Fig 2. HIF-2α stabilizes B-Myb.

(A) Control- or 3×FLAG-pVHL-expressing 786-O cell lines were cultured in the presence or absence of CoCl₂ (400 μM) for 7 h, and cell lysates were immunoblotted with anti-B-Myb, anti-HIF-2α, or anti-pVHL antibodies. Forced expression of 3×FLAG-pVHL downregulates HIF-2α and B-Myb protein levels under normoxic conditions, and CoCl₂ treatment rescues HIF-2α and B-Myb protein levels. Hsp90 was used as loading control. Representative data from three independent experiments are shown. (B) Knockdown of HIF-2α suppresses the accumulation of B-Myb under hypoxic-like conditions. Control or pVHL-expressing 786-O cell lines were transfected with non-specific or HIF-2α-targeting siRNA and cultured for 2 days, and cell lysates were immunoblotted with anti-B-Myb, pVHL, or anti-HIF-2α antibodies. Cells were incubated in the presence or absence of CoCl₂ (400 μM) for 7 h before they were harvested. Hsp90 was used as a loading control. Representative data from three independent experiments are shown.