Nup2p, a Yeast Nucleoporin, Functions in Bidirectional Transport of Importin α

Solsbacher, Jens; Maurer, Patrick; Vogel, Frank; Schlenstedt, Gabriel

doi:10.1128/mcb.20.22.8468-8479.2000

Cited by 88 publications

(131 citation statements)

References 53 publications

(85 reference statements)

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“…Cse1p Also Accelerates Dissociation of Kap60p from Nup2p via a KaRF Mechanism-It has been previously shown that Cse1p⅐Gsp1p antagonizes the interaction of Kap60p with Nup2p, and it was proposed that Nup2p serves as a platform for Cse1p-mediated export of Kap60p from the nucleus (10,11,28). We therefore examined whether Cse1p⅐Gsp1p can also accelerate the rate of dissociation of Kap60p from Nup2p.…”

Section: Resultsmentioning

confidence: 99%

“…Also, the autoinhibitory mechanism is not necessarily a nucleoplasmic event, since it can happen anywhere in the cell where Kap60p⅐NLS complexes form. Perhaps as a consequence, cells recruited two additional proteins, Cse1p and Nup2p, to aid in the disruption of Kap60p⅐NLS complexes in the nucleoplasm (10,15).…”

Section: Discussionmentioning

confidence: 99%

“…As a first clue, we noted that Nup1p and Nup2p are unique among yeast FG Nups in that they bind Kap60p monomers directly (10,11,28,29) (see Fig. 5A for an example).…”

Section: A Conserved Sequence Motif In a Compact Domain Of Nup2pmentioning

confidence: 99%

“…After 5 min at room temperature, beads were collected, and the amount of radiolabeled Kap60p remaining bound was determined by liquid scintillation. The percentage dissociation of Kap60p⅐NLS-cargo complexes (or KaRF activity) was determined as described in the legend to N-terminal 172 amino acids of Nup2p (10,28). A comparison of the two regions revealed a sequence motif, VMXXRKIA, which is highly conserved between Nup1p and Nup2p (Fig.…”

Section: A Conserved Sequence Motif In a Compact Domain Of Nup2pmentioning

confidence: 99%

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Molecular Basis for the Rapid Dissociation of Nuclear Localization Signals from Karyopherin α in the Nucleoplasm

Gilchrist

Rexach

2003

Journal of Biological Chemistry

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The yeast karyopherin heterodimer Kap60p⅐Kap95p facilitates nuclear import of proteins bearing a classic nuclear localization signal (NLS). The ␣ subunit Kap60p binds to the NLS of cargo molecules in the cytoplasm, forming stable complexes that must ultimately dissociate in the nucleoplasm. Although Kap60p can release NLSs on its own using an autoinhibitory sequence (AIS) motif that can occupy the NLS binding site, that mechanism is too slow to support rapid nuclear import. We previously showed that the nuclear basket nucleoporin Nup2p and the exportin complex Cse1p⅐Gsp1p⅐GTP function as karyopherin release factors (KaRFs) because they can accelerate the rate of dissociation of NLSs from Kap60p. Here we dissect the molecular mechanics of their KaRF activity. We show that Cse1p accelerates dissociation of Kap60p⅐NLS-cargo complexes and Kap60p⅐Nup2p complexes by increasing the affinity of Kap60p for its AIS motif. In contrast, Nup2p uses a conserved sequence motif (VMXXRKIA) coupled to an AISlike motif to accelerate dissociation of Kap60p⅐NLS complexes in a vectorial reaction mechanism. Mutation of either motif in Nup2p leads to a loss of KaRF activity and to the accumulation of Kap60p⅐NLS-cargo complexes in the nucleoplasm of yeast. We discuss a model whereby Nup2p, Cse1p, and Gsp1p cooperate to establish directionality in the movement of Kap60p and NLScargos across the nuclear pore complex.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

“…As a first clue, we noted that Nup1p and Nup2p are unique among yeast FG Nups in that they bind Kap60p monomers directly (10,11,28,29) (see Fig. 5A for an example).…”

Section: A Conserved Sequence Motif In a Compact Domain Of Nup2pmentioning

confidence: 99%

Section: A Conserved Sequence Motif In a Compact Domain Of Nup2pmentioning

confidence: 99%

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Molecular Basis for the Rapid Dissociation of Nuclear Localization Signals from Karyopherin α in the Nucleoplasm

Gilchrist

Rexach

2003

Journal of Biological Chemistry

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“…This model for protein import requires precise tuning of the thermodynamic interactions between the various species for the reaction to proceed efficiently in a single direction. In addition, it will be interesting to determine what contribution other release factors, such as Cse1p and Nup2p (18,50), make to cargo release in vivo.…”

Section: Table II Binding Of Importin ␣ Proteins To Nls-cargomentioning

confidence: 99%

The Auto-inhibitory Function of Importin α Is Essentialin Vivo

Harreman

Hodel²,

Fanara³

et al. 2003

Journal of Biological Chemistry

104

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Proteins that contain a classical nuclear localization signal (NLS) are recognized in the cytoplasm by a heterodimeric import receptor composed of importin/ karyopherin ␣ and ␤. The importin ␣ subunit recognizes classical NLS sequences, and the importin ␤ subunit directs the complex to the nuclear pore. Recent work shows that the N-terminal importin ␤ binding (IBB) domain of importin ␣ regulates NLS-cargo binding in the absence of importin ␤ in vitro. To analyze the in vivo functions of the IBB domain, we created a series of mutants in the Saccharomyces cerevisiae importin ␣ protein. These mutants dissect the two functions of the N-terminal IBB domain, importin ␤ binding and autoinhibition. One of these importin ␣ mutations, A3, decreases auto-inhibitory function without impacting binding to importin ␤ or the importin ␣ export receptor, Cse1p. We used this mutant to show that the auto-inhibitory function is essential in vivo and to provide evidence that this auto-inhibitory-defective importin ␣ remains bound to NLS-cargo within the nucleus. We propose a model where the auto-inhibitory activity of importin ␣ is required for NLS-cargo release and the subsequent Cse1p-dependent recycling of importin ␣ to the cytoplasm.In eukaryotes, the nuclear envelope provides an essential barrier that separates the nuclear genome from the intermediary metabolism, signaling systems, and translation machinery of the cytoplasm. Selective bi-directional transport of macromolecules across this nuclear envelope regulates critical cellular processes such as gene expression (1, 2). All nucleocytoplasmic transport of macromolecules occurs through large proteinaceous structures, called nuclear pore complexes (NPC), 1 that perforate the nuclear envelope (3, 4). These macromolecular cargoes are specifically targeted to and transported through NPCs by a family of soluble nuclear transport receptors (5, 6).The small GTPase, Ran, governs the interactions between the nuclear transport receptors and macromolecular cargoes (5, 7). Import receptors bind cargo in the absence of RanGTP, whereas export receptors bind cargo in a trimeric complex with RanGTP (5,8). This mode of regulation requires an asymmetric distribution of RanGTP, with more RanGTP in the nucleus than in the cytoplasm. To achieve this asymmetry, the Ran regulatory proteins are compartmentalized with the GTPase activating protein (RanGAP), which generates RanGDP, in the cytoplasm (9) and the guanine nucleotide exchange factor (RCC1), which generates RanGTP, in the nucleus (10).The best-characterized nuclear import process occurs via receptor recognition of a classical nuclear localization signal (NLS). This classical NLS is typified by a cluster of basic amino acids (monopartite) or two clusters of basic amino acids separated by a 10 -12 amino acid linker (bipartite) (11, 12). A heterodimeric import receptor, composed of importins ␣ and ␤ (also known as karyopherin ␣ and ␤), mediates the nuclear import of proteins that contain a classical NLS (13-15). Over the last several years many...

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Current Awareness

2001

Yeast

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In order to keep subscribers up‐to‐date with the latest developments in their field, this current awareness service is provided by John Wiley & Sons and contains newly‐published material on yeasts. Each bibliography is divided into 10 sections. 1 Books, Reviews & Symposia; 2 General; 3 Biochemistry; 4 Biotechnology; 5 Cell Biology; 6 Gene Expression; 7 Genetics; 8 Physiology; 9 Medical Mycology; 10 Recombinant DNA Technology. Within each section, articles are listed in alphabetical order with respect to author. If, in the preceding period, no publications are located relevant to any one of these headings, that section will be omitted. (5 weeks journals ‐ search completed 6th Dec. 2000)

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Nup2p, a Yeast Nucleoporin, Functions in Bidirectional Transport of Importin α

Cited by 88 publications

References 53 publications

Molecular Basis for the Rapid Dissociation of Nuclear Localization Signals from Karyopherin α in the Nucleoplasm

Molecular Basis for the Rapid Dissociation of Nuclear Localization Signals from Karyopherin α in the Nucleoplasm

The Auto-inhibitory Function of Importin α Is Essentialin Vivo

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