EGCG prevents proteasome inhibition and ER stress induction when combined with BZM and MG-262, but not MG-132. (A) LN229 cells were cultured for 30 hours with 20 nM BZM, 1.5 μM MG-132, or 20 nM MG-262 in the presence or absence of 20 μM EGCG. (Top) Percentage proteasome activity in total cell lysates was determined via 20S proteasome activity assay. (Bottom) The relative amount of polyubiquitinated proteins and the ER stress marker GRP78 was determined via Western blot analysis with specific antibodies against ubiquitin and GRP78, respectively. Western blot to actin was used as a loading control. (B) RPMI/8226 cells were cultured for 30 hours with 20 nM BZM in the presence or absence of 20 μM EGCG. (Top) Percentage proteasome activity from total cell lysates was determined via 20S proteasome activity assay. (Bottom) The relative amount of polyubiquitinated proteins, activated caspase-7 (cleaved casp-7), and cleavage of poly (ADP ribose) polymerase (PARP, which indicates ongoing apoptosis) in RPMI/8226 cells was determined via Western blot analysis with specific antibodies against ubiquitin, caspase-7, and PARP, respectively. (C) Highly purified proteasomes were treated with increasing concentrations of BZM in the presence or absence of 5 μM EGCG. Percentage activity was determined via 20S proteasome activity assay. Shown is percentage proteasome activity (mean ± SE; n ≥ 3), where activity in the absence of drug was set to 100%.