AIF translocates to the nucleus upon treatment of cells with UCD38B.
(A) MDA-MB-231 cells were treated with 500 µM amiloride, or 250 µM UCD38B for 2 hours, and cells were fractionated into nuclear (N) and cytosolic (C) components. Fractions were immunoblotted for CREB and histone H3 as nuclear markers, GAPDH as a cytosolic marker, and AIF. (B) Bands from the blot depicted in (A) were digitally quantified, and the nuclear AIF as a fraction of the total AIF was plotted for each condition. (C) Cells were treated for 2 hours with DMSO control or 250 µM amiloride, UCD74A or UCD38B. Cells were then fixed and stained with DAPI to visualize the nucleus (blue) and antibodies to AIF (green), and examined by confocal fluorescence microscopy. Results depicted in each panel are representative of 2–3 independent experiments.