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Multi-site rate control analysis identifies ribosomal scanning as the sole high-capacity/low-flux-control step in mRNA translation
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Firczuk, Helena, Teahan, James, Mendes, Pedro and McCarthy, John E. G. (2020) Multi-site rate control analysis identifies ribosomal scanning as the sole high-capacity/low-flux-control step in mRNA translation. FEBS Journal, 287 (5). pp. 925-940. doi:10.1111/febs.15059
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WRAP-multi-site-rate-control-analysis-ribosomal-sole-mRNA-McCarthy-2019.pdf - Accepted Version - Requires a PDF viewer. Download (3340Kb) | Preview |
Official URL: https://doi.org/10.1111/febs.15059
Abstract
Control of complex intracellular pathways such as protein synthesis is critical to organism survival, but is poorly understood. Translation of a reading frame in eukaryotic mRNA is preceded by a scanning process in which a subset of translation factors helps guide ribosomes to the start codon. Here, we perform comparative analysis of the control status of this scanning step that sits between recruitment of the small ribosomal subunit to the m7GpppG‐capped 5′end of mRNA and of the control exerted by downstream phases of polypeptide initiation, elongation and termination. We have utilized a detailed predictive model as guidance for designing quantitative experimental interrogation of control in the yeast translation initiation pathway. We have built a synthetic orthogonal copper‐responsive regulatory promoter (PCuR3) that is used here together with the tet07 regulatory system in a novel dual‐site in vivo rate control analysis strategy. Combining this two‐site strategy with calibrated mass spectrometry to determine translation factor abundance values, we have tested model‐based predictions of rate control properties of the in vivo system. We conclude from the results that the components of the translation machinery that promote scanning collectively function as a low‐flux‐control system with a capacity to transfer ribosomes into the core process of polypeptide production that exceeds the respective capacities of the steps of polypeptide initiation, elongation and termination. In contrast, the step immediately prior to scanning, that is, ribosome recruitment via the mRNA 5′ cap‐binding complex, is a high‐flux‐control step.
| Item Type: | Journal Article | |||||||||||||||
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| Subjects: | Q Science > QH Natural history Q Science > QP Physiology Q Science > QR Microbiology |
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| Divisions: | Faculty of Science, Engineering and Medicine > Science > Life Sciences (2010- ) | |||||||||||||||
| Library of Congress Subject Headings (LCSH): | Proteins -- Synthesis, Ribosomes, Polypeptides, Yeast -- Analysis, Messenger RNA | |||||||||||||||
| Journal or Publication Title: | FEBS Journal | |||||||||||||||
| Publisher: | Blackwell | |||||||||||||||
| ISSN: | 1742-4658 | |||||||||||||||
| Official Date: | March 2020 | |||||||||||||||
| Dates: |
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| Volume: | 287 | |||||||||||||||
| Number: | 5 | |||||||||||||||
| Page Range: | pp. 925-940 | |||||||||||||||
| DOI: | 10.1111/febs.15059 | |||||||||||||||
| Status: | Peer Reviewed | |||||||||||||||
| Publication Status: | Published | |||||||||||||||
| Reuse Statement (publisher, data, author rights): | "This is the peer reviewed version of the following article: Firczuk, H., Teahan, J., Mendes, P. and McCarthy, J.E. (2019), Multisite rate control analysis identifies ribosomal scanning as the sole high‐capacity/low‐flux‐control step in mRNA translation. FEBS J., which has been published in final form at https://doi.org/10.1111/febs.15059. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived Versions." | |||||||||||||||
| Access rights to Published version: | Open Access | |||||||||||||||
| RIOXX Funder/Project Grant: |
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