FIG. 4.

TSA-induced HIF-1α degradation is independent of ubiquitination. A. TSA degrades HIF-1α in E1-deficient cells. Normoxic Ts20 cells, expressing a temperature-sensitive E1 enzyme, were exposed to 39°C for 8 h in the presence or absence of TSA and in the presence of the proteasome inhibitor MG132 (5 μM) or the indicated concentrations of epoxomicin (Epox), the caspase inhibitor Z-VAD-FMK (50 μM), or the calpain inhibitor Z-Leu-Leu-Cho (40 μM). HIF-1α expression was analyzed by Western blotting. Tub, tubulin. B. E1-competent Ts20 cells respond to TSA. Ts20 cells cultured at 35°C were exposed to hypoxia (1% O₂) in the presence of TSA and the inhibitors described for panel A. C. 17AAG degrades HIF-1α in Ts20 cells. Normoxic Ts20 cells were exposed to 39°C for 8 h in the presence or absence of 17AAG (1 μM) and in the presence or absence of the proteasome inhibitor MG132 (5 μM).