The Nuclear Pore Complex as a Transport Machine

Abstract

Nucleocytoplasmic Transport and the NuclearPore Complex Unlike their prokaryotic counterparts, eukaryotic cells separate the nuclear synthesis of DNA and RNA from cytoplasmic protein synthesis with a barrier termed the nuclear envelope (NE).1 The NE is perforated by large proteinaceous assemblies, called nuclear pore complexes (NPCs), which act as the sole gatekeepers controlling the exchange of material between the two locales (reviewed in Ref. 1). NPCs are freely permeable to small molecules (such as water and ions), but they restrict the movement of larger molecules (such as proteins and RNAs) across the NE. To overcome this barrier, macromolecules carry specific signals that allow them to access the nucleocytoplasmic transport machinery of the cell. In this way the cell ensures that only selected macromolecules can travel between the nucleus and cytoplasm (reviewed in Ref. 2).Operationally, NPCs are composed of proteins called nucleoporins (or Nups) forming the stationary phase for nucleocytoplasmic exchange, whereas the mobile phase consists of soluble transport factors and their cargoes. As nucleocytoplasmic transport is driven by a series of specific interactions between components of both phases, it is frequently difficult to determine which proteins are permanent constituents of the NPC. Nevertheless, to understand how transport occurs, we must characterize the players in both phases and understand how their interplay leads to the coordinated vectorial exchange of macromolecules across the NE. In this review we focus on recent results that shed light on how some of these proteins interact to contribute to the elaborate NPC architecture and its function as a transport machine. Architecture of the NPCNPCs from different organisms share a common fundamental architecture (3). These similarities likely provide clues as to the key features common to a functioning transport machine. The NPC is a large octagonally symmetric cylindrical structure. In yeast it estimated to be ϳ50 MDa, whereas in metazoans it is over twice this mass (3,4). Given that a ribosome at 4 MDa contains ϳ80 proteins, it might be expected that the NPC would contain hundreds of different nucleoporins. However, it has recently been shown that the yeast NPC contains only ϳ30 different proteins and the vertebrate NPC contains perhaps a few more (5). This raises the question of how such a large complex can be constructed from so few component parts. The answer appears to lie in the symmetry of the structure (Figs. 1-3). The NPC is comprised of a cylindrical core from which numerous peripheral filaments project toward the nucleus and cytoplasm (reviewed in Ref. 6) (Figs. 1 and 2). The remarkable symmetry of the NPC is most apparent in the central core. Not only is it composed of eight identical spokes, but each spoke is also seemingly mirror symmetrical both in a plane parallel to the NE and in a perpendicular plane running through the cylindrical axis. As predicted from this symmetry, all nucleoporins examined thus far are present in multiple...

Visualizing single molecules interacting with nuclear pore complexes by narrow-field epifluorescence microscopy

“…We found that the geometrically averaged noise works well for our purposes, i.e. : (2) where I o and I b are the average measured intensities in the presence and absence of the fluorescent object, respectively, and σ o is the standard deviation of I o . This definition does not require σ o to be well-defined (if ill-or un-defined, σ o can be approximated with σ b ), and the limiting noise level at low signal is σ b , as expected.…”

“…First, high localization precision is necessary. NPCs are large (∼60-120 MDa), octagonally rotationally symmetric structures comprised of at least 30 different nuclear pore proteins (Nups) [1][2][3]. The pore itself is ∼90 nm in length and is ∼50 nm wide at its narrowest point.…”

“…An extensive network of thousands of phenylalanine-glycine (FG) repeat motifs located on almost half the Nups (FG-Nups) provides binding sites for importins and exportins and thereby provide the NPC interaction sites for cargo complexes [2]. These FG-Nups are distributed throughout the NPC structure on both the cytoplasmic filaments and nuclear basket, and in the pore itself [3,10,11].…”

Regulation of Smad activities

“…The transport of macromolecules across the nuclear envelope is mediated by the nuclear pore complex (NPC), an assembly of multiple copies of over 30 proteins termed nucleoporins [42,43]. Translocation across the nuclear envelope entails three steps: anchorage to the NPC, migration through the NPC and release into either cyto-or nucleoplasm [44][45][46].…”

Nup214-Nup88 Nucleoporin Subcomplex Is Required for CRM1-mediated 60 S Preribosomal Nuclear Export

“…A large subset of nucleoporins contains long phenylalanine-glycine dipeptide-containing domains (FG repeats), which are thought to form a hydrophobic/kinetic meshwork creating a barrier to most macromolecules while allowing passage of transport receptor complexes (2,8). These complexes are thought to pass the NPC by interacting with FG repeats, thus permeating the NPC core (8 -11).…”