title: Ordered Regions of Channel Nucleoporins Nup62, Nup54, and Nup58 Form Dynamic Complexes in Solution.
abstract: Three out of ∼30 nucleoporins, Nup62, Nup54, and Nup58, line the nuclear pore channel. These "channel" nucleoporins each contain an ordered region of ∼150-200 residues, which is predicted to be segmented into 3-4 α-helical regions of ∼40-80 residues. Notably, these segmentations are evolutionarily conserved between uni- and multicellular eukaryotes. Strikingly, the boundaries of these segments match our previously reported mapping and crystal data, which collectively identified two "cognate" segments of Nup54, each interacting with cognate segments, one in Nup58 and the other one in Nup62. Because Nup54 and Nup58 cognate segments form crystallographic hetero- or homo-oligomers, we proposed that these oligomers associate into inter-convertible "mid-plane" rings: a single large ring (40-50 nm diameter, consisting of eight hetero-dodecamers) or three small rings (10-20 nm diameter, each comprising eight homo-tetramers). Each "ring cycle" would recapitulate "dilation" and "constriction" of the nuclear pore complex's central transport channel. As for the Nup54·Nup62 interactome, it forms a 1:2 triple helix ("finger"), multiples of which project alternately up and down from mid-plane ring(s). Collectively, our previous crystal data suggested a copy number of 128, 64, and 32 for Nup62, Nup54, and Nup58, respectively, that is, a 4:2:1 stoichiometry. Here, we carried out solution analysis utilizing the entire ordered regions of Nup62, Nup54, and Nup58, and demonstrate that they form a dynamic "triple complex" that is heterogeneously formed from our previously characterized Nup54·Nup58 and Nup54·Nup62 interactomes. These data are consistent both with our crystal structure-deduced copy numbers and stoichiometries and also with our ring cycle model for structure and dynamics of the nuclear pore channel.
authors.affiliation: From the Laboratory of Cell Biology, Howard Hughes Medical Institute, The Rockefeller University, New York, New York 10065.
authors.forename: Alok
authors.initials: A
authors.lastname: Sharma
authors.affiliation: From the Laboratory of Cell Biology, Howard Hughes Medical Institute, The Rockefeller University, New York, New York 10065.
authors.forename: Sozanne R
authors.initials: SR
authors.lastname: Solmaz
authors.affiliation: From the Laboratory of Cell Biology, Howard Hughes Medical Institute, The Rockefeller University, New York, New York 10065 blobel@rockefeller.edu.
authors.forename: Günter
authors.initials: G
authors.lastname: Blobel
authors.affiliation: From the Laboratory of Cell Biology, Howard Hughes Medical Institute, The Rockefeller University, New York, New York 10065 melcaki@rockefeller.edu.
authors.forename: Ivo
authors.initials: I
authors.lastname: Melčák
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000235:genetics; D000818:Animals; D002942:Circular Dichroism; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism; D022022:Nuclear Pore / Q000737:chemistry* / Q000235:genetics; D028861:Nuclear Pore Complex Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D010957:Plasmids; D051381:Rats; D012996:Solutions / Q000737:chemistry
chemical_list: D008562:Membrane Glycoproteins; D046912:Multiprotein Complexes; D028861:Nuclear Pore Complex Proteins; C582177:Nup54 protein, rat; C518579:Nup58 protein, rat; D012996:Solutions; C049312:nuclear pore protein p62
keywords: FG nucleoporins; Light scattering; Nuclear Pore Complex; Ring cycle hypothesis; Transport channel; biophysics; cell biology; nuclear pore; nuclear transport; protein assembly