Surface Properties Determining Passage Rates of Proteins through Nuclear Pores

Abstract

Nuclear pore complexes (NPCs) conduct nucleocytoplasmic transport through an FG domain-controlled barrier. We now explore how surface-features of a mobile species determine its NPC passage rate. Negative charges and lysines impede passage. Hydrophobic residues, certain polar residues (Cys, His), and, surprisingly, charged arginines have striking translocation-promoting effects. Favorable cation-π interactions between arginines and FG-phenylalanines may explain this apparent paradox. Application of these principles to redesign the surface of GFP resulted in variants that show a wide span of transit rates, ranging from 35-fold slower than wild-type to ∼500 times faster, with the latter outpacing even naturally occurring nuclear transport receptors (NTRs). The structure of a fast and particularly FG-specific GFP variant illustrates how NTRs can expose multiple regions for binding hydrophobic FG motifs while evading non-specific aggregation. Finally, we document that even for NTR-mediated transport, the surface-properties of the "passively carried" cargo can strikingly affect the translocation rate.

The liquid state of FG-nucleoporins mimics permeability barrier properties of nuclear pore complexes

“…3 B and corresponding Video 5). The rim stainings were indicative of formation of a barrier, reminiscent to previously observed rim staining under steady-state conditions of other fluorescent protein cargoes that had limited ability to penetrate into hydrogel particles (Frey et al, 2018). (3) An mCherry (≈30 kD) without the His tag did not show the rim effect (Fig.…”

Novel functional insights revealed by distinct protein-protein interactions of the residual SWI/SNF complex in SMARCA4-deficient small cell carcinoma of the ovary, hypercalcemic type

“…As several nuclear pore proteins have been shown to dynamically associate with nuclear pores and also be nucleoplasmic, the absence of nucleoporin (NUP) protein interactions we observed could be due to the nucleoplasmic pool of NUPs [64][65][66] . Several NUP proteins contain FG domains which may serve as low complexity domains and conceivably contribute to phase-separation of transcriptional compartments at nuclear pores [67][68][69][70] . NUP153, an FG domain containing NUP, only interacted with the wild-type SWI/SNF and may suggest dysregulation of phase separation in SCCOHT cells.…”

Nonclassical nuclear localization signals mediate nuclear import of CIRBP

“…Recently, we and others have reported a second function of importins, namely chaperoning and inhibiting liquid-liquid phase separation (LLPS) of aggregation-prone RNA-binding proteins (RBPs): for example, fused in sarcoma (FUS), TAF15, EWSR1, hnRNPA1, and hnRNPA2 by the importin Transportin-1 (TNPO1; also known as Karyopherin β2) and TDP-43 by importin α/β (1-4). Both nuclear import and chaperoning rely on the specific interaction of the importin with a nuclear localization signal (NLS) in its cargo protein (5)(6)(7). This interaction and hence, directionality of transport are usually regulated by the small guanosine triphosphate (GTP) hydrolase (GTPase) Ran as binding of Ran to the importin promotes release of the cargo in the nucleus (8).…”