Multi-site rate control analysis identifies ribosomal scanning as the sole high-capacity/low-flux-control step in mRNA translation
| Autor: | Pedro Mendes, John E.G. McCarthy, Helena Firczuk, James Teahan |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
protein synthesis yeast Biochemistry Ribosome polypeptide initiation 03 medical and health sciences 0302 clinical medicine Eukaryotic translation Start codon Protein biosynthesis Translation factor RNA Messenger Peptide Chain Initiation Translational Molecular Biology rate control Messenger RNA Chemistry QH Computational Biology Translation (biology) Cell Biology Original Articles Ribosomal RNA Models Theoretical QP 3. Good health Cell biology QR 030104 developmental biology 030220 oncology & carcinogenesis Protein Biosynthesis ribosomal scanning Original Article Ribosomes |
| Zdroj: | The Febs Journal |
| ISSN: | 1742-4658 |
| Popis: | 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. Translation of a reading frame on eukaryotic mRNA is preceded by a poorly understood scanning process in which a subset of translation factors helps guide ribosomes to the start codon. We use novel experimental and computational tools to study 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 the downstream phases of polypeptide initiation, elongation and termination. |
| Databáze: | OpenAIRE |
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