Comprehensive identification of translation start sites by tetracycline-inhibited ribosome profiling
| Autor: | Kenji Nakahigashi, Toru Nakayashiki, Yuki Takai, Michiko Kimura, Hirofumi Yoshikawa, Yuh Shiwa, Yasushi Ishihama, Barry L. Wanner, Hirotada Mori, Nozomi Abe |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
genome annotation Pseudogene 030106 microbiology Codon Initiator Biology translation initiation 03 medical and health sciences Open Reading Frames Eukaryotic translation Genetics Escherichia coli Ribosome profiling Molecular Biology N-terminal ribosome profiling Protein Synthesis Inhibitors TetRP Translation (biology) Molecular Sequence Annotation General Medicine Genome project Full Papers Tetracycline Open reading frame 030104 developmental biology GenBank Ribosomes |
| Zdroj: | DNA Research: An International Journal for Rapid Publication of Reports on Genes and Genomes |
| ISSN: | 1756-1663 1340-2838 |
| Popis: | Tetracycline-inhibited ribosome profiling (TetRP) provides a powerful new experimental tool for comprehensive genome-wide identification of translation initiation sites in bacteria. We validated TetRP by confirming the translation start sites of protein-coding genes in accordance with the 2006 version of Escherichia coli K-12 annotation record (GenBank U00096.2) and found ∼150 new start sites within 60 nucleotides of the annotated site. This analysis revealed 72 per cent of the genes whose initiation site annotations were changed from the 2006 GenBank record to the newer 2014 annotation record (GenBank U00096.3), indicating a high sensitivity. Also, results from reporter fusion and proteomics of N-terminally enriched peptides showed high specificity of the TetRP results. In addition, we discovered over 300 translation start sites within non-coding, intergenic regions of the genome, using a threshold that retains ∼2,000 known coding genes. While some appear to correspond to pseudogenes, others may encode small peptides or have previously unforeseen roles. In summary, we showed that ribosome profiling upon translation inhibition by tetracycline offers a simple, reliable and comprehensive experimental tool for precise annotation of translation start sites of expressed genes in bacteria. |
| Databáze: | OpenAIRE |
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