Fig. 1

Subcellular localization of PR. a) Representative confocal images of U-2 OS cells stably expressing wild-type NPM1 (WT), NPM1 with deletions of the nuclear export (NESΔ) or nuclear localization (NLSΔ); NPM1 proteins were fused to GFP (green). Each NPM1 construct was co-transfected with PR (red) and nuclei stained with DAPI (blue). Automated microscopy and image analysis was used to quantify levels of PR in the nucleus (b) or cytoplasm (c) as a function of area (average intensity). Statistical significance was assessed by one-way ANOVA and post-hoc test between each experimental group; n = 2 biological groups, 9 fields per group, ****P < 0.0001; error bars are SEM, dots represent single cells. d) Representative images of U-2 OS cells stained with DAPI (blue) expressing GFP-NPM1 fusion proteins (green) with deletions of the nuclear localization (NLSΔ) or nucleolar (NuLSΔ) signals to confer cytoplasmic localization. Cells were immunolabeled with an antibody against PR (red). e) Nuclear/Cytoplasmic mean fluorescence (Y-axis) for n > 30 cells per mutant cell line (X-axis), n = 3 biological replicates. *P < 0.05; ***P < 0.0005 student’s t-test; error bars are SEM. f) Cartoon (created in PyMOL) with top and side perspectives of the NPM1 (PDB 4N8M) pentamer