title: Domain topology of nucleoporin Nup98 within the nuclear pore complex.
abstract: Nuclear pore complexes (NPCs) facilitate selective transport of macromolecules across the nuclear envelope in interphase eukaryotic cells. NPCs are composed of roughly 30 different proteins (nucleoporins) of which about one third are characterized by the presence of phenylalanine-glycine (FG) repeat domains that allow the association of soluble nuclear transport receptors with the NPC. Two types of FG (FG/FxFG and FG/GLFG) domains are found in nucleoporins and Nup98 is the sole vertebrate nucleoporin harboring the GLFG-type repeats. By immuno-electron microscopy using isolated nuclei from Xenopus oocytes we show here the localization of distinct domains of Nup98. We examined the localization of the C- and N-terminal domain of Nup98 by immunogold-labeling using domain-specific antibodies against Nup98 and by expressing epitope tagged versions of Nup98. Our studies revealed that anchorage of Nup98 to NPCs through its C-terminal autoproteolytic domain occurs in the center of the NPC, whereas its N-terminal GLFG domain is more flexible and is detected at multiple locations within the NPC. Additionally, we have confirmed the central localization of Nup98 within the NPC using super resolution structured illumination fluorescence microscopy (SIM) to position Nup98 domains relative to markers of cytoplasmic filaments and the nuclear basket. Our data support the notion that Nup98 is a major determinant of the permeability barrier of NPCs.
authors.affiliation: Institute of Molecular Biology and Medicine, Université Libre de Bruxelles, Charleroi, Belgium.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Chatel
authors.forename: Sachin H
authors.initials: SH
authors.lastname: Desai
authors.forename: Alexa L
authors.initials: AL
authors.lastname: Mattheyses
authors.forename: Maureen A
authors.initials: MA
authors.lastname: Powers
authors.forename: Birthe
authors.initials: B
authors.lastname: Fahrenkrog
mesh_terms: D000818:Animals; D000906:Antibodies / Q000378:metabolism; D001692:Biological Transport; D015153:Blotting, Western; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure; D016253:Microscopy, Immunoelectron / Q000379:methods; D022022:Nuclear Pore / Q000737:chemistry* / Q000648:ultrastructure; D028861:Nuclear Pore Complex Proteins / Q000737:chemistry* / Q000648:ultrastructure; D009865:Oocytes / Q000378:metabolism; D017434:Protein Structure, Tertiary; D014981:Xenopus / Q000254:growth & development
chemical_list: D000906:Antibodies; D028861:Nuclear Pore Complex Proteins; C093350:nuclear pore complex protein 98
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