Now that it is likely that all yeast nucleoporins are known, one of the ultimate goals is the in vitro assembly of the entire nuclear pore complex from its approximately 30 individual components. Here, we report the reconstitution of seven proteins (Nup133p, Nup145p-C, Nup120p, Nup85p, Nup84p, Seh1p and Sec13p) into a heptameric 0.5 MDa nuclear pore subcomplex. We found that double plasmid transformation combined with bi-cistronic mRNA translation allow the expression and assembly of distinct subcomplexes of up to five nucleoporins in a single Escherichia coli cell. During the sequential reconstitution of the Nup84p complex, smaller assembly intermediates can be isolated, which exhibit modular structures determined by electron microscopy that finally make up the whole Y-shaped Nup84p complex. Importantly, a seventh subunit, Nup133p, was incorporated into the complex through its interaction with Nup84p, thereby elongating one arm of the Y-shaped assembly to an approximately 40 nm long stalk. Taken together, our data document that the Nup84p-Nup133p complex self-assembles in a modular concept from distinct smaller nucleoporin construction sets.
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Evidence ID | Analyze ID | Gene/Complex | Systematic Name/Complex Accession | Qualifier | Gene Ontology Term ID | Gene Ontology Term | Aspect | Annotation Extension | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Gene | Gene Systematic Name | Phenotype | Experiment Type | Experiment Type Category | Mutant Information | Strain Background | Chemical | Details | Reference |
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Evidence ID | Analyze ID | Gene | Gene Systematic Name | Disease Ontology Term | Disease Ontology Term ID | Qualifier | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Regulator | Regulator Systematic Name | Target | Target Systematic Name | Direction | Regulation of | Happens During | Regulator Type | Direction | Regulation Of | Happens During | Method | Evidence | Strain Background | Reference |
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Site | Modification | Modifier | Source | Reference |
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Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Allele | Assay | Annotation | Action | Phenotype | SGA score | P-value | Source | Reference | Note |
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Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Assay | Annotation | Action | Modification | Source | Reference | Note |
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Complement ID | Locus ID | Gene | Species | Gene ID | Strain background | Direction | Details | Source | Reference |
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Evidence ID | Analyze ID | Dataset | Description | Keywords | Number of Conditions | Reference |
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