O-GlcNAcylation enhances the permeability of the NPC. (A) Passive diffusion rate measurement. Left representative images: mEOS (top row) in the rectangular region (yellow box) inside the nucleus was selectively converted and continually imaged along with DNA (bottom row). Scale bar, 10 µm. Middle plot: A biexponential decay model was fitted to the nuclear intensity of converted mEOS to measure the passive permeation (perm) rate kp. Right box plot: Passive permeation rates for cells treated with DMSO, OSMI-4, or Thiamet-G (10 µM, 24 h). n ≥ 23 cells for each condition. (B) NPC counting. Top images: 3D SIM image of MAb414 immunostaining (red), Halo-H2A:JF646 (blue), and identified NPCs (white). Bottom box plots: Total number (left) and area density (right) of NPCs in cells treated with DMSO, OSMI-4, or Thiamet-G (10 µM, 24 h). n ≥ 17 cells for each condition. Scale bars, 10 µm (white) and 1 µm (yellow; magnified inset). n.s., P > 0.01; *, P < 1 × 10−2; ***, P < 1 × 10−4. (C) Proposed model for O-GlcNAc–dependent modulation of nuclear transport kinetics. Hydrophilic, bulky O-GlcNAc modifications hinder the cohesive interaction between FG repeats, thereby facilitating both passive and NTR-facilitated permeations of molecules through the NPC in both directions.
