Yeast nucleoporins involved in passive nuclear envelope permeability.
The vertebrate nuclear pore complex (NPC) harbors an approximately 10-nm diameter diffusion channel that is large enough to admit 50-kD polypeptides. We have analyzed the permeability properties of the Saccharomyces cerevisiae nuclear envelope (NE) using import (NLS) and export (NES) signal-containing green fluorescent protein (GFP) reporters. Compared with wild-type, passive export rates of a classical karyopherin/importin (Kap) Kap60p/Kap95p-targeted NLS-GFP reporter (cNLS-GFP) were significantly faster in nup188-Delta and nup170-Delta cells. Similar results were obtained using two other NLS-GFP reporters, containing either the Kap104p-targeted Nab2p NLS (rgNLS) or the Kap121p-targeted Pho4p NLS (pNLS). Elevated levels of Hsp70 stimulated cNLS-GFP import, but had no effect on the import of rgNLS-GFP. Thus, the role of Hsp70 in NLS-directed import may be NLS- or targeting pathway-specific. Equilibrium sieving limits for the diffusion channel were assessed in vivo using NES-GFP reporters of 36-126 kD and were found to be greater than wild-type in nup188-Delta and nup170-Delta cells. We propose that Nup170p and Nup188p are involved in establishing the functional resting diameter of the NPC's central transport channel.
Shulga
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Department of Biology, University of Rochester, Rochester, New York 14627, USA.
Mosammaparast
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Wozniak
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Goldfarb
D S
DS
D001692:Biological Transport / Q000502:physiology; D003080:Cold Temperature; D003593:Cytoplasm / Q000378:metabolism; D004058:Diffusion; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents / Q000378:metabolism; D007700:Kinetics; D008164:Luminescent Proteins / Q000235:genetics; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism*; D016296:Mutagenesis / Q000502:physiology; D009685:Nuclear Envelope / Q000378:metabolism*; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism*; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D015398:Signal Transduction / Q000502:physiology*
D007202:Indicators and Reagents; D008164:Luminescent Proteins; D008565:Membrane Proteins; C097178:NUP170 protein, S cerevisiae; C100410:NUP188 protein, S cerevisiae; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins