scholarly journals Avoidance of stochastic RNA interactions can be harnessed to control protein expression levels in bacteria and archaea

2015 ◽  
Author(s):  
Sinan Uğur Umu ◽  
Anthony M. Poole ◽  
Renwick C. J. Dobson ◽  
Paul P. Gardner
Keyword(s):  
Codon Usage ◽  
Gene Expression Analysis ◽  
Fluorescent Protein ◽  
Translational Efficiency ◽  
Protein Abundance ◽  
Global Factor ◽  
Mrna Structure ◽  
Selection For ◽  
Green Fluorescent ◽  
Control Protein

AbstractA critical assumption of gene expression analysis is that mRNA abundances broadly correlate with protein abundance, but these two are often imperfectly correlated. Some of the discrepancy can be accounted for by two important mRNA features: codon usage and mRNA secondary structure. We present a new global factor, called mRNA:ncRNA avoidance, and provide evidence that avoidance increases translational efficiency. We also demonstrate a strong selection for avoidance of stochastic mRNA:ncRNA interactions across prokaryotes, and that these have a greater impact on protein abundance than mRNA structure or codon usage. By generating synonymously variant green fluorescent protein (GFP) mRNAs with different potential for mRNA:ncRNA interactions, we demonstrate that GFP levels correlate well with interaction avoidance. Therefore, taking stochastic mRNA:ncRNA interactions into account enables precise modulation of protein abundance.

scholarly journals Avoidance of stochastic RNA interactions can be harnessed to control protein expression levels in bacteria and archaea

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Sinan Uğur Umu ◽  
Anthony M Poole ◽  
Renwick CJ Dobson ◽  
Paul P Gardner
Keyword(s):  
Codon Usage ◽  
Gene Expression Analysis ◽  
Fluorescent Protein ◽  
Translational Efficiency ◽  
Protein Abundance ◽  
Global Factor ◽  
Mrna Structure ◽  
Selection For ◽  
Green Fluorescent ◽  
Control Protein

A critical assumption of gene expression analysis is that mRNA abundances broadly correlate with protein abundance, but these two are often imperfectly correlated. Some of the discrepancy can be accounted for by two important mRNA features: codon usage and mRNA secondary structure. We present a new global factor, called mRNA:ncRNA avoidance, and provide evidence that avoidance increases translational efficiency. We also demonstrate a strong selection for the avoidance of stochastic mRNA:ncRNA interactions across prokaryotes, and that these have a greater impact on protein abundance than mRNA structure or codon usage. By generating synonymously variant green fluorescent protein (GFP) mRNAs with different potential for mRNA:ncRNA interactions, we demonstrate that GFP levels correlate well with interaction avoidance. Therefore, taking stochastic mRNA:ncRNA interactions into account enables precise modulation of protein abundance.

scholarly journals A Photostable Green Fluorescent Protein Variant for Analysis of Protein Localization in Candida albicans

2009 ◽  
Vol 9 (1) ◽  
pp. 224-226 ◽  
Author(s):  
Chengda Zhang ◽  
James B. Konopka
Keyword(s):  
Candida Albicans ◽  
Green Fluorescent Protein ◽  
Codon Usage ◽  
Signal Intensity ◽  
Fluorescent Protein ◽  
Protein Localization ◽  
Protein Variant ◽  
Green Fluorescent ◽  
Green Fluorescent Protein Variant

ABSTRACT Fusions to the green fluorescent protein (GFP) are an effective way to monitor protein localization. However, altered codon usage in Candida species has delayed implementation of new variants. Examination of three new GFP variants in Candida albicans showed that one has higher signal intensity and increased resistance to photobleaching.

scholarly journals Widespread positive selection for mRNA secondary structure at synonymous sites in domesticated yeast

2019 ◽  
Author(s):  
Minghao Yu ◽  
Wenna Guo ◽  
Qiang Wang ◽  
Jian-Qun Chen
Keyword(s):  
Secondary Structure ◽  
Positive Selection ◽  
Yeast Genome ◽  
Cellular Respiration ◽  
Translational Efficiency ◽  
Mrna Secondary Structure ◽  
Frequency Spectra ◽  
Mrna Structure ◽  
Synonymous Sites ◽  
Selection For

AbstractmRNA secondary structure assumes a critical role in gene regulation, especially for translational efficiency. Previous studies have a growing appreciation of purifying selection for the conserved mRNA structure across lineages of different species. However, the effect of mRNA structure on positive evolution remains unclear. Here, we construct a large-scale dataset of single nucleotide polymorphisms (SNPs) at synonymous sites in the population of Saccharomyces cerevisiae, combined with the experimental assessment of mRNA structure, and perform empirical population genetics data analysis through unfolded site-frequency spectra. Our results suggest that functional mRNA stem drives faster evolution of increasing GC contents itself with the purpose of regulating translational speed, which is greatly influenced by length. At the synonymous site without codon usage bias, this kind of positive selection still exists. Furthermore, mRNA secondary structure is subject to positive selection widespread among the yeast genome, particularly related to mitochondria activities, which is possibly aimed to achieve a balance between cellular respiration and alcoholic fermentation precisely at a non-protein level. It is conducive to the adaption of the dramatic environment alterations from wild to man-made environments during the domestication.

scholarly journals Genetics of single-cell protein abundance variation in large yeast populations

2013 ◽  
Author(s):  
Frank Albert ◽  
Sebastian Treusch ◽  
Arthur H Shockley ◽  
Joshua S Bloom ◽  
Leonid Kruglyak
Keyword(s):  
Single Cell ◽  
Fluorescent Protein ◽  
Single Cell Protein ◽  
Cell Protein ◽  
Cell Physiology ◽  
Protein Abundance ◽  
Large Populations ◽  
Abundance Variation ◽  
Green Fluorescent ◽  
Fluorescent Protein Tags

Many DNA sequence variants influence phenotypes by altering gene expression. Our understanding of these variants is limited by sample sizes of current studies and by measurements of mRNA rather than protein abundance. We developed a powerful method for identifying genetic loci that influence protein expression in very large populations of the yeast Saccharomyes cerevisiae. The method measures single-cell protein abundance through the use of green-fluorescent-protein tags. We applied this method to 160 genes and detected many more loci per gene than previous studies. We also observed closer correspondence between loci that influence protein abundance and loci that influence mRNA abundance of a given gene. Most loci cluster at hotspot locations that influence multiple proteins—in some cases, more than half of those examined. The variants that underlie these hotspots have profound effects on the gene regulatory network and provide insights into genetic variation in cell physiology between yeast strains.

scholarly journals Biochemical and functional characterization of a mitochondrial citrate carrier in Arabidopsis thaliana

2020 ◽  
Vol 477 (9) ◽  
pp. 1759-1777 ◽  
Author(s):  
Danielle S. Brito ◽  
Gennaro Agrimi ◽  
Lennart Charton ◽  
Dominik Brilhaus ◽  
Maria Gabriella Bitetto ◽  
...  
Keyword(s):  
Gene Expression Analysis ◽  
Nitrogen Assimilation ◽  
Fluorescent Protein ◽  
Developmental Stages ◽  
Functional Characterization ◽  
Radicle Growth ◽  
Citrate Carrier ◽  
Green Fluorescent ◽  
Lower Expression

A homolog of the mitochondrial succinate/fumarate carrier from yeast (Sfc1p) has been found in the Arabidopsis genome, named AtSFC1. The AtSFC1 gene was expressed in Escherichia coli, and the gene product was purified and reconstituted in liposomes. Its transport properties and kinetic parameters demonstrated that AtSFC1 transports citrate, isocitrate and aconitate and, to a lesser extent, succinate and fumarate. This carrier catalyzes a fast counter-exchange transport as well as a low uniport of substrates, exhibits a higher transport affinity for tricarboxylates than dicarboxylates, and is inhibited by pyridoxal 5′-phosphate and other inhibitors of mitochondrial carriers to various degrees. Gene expression analysis indicated that the AtSFC1 transcript is mainly present in heterotrophic tissues, and fusion with a green-fluorescent protein localized AtSFC1 to the mitochondria. Furthermore, 35S-AtSFC1 antisense lines were generated and characterized at metabolic and physiological levels in different organs and at various developmental stages. Lower expression of AtSFC1 reduced seed germination and impaired radicle growth, a phenotype that was related to reduced respiration rate. These findings demonstrate that AtSFC1 might be involved in storage oil mobilization at the early stages of seedling growth and in nitrogen assimilation in root tissue by catalyzing citrate/isocitrate or citrate/succinate exchanges.

scholarly journals Differential proteostatic regulation of insoluble and abundant proteins

2019 ◽  
Vol 35 (20) ◽  
pp. 4098-4107 ◽  
Author(s):  
Reshmi Ramakrishnan ◽  
Bert Houben ◽  
Frederic Rousseau ◽  
Joost Schymkowitz
Keyword(s):  
Fluorescent Protein ◽  
Meta Analysis ◽  
Trigger Factor ◽  
Supplementary Information ◽  
Translational Efficiency ◽  
Translation Rate ◽  
Synonymous Mutations ◽  
Metabolic Functions ◽  
Client Proteins ◽  
Green Fluorescent

Abstract Motivation Despite intense effort, it has been difficult to explain chaperone dependencies of proteins from sequence or structural properties. Results We constructed a database collecting all publicly available data of experimental chaperone interaction and dependency data for the Escherichia coli proteome, and enriched it with an extensive set of protein-specific as well as cell-context-dependent proteostatic parameters. Employing this new resource, we performed a comprehensive meta-analysis of the key determinants of chaperone interaction. Our study confirms that GroEL client proteins are biased toward insoluble proteins of low abundance, but for client proteins of the Trigger Factor/DnaK axis, we instead find that cellular parameters such as high protein abundance, translational efficiency and mRNA turnover are key determinants. We experimentally confirmed the finding that chaperone dependence is a function of translation rate and not protein-intrinsic parameters by tuning chaperone dependence of Green Fluorescent Protein (GFP) in E.coli by synonymous mutations only. The juxtaposition of both protein-intrinsic and cell-contextual chaperone triage mechanisms explains how the E.coli proteome achieves combining reliable production of abundant and conserved proteins, while also enabling the evolution of diverging metabolic functions. Availability and implementation The database will be made available via http://phdb.switchlab.org. Supplementary information Supplementary data are available at Bioinformatics online.

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