(A) Protein levels of HIF1α, HIF2α, FIH and PHD2
in U2OS cells upon hypoxia (1% O2) treatment at the indicated time
points. GAPDH was used as a loading control.
(B) Immunofluorescence staining of FIH (green) in U2OS cells
upon hypoxia (1% O2) at the indicated time points. TO-PRO-3 (blue)
was used to stain nuclei. Scale bar: 20 µm.
(C) Protein levels of FIH and HIF1α from cytoplasmic or
nuclear fractions in U2OS cells in normoxia or hypoxia (1% O2, 3
hours). β-tubulin and PARP were used as loading controls for the
cytoplasmic and nuclear fractions respectively. Figures beneath lanes 2 and 4
are relative intensities of nuclear FIH in normoxia and hypoxia. Note that
different quantity of cytoplasmic versus nuclear extract were loaded.