The essential role of mRNA degradation in understanding and engineering E. coli metabolism

Roux, C; Etienne, Thibault A.; Hajnsdorf, Eliane; Ropers, Delphine; Carpousis, Agamemnon J.; Cocaign‐Bousquet, Muriel; Girbal, Laurence

doi:10.1016/j.biotechadv.2021.107805

Cited by 7 publications

(3 citation statements)

References 158 publications

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“…Improving the understanding of mRNA degradation can play a critical role in optimizing metabolic engineering strategies (Roux et al, 2021 ). Targeting mRNA degradation has been shown to enhance protein production without competing for cellular resources in the host strain (Mao & Inouye, 2012 ; Venturelli et al, 2017 ; Wu et al, 2020 ).…”

Section: Discussionmentioning

confidence: 99%

Engineering resource allocation in artificially minimized cells: Is genome reduction the best strategy?

Marquez‐Zavala

Utrilla

2023

Microbial Biotechnology

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The elimination of the expression of cellular functions that are not needed in a certain well-defined artificial environment, such as those used in industrial production facilities, has been the goal of many cellular minimization projects. The generation of a minimal cell with reduced burden and less host-function interactions has been pursued as a tool to improve microbial production strains. In this work, we analysed two cellular complexity reduction strategies: genome and proteome reduction. With the aid of an absolute proteomics data set and a genome-scale model of metabolism and protein expression (MEmodel), we quantitatively assessed the difference of reducing genome to the correspondence of reducing proteome. We compare the approaches in terms of energy consumption, defined in ATP equivalents. We aim to show what is the best strategy for improving resource allocation in minimized cells. Our results show that genome reduction by length is not proportional to reducing resource use. When we normalize calculated energy savings, we show that strains with the larger calculated proteome reduction show the largest resource use reduction. Furthermore, we propose that reducing highly expressed proteins should be the target as the translation of a gene uses most of the energy. The strategies proposed here should guide cell design when the aim of a project is to reduce the maximum amount or cellular resources.

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Section: Discussionmentioning

confidence: 99%

Engineering resource allocation in artificially minimized cells: Is genome reduction the best strategy?

Marquez‐Zavala

Utrilla

2023

Microbial Biotechnology

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“…Thus, we speculated that RNA degradation led to the accumulation of adenosine and guanosine, but this degradation could have occurred in Synechocystis as well (Zhang et al, 2022). Interestingly, mRNA degradation regulates the activity of the central carbon metabolism and metabolic responses to stress (Roux et al, 2022). As arginine and leucine have the potential to control almost all amino acid biosynthesis pathways (Radoš et al, 2022), we hypothesized that their abundance is related to the capacity of cells to begin growing as promptly as the environmental conditions return favorably.…”

Section: Drought Changes the Composition Structure And Metabolic Prof...mentioning

confidence: 99%

A desiccated dual-species subaerial biofilm reprograms its metabolism and affects water dynamics in limestone

Villa

Ludwig

Mazzini

et al. 2023

Science of The Total Environment

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“…Although RNAseE has broad substrate specificity, various sequence features have been shown to increase its relative specific activity on individual mRNA species, including unstructured AU rich regions, and, most critically, the presence of a 5′-monophosphate (Bae et al, 2023;Callaghan et al, 2005;Richards & Belasco, 2023). However, other molecule-specific features, such as RNA-binding protein binding sites, codon usage, and secondary structure profiles can reduce RNAse E mediated turnover of endogenous mRNA (Börner et al, 2023;Roux et al, 2022). For protein production in E. coli, recombinant mRNA stability is commonly enhanced by using bacterial 5′ untranslated regions (UTRs) containing secondary structures that prevent RNAse binding (Viegas et al, 2018), however, these elements are incompatible with manufacture of synthetic mRNA for mammalian cell applications.…”

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confidence: 99%

Engineering an Escherichia coli based in vivo mRNA manufacturing platform

Curry,

Muir,

et al. 2024

Biotech & Bioengineering

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Synthetic mRNA is currently produced in standardized in vitro transcription systems. However, this one‐size‐fits‐all approach has associated drawbacks in supply chain shortages, high reagent costs, complex product‐related impurity profiles, and limited design options for molecule‐specific optimization of product yield and quality. Herein, we describe for the first time development of an in vivo mRNA manufacturing platform, utilizing an Escherichia coli cell chassis. Coordinated mRNA, DNA, cell and media engineering, primarily focussed on disrupting interactions between synthetic mRNA molecules and host cell RNA degradation machinery, increased product yields >40‐fold compared to standard “unengineered” E. coli expression systems. Mechanistic dissection of cell factory performance showed that product mRNA accumulation levels approached theoretical limits, accounting for ~30% of intracellular total RNA mass, and that this was achieved via host‐cell's reallocating biosynthetic capacity away from endogenous RNA and cell biomass generation activities. We demonstrate that varying sized functional mRNA molecules can be produced in this system and subsequently purified. Accordingly, this study introduces a new mRNA production technology, expanding the solution space available for mRNA manufacturing.

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The essential role of mRNA degradation in understanding and engineering E. coli metabolism

Cited by 7 publications

References 158 publications

Engineering resource allocation in artificially minimized cells: Is genome reduction the best strategy?

Engineering resource allocation in artificially minimized cells: Is genome reduction the best strategy?

A desiccated dual-species subaerial biofilm reprograms its metabolism and affects water dynamics in limestone

Engineering an Escherichia coli based in vivo mRNA manufacturing platform

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