title: Intermolecular disulfide bonds between nucleoporins regulate karyopherin-dependent nuclear transport.
abstract: Disulfide (S-S) bonds play important roles in the regulation of protein function and cellular stress responses. In this study, we demonstrate that distinct sets of nucleoporins (Nups), components of the nuclear pore complex (NPC), form S-S bonds and regulate nuclear transport through the NPC. Kinetic analysis of importin β demonstrated that the permeability of the NPC was increased by dithiothreitol treatment and reduced by oxidative stress. The permeability of small proteins such as GFP was not affected by either oxidative stress or a reducing reagent. Immunoblot analysis revealed that the oxidative stress significantly induced S-S bond formation in Nups 358, 155, 153 and 62 but not 88 and 160. The direct involvement of cysteine residues in the formation of S-S bonds was confirmed by mutating conserved cysteine residues in Nup62, which abolished the formation of S-S bonds and enhanced the permeability of the NPC. Knocking down Nup62 reduced the stress-inducible S-S bonds of Nup155, suggesting that Nup62 and Nup155 are covalently coupled via S-S bonds. From these results, we propose that the inner channel of the NPC is somehow insulated from the cytoplasm and is more sensitive than the cytoplasm to the intracellular redox state.
authors: Graduate School of Biostudies, Kyoto University, Yoshida-konoe-cho, Sakyo-ku, Kyoto 606-8501, Japan. yoshimura@lif.kyoto-u.ac.jp
authors: Shige H
authors: SH
authors: Yoshimura
authors: Shotaro
authors: S
authors: Otsuka
authors: Masahiro
authors: M
authors: Kumeta
authors: Mariko
authors: M
authors: Taga
authors: Kunio
authors: K
authors: Takeyasu
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000235:genetics; D002463:Cell Membrane Permeability / Q000187:drug effects / Q000235:genetics; D003545:Cysteine / Q000737:chemistry / Q000235:genetics; D004229:Dithiothreitol / Q000494:pharmacology; D055785:Gene Knockdown Techniques; D006367:HeLa Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D022022:Nuclear Pore / Q000378:metabolism*; D028861:Nuclear Pore Complex Proteins / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction; D017382:Reactive Oxygen Species / Q000378:metabolism; D028961:beta Karyopherins / Q000378:metabolism*
chemical_list: D028861:Nuclear Pore Complex Proteins; D017382:Reactive Oxygen Species; D028961:beta Karyopherins; D006861:Hydrogen Peroxide; D003545:Cysteine; D004229:Dithiothreitol
keywords: Disulfide bridge; Importin β; Nuclear pore; Nucleocytoplasmic transport; Oxidative stress