We describe post-transcriptional gene regulation in yeast based on direct RNA-ligand interaction. Tetracycline-dependent translational regulation could be imposed via specific aptamers inserted at two different positions in the 5' untranslated region (5'UTR). Translation in vivo was suppressed up to ninefold upon addition of tetracycline. Repression via an aptamer located near the start codon (cap-distal) in the 5'UTR was more effective than repression via a cap-proximal position. On the other hand, suppression in a cell-free system reached maximally 50-fold and was most effective via a cap-proximal aptamer. Examination of the kinetics of tetracycline-dependent translational inhibition in vitro revealed that preincubation of tetracycline and mRNA before starting translation led not only to the fastest onset of inhibition but also the most effective repression. The differences between the behaviour of the regulatory system in vivo and in vitro are likely to be related to distinct properties of mRNP structure and mRNA accessibility in intact cells as opposed to cell-extracts. Tetracycline-dependent regulation was also observed after insertion of an uORF sequence upstream of the aptamer, indicating that our system also targets reinitiating ribosomes. Polysomal gradient analyses provided insight into the mechanism of regulation. Cap-proximal insertion inhibits binding of the 43S complex to the cap structure whereas start-codon-proximal aptamers interfere with formation of the 80S ribosome, probably by blocking the scanning preinitiation complex.
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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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