Phase separation of intrinsically disordered FG-Nups is driven by highly dynamic FG motifs

Abstract

The intrinsically disordered FG-Nups in the central channel of the nuclear pore complex (NPC) form a selective permeability barrier, allowing small molecules to traverse by passive diffusion, while large molecules can only translocate with the help of nuclear transport receptors. The exact phase state of the permeability barrier remains elusive. In vitro experiments have shown that some FG-Nups can undergo phase separation into condensates that display NPC-like permeability barrier properties. Here, we use molecular dynamics simulations at amino acid resolution to study the phase separation characteristics of each of the disordered FG-Nups of the yeast NPC. We find that GLFG-Nups undergo phase separation and reveal that the FG motifs act as highly dynamic hydrophobic stickers that are essential for the formation of FG-Nup condensates featuring droplet-spanning percolated networks. Additionally, we study phase separation in an FG-Nup mixture that resembles the NPC stoichiometry and observe that an NPC condensate is formed containing multiple GLFG-Nups. We find that the phase separation of this NPC condensate is also driven by FG–FG interactions, similar to the homotypic FG-Nup condensates. Based on the observed phase separation behavior, the different FG-Nups of the yeast NPC can be divided into two classes: The FG-Nups (mostly GLFG-type) located in the central channel of the NPC form a highly dynamic percolated network formed by many short-lived FG–FG interactions, while the peripheral FG-Nups (mostly FxFG-type) at the entry and exit of the NPC channel likely form an entropic brush.

Selective phase separation of transcription factors is driven by orthogonal molecular grammar

“…In all cases highly dynamic contacts are seen with contact lifetimes in the range 4-6 ns (Fig. S9), showing that highly dynamical cross links are present [46, 47].…”

“…The intermolecular interactions in the TF condensates, however, are not at all permanent; they are highly dynamic with contact lifetimes on the order of 5 ns (see Fig. S9) such that the cross-links between TFs are transient in nature, similar to FG-nucleoporin condensates [47]. The nature of this phase state can best be categorized as viscoelastic: on one hand the continuous percolations suggest elastic properties, while their transient nature is indicative of a fluid, with mostly spherical morphologies and the ability of two droplets to merge into one [50].…”

“…In this work we use the 1BPA model that was previously developed for the study of intrinsically disordered proteins in the nuclear pore complex (NPC) [53,54]. It has been used extensively to model the behaviour of the disordered nucleoporin regions which fill the center of the yeast NPC and provide a selective barrier to the transport of cargo between the cytoplasm and nucleoplasm [47,55,56]. In this work an updated 1BPA model is developed (1BPA-2.1) that extends the applicability domain beyond the intrinsically disordered domains of yeast nucleoporins, by incorporating a greater training set for parameterisation (collating experimental data from [35,[57][58][59][60]), and further improves the performance of the previously used 1BPA-2.0 model [45,61].…”

RNA regulates cohesiveness and porosity of a biological condensate

“…MD studies of biomolecular inter-actions enabling the LLPS phenomena differ in their resolution from explicit solvent all-atom models [25][26][27][28] to coarse-grained models that in turn differ from one another by the parameterization of the intermolecular interactions and their treatment of solvent [29][30][31] . Among the most popular one-bead-per-amino-acid models used for MD simulations of biomolecular condensates are the Kim-Hummer (KH) model [32][33][34] , the HPS model 33 and its variants [35][36][37] , the AWSEM-IDP model 38 , and more specialized models targeting a particular type of disordered proteins 39 . We note, however, that the majority of the CG MD simulations to date have focused on characterization of the protein components of the condensates, in part because of the lack of CG models of RNA compatible with the existing CG models of intrinsically disordered proteins.…”

Coordinating nucleoporin condensation and nuclear pore complex assembly

“…Conversely, excess NTRs result in a strengthening of the barrier [33]. How the selective permeability barrier is formed and how this facilitates the fast nucleocytoplasmic transport mechanism continue to be an active area of investigation [15,21,22,34–39].…”