Characterisation of the passive permeability barrier of nuclear pore complexes

Abstract

Nuclear pore complexes (NPCs) restrict uncontrolled nucleocytoplasmic fluxes of inert macromolecules but permit facilitated translocation of nuclear transport receptors and their cargo complexes. We probed the passive barrier of NPCs and observed sieve-like properties with a dominating mesh or channel radius of 2.6 nm, which is narrower than proposed earlier. A small fraction of diffusion channels has a wider opening, explaining the very slow passage of larger molecules. The observed dominant passive diameter approximates the distance of adjacent hydrophobic clusters of FG repeats, supporting the model that the barrier is made of FG repeat domains cross-linked with a spacing of an FG repeat unit length. Wheat germ agglutinin and the dominant-negative importin b 45-462 fragment were previously regarded as selective inhibitors of facilitated NPC passage. We now observed that they do not distinguish between the passive and the facilitated mode. Instead, their inhibitory effect correlates with the size of the NPC-passing molecule. They have little effect on small species, inhibit the passage of green fluorescent proteinsized objects 410-fold and virtually block the translocation of larger ones. This suggests that passive and facilitated NPC passage proceed through one and the same permeability barrier.

NRAS isoforms differentially affect downstream pathways, cell growth, and cell transformation

“…As isoforms 3 and 5 are very small, we hypothesized that they may translocate into the nucleus despite not having nuclear localization signals (15). Indeed, confocal microscopy of COS-7 cells expressing each of the isoforms revealed that isoforms 1, 2, and 4 were localized entirely in the cytoplasm, but, strikingly, isoforms 3 and 5 were also detected in the nucleus (Fig.…”

How to operate a nuclear pore complex by Kap-centric control

“…40 Similar bimodal kinetics has also been reported for GFP-Kapb1 42 in permeabilized cells and mRNA export in living cells. 57 Interestingly, it was found that additions of exogenous Kapb1, 40 as well as other receptors 58 further reduced NPC permeability against passive cargoes, thereby recapitulating the role of Kaps as bona fide constituents of the NPC. Moreover, Yang and Musser, 36 as well as Timney et al 45 showed in vitro and in vivo respectively, that increasing amounts of Kaps expedited cargo import rates, rather than slowing it down.…”

Nucleocytoplasmic Distribution Is Required for Activation of Resistance by the Potato NB-LRR Receptor Rx1 and Is Balanced by Its Functional Domains

“…Proteins with a molecular mass higher than 40 kD (or a Stokes radius over 2.7 nm, as dimensions are more important than molecular mass) are limited in their passive passage through the nuclear pore complex (Paine et al, 1975;Merkle, 2003;Mohr et al, 2009). Therefore, it is likely that the transport of NB-LRR resistance proteins in and out of the nucleus needs to be actively facilitated.…”