Super-resolved 3D Tracking of Cargo Transport Through Nuclear Pore Complexes

Nuclear pore complexes (NPCs) embedded within the nuclear envelope (NE) mediate rapid, selective, and bidirectional traffic between the cytoplasm and the nucleoplasm. Deciphering the mechanism and dynamics of this process is challenged by the need for high spatial and temporal precision. We report here a multi-color imaging approach that enables direct 3D visualization of cargo transport trajectories relative to a super-resolved octagonal double-ring structure of the NPC scaffold. The success of this approach is enabled by the high positional stability of NPCs within permeabilized cells, as verified by a combined experimental and simulation analysis. Hourglass-shaped translocation conduits for two cargo complexes representing different nuclear transport receptor (NTR) pathways indicates rapid migration through the permeability barrier on or near the NPC scaffold. Binding sites for cargo complexes extend over 100 nm from the pore openings, consistent with a wide distribution of the FG-polypeptides that bind NTRs.