Amyloid-like interactions within nucleoporin FG hydrogels

Abstract

The 62 kDa FG repeat domain of the nucleoporin Nsp1p forms a hydrogel-based, sieve-like permeability barrier that excludes inert macromolecules but allows rapid entry of nuclear transport receptors (NTRs). We found that the N-terminal part of this domain, which is characterized by Asn-rich inter-FG spacers, forms a tough hydrogel. The C-terminal part comprises charged inter-FG spacers, shows low gelation propensity on its own, but binds the Nterminal part and passivates the FG hydrogel against nonselective interactions. It was previously shown that a hydrophobic collapse involving Phe residues is required for FG hydrogel formation. Using solid-state NMR spectroscopy, we now identified two additional types of intragel interactions, namely, transient hydrophobic interactions between Phe and methyl side chains as well as intermolecular β-sheets between the Asn-rich spacer regions. The latter appear to be the kinetically most stable structures within the FG hydrogel. They are also a central feature of neuronal inclusions formed by Asn/Gln-rich amyloid and prion proteins. The cohesive properties of FG repeats and the Asn/Gln-rich domain from the yeast prion Sup35p appear indeed so similar to each other that these two modules interact in trans. Our data, therefore, suggest a fully unexpected cellular function of such interchain β-structures in maintaining the permeability barrier of nuclear pores. They provide an explanation for how contacts between FG repeats might gain the kinetic stability to suppress passive fluxes through nuclear pores and yet allow rapid NTR passage.Beta-sheet | nuclear pore complex | Nup | Prion | solid-state NMR N uclear pore complexes (NPCs) control all nucleocytoplasmic exchange (1-4). Their permeability barrier allows free passage of small molecules but suppresses the flux of macromolecules larger than 30 kDa and thereby prevents an uncontrolled intermixing of nuclear and cytoplasmic contents. However, the permeability barrier also permits a rapid passage of even large cargoes, provided these are bound to appropriate nuclear transport receptors (NTRs) (2-5). NTRs thereby supply nuclei with proteins and the cytoplasm with ribosomes and other nuclear products.FG repeat domains are essential building blocks of NPCs (6). They are considered to be natively unfolded and contain up to 50 repeat units, in which a characteristic hydrophobic patch, typically with the sequence FG, FxFG, or GLFG, is surrounded by more hydrophilic spacer sequences (7-9). These hydrophobic patches transiently bind NTRs during facilitated NPC passage (10-15).Recent evidence suggests that the permeability barrier is a hydrogel derived from FG repeat domains (FG hydrogel) (13,(15)(16)(17)(18). FG hydrogels have been predicted by the selective phase model (15,16,19) and indeed have been reconstituted from the FG/FxFG domain of the yeast nucleoporin Nsp1p (13,16) or the GLFG domains from Nup49p and Nup57p (17). All these gels showed permeability properties very similar to those of NPCs themselves: they allowed an up to 20,000...

Toxic PR n poly-dipeptides encoded by the C9orf72 repeat expansion block nuclear import and export

“…proteins have described polymerization of FG domains into cross-β fibers (22)(23)(24). We observed similar evidence of the polymerization of recombinant proteins composed of the FG domains of the human Nup54 and Nup98 nucleoporin proteins.…”

The self-interaction of native TDP-43 C terminus inhibits its degradation and contributes to early proteinopathies

“…Comparisons among the cellular and misfolded properties of the known Q/N-rich domain are shown in Table 1 [20][21][22][23][24][25][26][27][28][29][30][31] . The cellularfolded Q/N-rich domain shared key properties characteristic of the misfolded proteins, including insolubility and a tendency to form aggregates via intrinsic element-and self-interactions.…”

Converging on the function of intrinsically disordered nucleoporins in the nuclear pore complex

“…As suggested by the ROD model (Peters, 2005(Peters, , 2009a, this implies that the FG-domains are less extended (i.e., or partially collapsed) under physiological transport conditions. Next, two separate papers (Ader et al, 2010;Yamada et al, 2010) now show corroborating evidence that could help to dispel the conflicting experimental evidence that yeast Nsp1 is both cohesive (Frey et al, 2006;Gorlich, 2007, 2009) and non-cohesive (Patel et al, 2007). Here, both groups report that the FG-domain of Nsp1 contains both a cohesive and non-cohesive region in its N-and C-terminal parts, respectively.…”