Molecular mechanism of HIF-1α protein stabilization by CuCl₂. (A) Hep3B hepatoma reporter cells (subline HRB5) were treated with or without 100 μM CuCl₂ for 24 hours in the presence or absence of the indicated kinase inhibitors under normoxic conditions. Shown are mean induction factors of luciferase activity ± SD of n = 3 independent experiments. (B) Hep3B cells were kept under hypoxic conditions in the presence or absence of 150 μM CuCl₂ before the addition of 20 μM cycloheximide (CHX) to block translation. Following reoxygenation for the indicated time periods, HIF-1α levels were determined by immunoblotting as in Figure 2. Subsequent detection of β-actin served as control for equal loading.