The 5’-end codon bias minimizes fitness costs to facilitate the persistent dissemination of mcr-3 and investigation of novel MCR inhibitors
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PROVIDER: PRJNA1281520 | ENA |
REPOSITORIES: ENA
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Project description:Background: The rapid evolution and dissemination of mobilized colistin resistance gene (mcr) family has revealed as a severe threat to the global public health. Nevertheless, dramatic reduction in the prevalence of mcr-1, the major member of mcr family, was observed after the withdrawal of colistin in animal fodder in China since 2017, demonstrating that colistin acts as a selective stress to promote the dissemination of mcr-1. As the second largest lineage, mcr-3 was firstly discovered in 2017 and has been identified from numerous sources. However, whether the spreading of mcr-3 is driven by colistin remains unknown. Methods: To this end, we investigated the global prevalence of mcr-3 from 2005 to 2022 by an up-to-date systematic review, along with a nation-wide epidemiological study to establish the change of mcr-3 prevalence in China before and after 2017. To investigate the fitness cost imposed by MCR-3 upon bacterial host, in vitro and in vivo competitive assays were employed, along with morphological study and fluorescent observation. Moreover, by replacing non-optimal codons with optimal codons, synonymous mutations were introduced into the 5’-coding region of mcr-3 to study mechanisms accounting for the distinct fitness cost conferred by MCR-1 and MCR-3. Furthermore, by combining AlphaFold and molecular dynamics (MD) simulation, we provided a complete characterization on the putative lipid A binding pocket localized at the linker domain of MCR-3. Crucially, inhibitors targeting at the putative binding pocket of MCR-1 or MCR-3 were identified from small molecules library using the pipeline of virtual screening. Findings: The global prevalence of mcr-3 increased continuously from 2005 to 2022. The average prevalence was 0.18% during 2005-2014 and rapidly increased to 3.41% during 2020-2022. The prevalence of mcr-3 in China increased from 0.79% in 2016 to 5.87% in 2019. We found that the fitness of mcr-3-bearing E. coli and empty plasmid control was comparable but higher than that of mcr-1-positive strain. Although the putative lipid A binding pocket of MCR-3 was similar to that of in MCR-1, mcr-3 occupies remarkable codon bias at the 5’-end of coding region that disrupted the stability of mRNA, further reduced its protein expression in E. coli, resulting in the low fitness burden of bacterial host. Moreover, the 5’-end codon usage frequency appeared as a critical factor related with the evolution of mcr family. Furthermore, based on the similar lipid A binding pocket among MCR family protein, we identified three novel MCR inhibitors targeting at such pocket by screening from small-molecule library, which effectively restored the colistin susceptibility of mcr-bearing E. coli. Interpretation: For the first time, we found that the prevalence of mcr-3 increased continuously during 2016-2019 in China, demonstrating that the withdrawal of colistin in husbandry failed to prevent the dissemination of mcr-3. Our study evidenced that the 5’-end codon bias appeared as a crucial regulator upon the fitness cost conferred by horizontally transferred genes. Most importantly, the putative lipid A binding pocket verified from current study was a promising target site for designing inhibitors against mcr-positive strains.
2025-06-29 | GSE300593 | GEO
Project description:Usage of synonymous codons represents a characteristic pattern of preference in each organism. It has been inferred that such bias of codon usage has evolved as a result of adaptation for efficient synthesis of proteins. Here we examined synonymous codon usage in genes of the fission yeast Schizosaccharomyces pombe, and compared codon usage bias with expression levels of the gene. In this organism, synonymous codon usage bias was correlated with expression levels of the gene; the bias was most obvious in two-codon amino acids. A similar pattern of the codon usage bias was also observed in Saccharomyces cerevisiae, Arabidopsis thaliana, and Caenorhabditis elegans, but was not obvious in Oryza sativa, Drosophila melanogaster, Takifugu rubripes and Homo sapiens. As codons of the highly expressed genes have greater influence on translational efficiency than codons of genes expressed at lower levels, it is likely that codon usage in the S. pombe genome has been optimized by translational selection through evolution. Relative amounts of mRNA for each ORF were measured by DNA microarray using genomic DNA as a reference, and the copy number of mRNA was calculated using an estimate of the total mRNA number in the cell as 100,000 copies.
2010-06-20 | E-GEOD-13554 | biostudies-arrayexpress
Project description:Usage of synonymous codons represents a characteristic pattern of preference in each organism. It has been inferred that such bias of codon usage has evolved as a result of adaptation for efficient synthesis of proteins. Here we examined synonymous codon usage in genes of the fission yeast Schizosaccharomyces pombe, and compared codon usage bias with expression levels of the gene. In this organism, synonymous codon usage bias was correlated with expression levels of the gene; the bias was most obvious in two-codon amino acids. A similar pattern of the codon usage bias was also observed in Saccharomyces cerevisiae, Arabidopsis thaliana, and Caenorhabditis elegans, but was not obvious in Oryza sativa, Drosophila melanogaster, Takifugu rubripes and Homo sapiens. As codons of the highly expressed genes have greater influence on translational efficiency than codons of genes expressed at lower levels, it is likely that codon usage in the S. pombe genome has been optimized by translational selection through evolution.
2009-04-01 | GSE13554 | GEO
Project description:Differences in codon frequency between genomes, genes, or positions along a gene, modulate transcription and translation efficiency, leading to phenotypic and functional differences. Integrative studies quantifying the phenotypic consequences of codon usage bias at different molecular and cellular levels in human cells are lacking. Here, we present a multiscale analysis of the effects of synonymous codon recoding during heterologous gene expression in human cells. Six synonymous versions of an antibiotic resistance gene were generated, fused to a fluorescent reporter, and independently expressed in HEK293 cells. Multiscale phenotype was analysed by means of: quantitative transcriptome and proteome assessment, as proxies for gene expression; cellular fluorescence, as a proxy for single-cell level expression; and real-time cell proliferation in absence or presence of antibiotic, as a proxy for the cell fitness. We show that differences in codon usage bias strongly impact the molecular and cellular phenotype: (i) they result in large differences in mRNA and in protein levels, as well in mRNA-to-protein ratio; (ii) they introduce unpredicted splicing events; (iii) they lead to reproducible phenotypic heterogeneity; and (iv) they lead to a trade-off between the benefit of antibiotic resistance and the burden of heterologous expression. In human cells in culture, codon usage bias modulates gene expression by modifying mRNA availability and suitability for translation, leading to differences in protein levels and eventually eliciting functional phenotypic changes.
2024-05-24 | PXD038324 | Pride
Project description:Ribosome profiling data reports on the distribution of translating ribosomes, at steady-state, with codon-level resolution. We present a robust method to extract codon translation rates and protein synthesis rates from these data, and identify causal features associated with elongation and translation efficiency in physiological conditions in yeast. We show that neither elongation rate nor translational efficiency is improved by experimental manipulation of the abundance or body sequence of the rare AGG tRNA. Deletion of three of the four copies of the heavily used ACA tRNA shows a modest efficiency decrease that could be explained by other rate-reducing signals at gene start. This suggests that correlation between codon bias and efficiency arises as selection for codons to utilize translation machinery efficiently in highly translated genes. We also show a correlation between efficiency and RNA structure calculated both computationally and from recent structure probing data, as well as the Kozak initiation motif, which may comprise a mechanism to regulate initiation. We test whether tRNA abundance affects elongation or translation efficiency by changing the tRNA levels through deletion or over expression and measuring the ribosomal dwell time at each codon using a robust statistical method that accounts for flow conservation.
2015-01-05 | E-GEOD-63789 | biostudies-arrayexpress
Project description:Ribosome profiling data reports on the distribution of translating ribosomes, at steady-state, with codon-level resolution. We present a robust method to extract codon translation rates and protein synthesis rates from these data, and identify causal features associated with elongation and translation efficiency in physiological conditions in yeast. We show that neither elongation rate nor translational efficiency is improved by experimental manipulation of the abundance or body sequence of the rare AGG tRNA. Deletion of three of the four copies of the heavily used ACA tRNA shows a modest efficiency decrease that could be explained by other rate-reducing signals at gene start. This suggests that correlation between codon bias and efficiency arises as selection for codons to utilize translation machinery efficiently in highly translated genes. We also show a correlation between efficiency and RNA structure calculated both computationally and from recent structure probing data, as well as the Kozak initiation motif, which may comprise a mechanism to regulate initiation.
2015-01-05 | GSE63789 | GEO
Project description:The Moutan Cortex Radicis (MCR) has been used as an analgesic, sedative and anti-inflammatory agent. This study investigated the changes in gene expression by MCR treatment when stimulated with lipopolysaccharide (LPS) in cultured human gingival fibroblasts (HGFs) and the gene expression changes by the MCR when challenged with LPS using a microarray chip.
2012-06-28 | GSE38977 | GEO
Project description:The uneven use of synonymous codons in the transcriptome regulates the efficiency and fidelity of protein translation rates. Yet, the importance of this codon bias on regulating cell state-specific expression programs is currently debated. Here, we asked whether the gene expression program in the well-defined cell states of self-renewal and differentiation in embryonic stem cells is driven by optimized codon usage. Using ribosome and transcriptome profiling, we identified distinct codon signatures for human self-renewing and differentiating embryonic stem cells. One driver for the cell state-specific codon bias was the genomic GC-content of the differentially expressed genes and thus, determined by transcription rather than translation. However, by measuring the codon frequencies at the ribosome’s active sites interacting with transfer RNAs (tRNA), we discovered that the wobble position tRNA modification inosine strongly influenced the codon optimization in self-renewing embryonic stem cells. This effect was conserved in mice and independent of the differentiation stimulus. In summary, we newly reveal how translational mechanisms based on RNA modifications can shape optimized codon usage in embryonic stem cells.
2019-05-17 | GSE112085 | GEO
Project description:The uneven use of synonymous codons in the transcriptome regulates the efficiency and fidelity of protein translation rates. Yet, the importance of this codon bias on regulating cell state-specific expression programs is currently debated. Here, we asked whether the gene expression program in the well-defined cell states of self-renewal and differentiation in embryonic stem cells is driven by optimized codon usage. Using ribosome and transcriptome profiling, we identified distinct codon signatures for human self-renewing and differentiating embryonic stem cells. One driver for the cell state-specific codon bias was the genomic GC-content of the differentially expressed genes and thus, determined by transcription rather than translation. However, by measuring the codon frequencies at the ribosome’s active sites interacting with transfer RNAs (tRNA), we discovered that the wobble position tRNA modification inosine strongly influenced the codon optimization in self-renewing embryonic stem cells. This effect was conserved in mice and independent of the differentiation stimulus. In summary, we newly reveal how translational mechanisms based on RNA modifications can shape optimized codon usage in embryonic stem cells.
2019-05-17 | GSE123608 | GEO
Project description:The Moutan Cortex Radicis (MCR) has been used as an analgesic, sedative and anti-inflammatory agent. This study investigated the changes in gene expression by MCR treatment when stimulated with lipopolysaccharide (LPS) in cultured human gingival fibroblasts (HGFs) and the gene expression changes by the MCR when challenged with LPS using a microarray chip. Human gingival fibroblast were divided into three experimental groups; 1, C: Control, 2, LPS: LPS-treatment only, 3, MCR40: LPS- and MCR40-treatments. Total RNA was isolated from each experimental fibroblast (3 experimental group M-CM-^W 1 sample of each experimental group = total 3 samples).
2012-06-27 | E-GEOD-38977 | biostudies-arrayexpress