title: Entry into the nuclear pore complex is controlled by a cytoplasmic exclusion zone containing dynamic GLFG-repeat nucleoporin domains.
abstract: Nuclear pore complexes (NPCs) mediate nucleocytoplasmic movement. The central channel contains proteins with phenylalanine-glycine (FG) repeats, or variations (GLFG, glycine-leucine-phenylalanine-glycine). These are 'intrinsically disordered' and often represent weak interaction sites that become ordered upon interaction. We investigated this possibility during nuclear transport. Using electron microscopy of S. cerevisiae, we show that NPC cytoplasmic filaments form a dome-shaped structure enclosing GLFG domains. GLFG domains extend out of this structure and are part of an 'exclusion zone' that might act as a partial barrier to entry of transport-inert proteins. The anchor domain of a GLFG nucleoporin locates exclusively to the central channel. By contrast, the localisation of the GLFG domains varied between NPCs and could be cytoplasmic, central or nucleoplasmic and could stretch up to 80 nm. These results suggest a dynamic exchange between ordered and disordered states. In contrast to diffusion through the NPC, transport cargoes passed through the exclusion zone and accumulated near the central plane. We also show that movement of cargo through the NPC is accompanied by relocation of GLFG domains, suggesting that binding, restructuring and movement of these domains could be part of the translocation mechanism.
authors.affiliation: School of Biological and Biomedical Sciences, Durham University, Science Laboratories, South Road, Durham DH1 3LE, UK.
authors.forename: Jindriska
authors.initials: J
authors.lastname: Fiserova
authors.forename: Matthew
authors.initials: M
authors.lastname: Spink
authors.forename: Shane A
authors.initials: SA
authors.lastname: Richards
authors.forename: Christopher
authors.initials: C
authors.lastname: Saunter
authors.forename: Martin W
authors.initials: MW
authors.lastname: Goldberg
mesh_terms: D021581:Active Transport, Cell Nucleus; D002467:Cell Nucleus / Q000378:metabolism* / Q000648:ultrastructure; D003593:Cytoplasm / Q000378:metabolism* / Q000648:ultrastructure; D008854:Microscopy, Electron; D022022:Nuclear Pore / Q000737:chemistry* / Q000378:metabolism; D028861:Nuclear Pore Complex Proteins / Q000737:chemistry* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D020449:Repetitive Sequences, Amino Acid; D012441:Saccharomyces cerevisiae / Q000378:metabolism* / Q000648:ultrastructure; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: D028861:Nuclear Pore Complex Proteins; D029701:Saccharomyces cerevisiae Proteins
keywords: GLFG; Nuclear pore; Phenylalanine-glycine repeat; Transport