apoptosis of cancer cells, activating caspase 3 to induce cleavage of caspase 3 and PARP.