Fig 1.
Each of the Nuclear Pore Complexes depicted has a cylindrical structural domain (navy blue) which spans the lipid bilayers (grey) which make up the nuclear envelope, oriented such that the tops of the pores face the cytoplasm, while the bottoms of the pores face the nucleus. The nuclear basket, located on the nuclear face of the pore is not shown for simplicity. Additionally the cytoplasmic filaments of the pore, which are not directly responsible for gating [10], are shown in yellow. A: In the original DCBG model individual di-block FG nups (brown) have collapsed coil gel-like regions (dark brown) and extended coil brush-like domains (light brown), resulting in a microphase separation of these domains within the NPC. This results in a central plug-like structure supported by a polymer brush of extended disordered regions of FG nups. [11] B: The modified DCBG model which is the focus of this manuscript is the same as in part A except for the addition of a dense region of single-block FG nups which lie along the wall of the NPC. The core of our proposed Di-block Copolymer Brush Gate model is the idea that when particular transport factors are present which are able to outcompete the inter-FG domain “sticky tip” interactions, the polymer brush which fills the NPC is able to open up to a new free energy minimum that can accommodate the transit of cargo, while when interactions between sticky tips are able to recover into the several kT range, the pore is able to close.
Fig 2.
Biphasic FG nups and illustration of the Forest model.
A: Diagrams of the various FG nups in S. cerevisiae and their hydrodynamic radii. Blue or red lines depict either high (red) or low (blue) content of charged AAs along the disordered region of the FG nup. The small green triangles represent the anchor domain of each FG nup. Single-block FG nups, termed “shrubs” in the Forest model, were categorized as consisting of a continuos collapsed FG domain adjacent to an anchor domain, consisting of Nup57, Nup49, and Nup42. Di-block FG nups, or “trees” in the terminology of the Forest model, were categorized as having a collapsed FG domain at their free ends separated from the anchor domain by a extended coil domain B: A diagram of the Forest model NPC architecture. Orientation of the pore is such that top side faces the cytoplasm, while the bottom side faces the nucleus. FG nups are drawn to scale and positioned according to the relative location of their anchor domains along the z-axis of the NPC, as determined by immuno-localization [30].
Fig 3.
Coarse grained molecular modeling of single-block FG nups.
A: Left is a simulation snapshot of full length Nup57 showing the single polymer block FG domain. Right shows a snapshot of full length Nup1 demonstrating the di-block structure of FG and extended domain structures. B: Left, the contact probability map for Nup57 show the time-averaged contacts between all pairs of amino acids. A monolithic block diagonal structure fills the entire contact map, implying that this FG nup is comprised of a single polymer block FG domain. Right, in contrast, the contact map for human Nup1 shows one block for the FG domain and diagonal contacts for the extended polymer domain, with both domains having low probability of cross contact.
Fig 4.
Contact maps for the different single-block FG nups in Baker’s yeast from coarse grained molecular modeling.
A: Nup57’s contact map B: Nup49’s contact map C: Nup42’s contact map. Contact probability maps show the time averaged contacts between all pairs of amino acids. A block diagonal structure in the contact maps represents a collapsed coil structure, which describes the single-block FG nups well, except for a small region of around 80 amino acids in length where Nup42 anchors to the pore wall. Amino acid residues shown are with respect to the disordered domains of the simulated FG nups, while full protein amino indexes can be determined by domain definitions in Yamada et al [30].
Fig 5.
Mean field polymer brush model of the Nuclear Pore Complex.
A: Schematic of a polymer brush structure formed by di-block FG nups. Parameters H, height of the brush; R, radius of the pore; δ, diameter of the “sticky tips”; and d, the average distance between anchor points. Green circles represent the locations at which FG nups are grafted to the pore. B: Free energy of the Nsp1 brush, with and without single-block FG nups present on the pore wall, as a function of brush height for various values of the blob cohesive energy (ϵkBT). Brush height can extend to a maximum of around 22 nm, which is the radius R of the modeled pore minus the size of the sticky tips. Di-block FG nup tip to single-block FG nup cohesion is fixed at ϵs = 6kT. Right: Schematic diagram of the proposed Di-block Copolymer Brush Gate model at various minima of the brush free energy. When particular transport factors are present which are able to outcompete the inter-FG domain “sticky tip” interactions, the brush is able to open up to a new free energy minimum that can accommodate the cargo. When interactions between sticky tips are able to recover into the several kT range, the pore is able to close with a free energy minimum at H ∼ R − δ. We have previously estimated the self interaction energy level of the Nsp1 sticky tip to be 4.7 kT [11], which also sets the energy scale for blob-blob interactions of ϵkT. Single-block FG nups (shrubs) denoted by green compact polymers in the schematic, produce a new minimum in the free energy close to the wall, thus potentially allowing for wide-open states of the brush if tip-tip cohesiveness is lost or reduced.
Fig 6.
Overview of brush response to tip-tip and tip-transport factor interaction levels in the Di-block Copolymer Brush Gate (DCBG) model with single-block FG nups.
A: Changes in tip-tip cohesiveness result in equilibrium brush openings that have the nuclear pore in either significantly open or completely closed states, with a sharp transition at ∼1.6 kT when single-block FG nups (shrubs) are not present and ∼1.9 kT when single-block FG nups are present. The single-block FG nups almost double the equilibrium brush opening for weak tip-tip cohesiveness below ∼1.6 kT. Tip to single-block FG nup cohesion is fixed at ϵs = 6kT. B: Maximum brush opening assuming the energy of tip-cargo interaction is added to the brush free energy, resulting in an increase in the possible opening of the pore. Maximum brush opening is plotted for different amounts of free energy introduced into the brush system by tip-transport factor binding for two different values of the tip to tip cohesion ϵ. Tip to single-block FG nup cohesion is fixed at ϵs = 6kT.