The position and dynamics of mRNA-free NXF1 in NPCs. (A) STED images showing the position of endogenous NXF1 relative to endogenous Nup358 (top) or Tpr (bottom) in control, WGA-treated, and DRB-treated cells. NXF1-10RA mutant (anti-Myc) was also tested. (B and C) FRAP experiment on U2OS cells expressing GFP-NXF1-10RA or GFP-NXF1 under untreated and DRB-treated conditions. Cells were imaged and photobleached in the nucleoplasm (B) or at the NPCs (C). Fluorescence recovery was tracked over time and plotted. See Materials and methods for the number of cells shown. Fit of average of three repeated experiments is presented. (D) Photoactivatable PAGFP-NXF1 (left) and PAGFP-NXF1-10RA (right) expressed in cells under WGA or Dbp5-DN mRNA export block conditions. A nucleoplasmic region was photoactivated, and the signal accumulating in the NPCs was tracked over time. Scale bar, 5 µm.