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The FG-repeat asymmetry of the nuclear pore complex is dispensable for bulk nucleocytoplasmic transport in vivo.

Zeitler B, Weis K - J. Cell Biol. (2004)

Bottom Line: The mutant Nups localize properly within the NPC and exhibit exchanged binding specificity for the export factor Xpo1.Surprisingly, we were unable to detect any defects in the Kap95, Kap121, Xpo1, or mRNA transport pathways in cells expressing the mutant FG Nups.These findings suggest that the biased distribution of FG repeats is not required for major nucleocytoplasmic trafficking events across the NPC.

View Article: PubMed Central - PubMed

Affiliation: Division of Cell and Developmental Biology, Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.

ABSTRACT
Nucleocytoplasmic transport occurs through gigantic proteinaceous channels called nuclear pore complexes (NPCs). Translocation through the NPC is exquisitely selective and is mediated by interactions between soluble transport carriers and insoluble NPC proteins that contain phenylalanine-glycine (FG) repeats. Although most FG nucleoporins (Nups) are organized symmetrically about the planar axis of the nuclear envelope, very few localize exclusively to one side of the NPC. We constructed Saccharomyces cerevisiae mutants with asymmetric FG repeats either deleted or swapped to generate NPCs with inverted FG asymmetry. The mutant Nups localize properly within the NPC and exhibit exchanged binding specificity for the export factor Xpo1. Surprisingly, we were unable to detect any defects in the Kap95, Kap121, Xpo1, or mRNA transport pathways in cells expressing the mutant FG Nups. These findings suggest that the biased distribution of FG repeats is not required for major nucleocytoplasmic trafficking events across the NPC.

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poly(A) RNA export is not inhibited in the FG mutants. Log-phase cells were fixed and stained for poly(A) RNA using an oligo-d(T) probe and detected by indirect immunofluorescence. DNA was labeled with Hoechst dye to visualize nuclei. mex67-5 cells were shifted to 37°C for 1 h before fixation. Bar, 5 μm.
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fig5: poly(A) RNA export is not inhibited in the FG mutants. Log-phase cells were fixed and stained for poly(A) RNA using an oligo-d(T) probe and detected by indirect immunofluorescence. DNA was labeled with Hoechst dye to visualize nuclei. mex67-5 cells were shifted to 37°C for 1 h before fixation. Bar, 5 μm.

Mentions: It is presently unclear how mRNAs are directionally exported through the NPC, because this transport pathway appears to function independently of the RanGTP gradient (for review see Reed and Hurt, 2002). We tested whether the correct positioning of the FG repeats of Nup1 and Nup159 is required for mRNA export by using an in situ hybridization probe to poly(A) mRNA (Fig. 5). As expected, mRNA export was inhibited in the export mutant mex67-5 at the nonpermissive temperature. In contrast, no nuclear accumulation of poly(A) RNA was detected in wild-type cells or in any of the nup1 or nup159 FG mutants. Therefore, we conclude that the FG-asymmetry of Nup1 and Nup159 is not required for efficient mRNA export.


The FG-repeat asymmetry of the nuclear pore complex is dispensable for bulk nucleocytoplasmic transport in vivo.

Zeitler B, Weis K - J. Cell Biol. (2004)

poly(A) RNA export is not inhibited in the FG mutants. Log-phase cells were fixed and stained for poly(A) RNA using an oligo-d(T) probe and detected by indirect immunofluorescence. DNA was labeled with Hoechst dye to visualize nuclei. mex67-5 cells were shifted to 37°C for 1 h before fixation. Bar, 5 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172579&req=5

fig5: poly(A) RNA export is not inhibited in the FG mutants. Log-phase cells were fixed and stained for poly(A) RNA using an oligo-d(T) probe and detected by indirect immunofluorescence. DNA was labeled with Hoechst dye to visualize nuclei. mex67-5 cells were shifted to 37°C for 1 h before fixation. Bar, 5 μm.
Mentions: It is presently unclear how mRNAs are directionally exported through the NPC, because this transport pathway appears to function independently of the RanGTP gradient (for review see Reed and Hurt, 2002). We tested whether the correct positioning of the FG repeats of Nup1 and Nup159 is required for mRNA export by using an in situ hybridization probe to poly(A) mRNA (Fig. 5). As expected, mRNA export was inhibited in the export mutant mex67-5 at the nonpermissive temperature. In contrast, no nuclear accumulation of poly(A) RNA was detected in wild-type cells or in any of the nup1 or nup159 FG mutants. Therefore, we conclude that the FG-asymmetry of Nup1 and Nup159 is not required for efficient mRNA export.

Bottom Line: The mutant Nups localize properly within the NPC and exhibit exchanged binding specificity for the export factor Xpo1.Surprisingly, we were unable to detect any defects in the Kap95, Kap121, Xpo1, or mRNA transport pathways in cells expressing the mutant FG Nups.These findings suggest that the biased distribution of FG repeats is not required for major nucleocytoplasmic trafficking events across the NPC.

View Article: PubMed Central - PubMed

Affiliation: Division of Cell and Developmental Biology, Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.

ABSTRACT
Nucleocytoplasmic transport occurs through gigantic proteinaceous channels called nuclear pore complexes (NPCs). Translocation through the NPC is exquisitely selective and is mediated by interactions between soluble transport carriers and insoluble NPC proteins that contain phenylalanine-glycine (FG) repeats. Although most FG nucleoporins (Nups) are organized symmetrically about the planar axis of the nuclear envelope, very few localize exclusively to one side of the NPC. We constructed Saccharomyces cerevisiae mutants with asymmetric FG repeats either deleted or swapped to generate NPCs with inverted FG asymmetry. The mutant Nups localize properly within the NPC and exhibit exchanged binding specificity for the export factor Xpo1. Surprisingly, we were unable to detect any defects in the Kap95, Kap121, Xpo1, or mRNA transport pathways in cells expressing the mutant FG Nups. These findings suggest that the biased distribution of FG repeats is not required for major nucleocytoplasmic trafficking events across the NPC.

Show MeSH