scholarly journals RNA polyadenylation and its consequences in prokaryotes

2018 ◽  
Vol 373 (1762) ◽  
pp. 20180166 ◽  
Author(s):  
Eliane Hajnsdorf ◽  
Vladimir R. Kaberdin
Keyword(s):  
Gene Expression ◽  
State Level ◽  
Rna Degradation ◽  
Plasmid Copy Number ◽  
Plasmid Copy ◽  
Protein Coding ◽  
Mrna Metabolism ◽  
Isolation And Characterization ◽  
Polymerase I ◽  
The Impact

Post-transcriptional addition of poly(A) tails to the 3′ end of RNA is one of the fundamental events controlling the functionality and fate of RNA in all kingdoms of life. Although an enzyme with poly(A)-adding activity was discovered in Escherichia coli more than 50 years ago, its existence and role in prokaryotic RNA metabolism were neglected for many years. As a result, it was not until 1992 that E. coli poly(A) polymerase I was purified to homogeneity and its gene was finally identified. Further work revealed that, similar to its role in surveillance of aberrant nuclear RNAs of eukaryotes, the addition of poly(A) tails often destabilizes prokaryotic RNAs and their decay intermediates, thus facilitating RNA turnover. Moreover, numerous studies carried out over the last three decades have shown that polyadenylation greatly contributes to the control of prokaryotic gene expression by affecting the steady-state level of diverse protein-coding and non-coding transcripts including antisense RNAs involved in plasmid copy number control, expression of toxin–antitoxin systems and bacteriophage development. Here, we review the main findings related to the discovery of polyadenylation in prokaryotes, isolation, and characterization and regulation of bacterial poly(A)-adding activities, and discuss the impact of polyadenylation on prokaryotic mRNA metabolism and gene expression. This article is part of the theme issue ‘5′ and 3′ modifications controlling RNA degradation’.

scholarly journals A Plasmid System with Tunable Copy Number

2021 ◽  
Author(s):  
Miles V Rouches ◽  
Yasu Xu ◽  
Louis Cortes ◽  
Guillaume Lambert
Keyword(s):  
Gene Expression ◽  
Copy Number ◽  
Plasmid Copy Number ◽  
Metabolic Effects ◽  
Massively Parallel ◽  
Simple Relationship ◽  
Plasmid Copy ◽  
Control Of Gene Expression ◽  
Recombinant Gene Expression ◽  
The Impact

Plasmids are one of the most commonly used and time-tested molecular biology platforms for genetic engineering and recombinant gene expression in bacteria. Despite their ubiquity, little consideration is given to metabolic effects and fitness costs of plasmid copy numbers on engineered genetic systems. Here, we introduce two systems that allow for the finely-tuned control of plasmid copy number: a plasmid with an anhydrotetracycline-controlled copy number, and a massively parallel assay that is used to generate a continuous spectrum of ColE1-based copy number variants. Using these systems, we investigate the effects of plasmid copy number on cellular growth rates, gene expression, biosynthesis, and genetic circuit performance. We perform single-cell timelapse measurements to characterize plasmid loss, runaway plasmid replication, and quantify the impact of plasmid copy number on the variability of gene expression. Using our massively parallel assay, we find that each plasmid imposes a 0.063% linear metabolic burden on their hosts, hinting at a simple relationship between metabolic burdens and plasmid DNA synthesis. Our plasmid system with tunable copy number should allow for a precise control of gene expression and highlight the importance of tuning plasmid copy number as tool for the optimization of synthetic biological systems.

scholarly journals Characterization of the nuclear and cytosolic transcriptomes in human brain tissue reveals new insights into the subcellular distribution of RNA transcripts

2020 ◽  
Author(s):  
Ammar Zaghlool ◽  
Adnan Niazi ◽  
Åsa K. Björklund ◽  
Jakub Orzechowski Westholm ◽  
Adam Ameur ◽  
...  
Keyword(s):  
Gene Expression ◽  
Subcellular Localization ◽  
Human Brain ◽  
Expression Analysis ◽  
Differential Expression Analysis ◽  
Adult Brain ◽  
Sequencing Data ◽  
Human Brain Tissue ◽  
Protein Coding ◽  
The Impact

AbstractTranscriptome analysis has mainly relied on analyzing RNA sequencing data from whole cells, overlooking the impact of subcellular RNA localization and its influence on our understanding of gene function, and interpretation of gene expression signatures in cells. Here, we performed a comprehensive analysis of cytosolic and nuclear transcriptomes in human fetal and adult brain samples. We show significant differences in RNA expression for protein-coding and lncRNA genes between cytosol and nucleus. Transcripts displaying differential subcellular localization belong to particular functional categories and display tissue-specific localization patterns. We also show that transcripts encoding the nuclear-encoded mitochondrial proteins are significantly enriched in the cytosol compared to the rest of protein-coding genes. Further investigation of the use of the cytosolic or the nuclear transcriptome for differential gene expression analysis indicates important differences in results depending on the cellular compartment. These differences were manifested at the level of transcript types and the number of differentially expressed genes. Our data provide a resource of RNA subcellular localization in the human brain and highlight differences in using the cytosolic or the nuclear transcriptomes for differential expression analysis.

scholarly journals Codon usage is an important determinant of gene expression levels largely through its effects on transcription

2016 ◽  
Vol 113 (41) ◽  
pp. E6117-E6125 ◽  
Author(s):  
Zhipeng Zhou ◽  
Yunkun Dang ◽  
Mian Zhou ◽  
Lin Li ◽  
Chien-hung Yu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Codon Usage ◽  
Important Determinant ◽  
Mrna Translation ◽  
Mrna Levels ◽  
Protein Coding ◽  
Expression Levels ◽  
Highly Expressed Genes ◽  
The Impact ◽  
Gene Expression Levels

Codon usage biases are found in all eukaryotic and prokaryotic genomes, and preferred codons are more frequently used in highly expressed genes. The effects of codon usage on gene expression were previously thought to be mainly mediated by its impacts on translation. Here, we show that codon usage strongly correlates with both protein and mRNA levels genome-wide in the filamentous fungus Neurospora. Gene codon optimization also results in strong up-regulation of protein and RNA levels, suggesting that codon usage is an important determinant of gene expression. Surprisingly, we found that the impact of codon usage on gene expression results mainly from effects on transcription and is largely independent of mRNA translation and mRNA stability. Furthermore, we show that histone H3 lysine 9 trimethylation is one of the mechanisms responsible for the codon usage-mediated transcriptional silencing of some genes with nonoptimal codons. Together, these results uncovered an unexpected important role of codon usage in ORF sequences in determining transcription levels and suggest that codon biases are an adaptation of protein coding sequences to both transcription and translation machineries. Therefore, synonymous codons not only specify protein sequences and translation dynamics, but also help determine gene expression levels.

scholarly journals Small RNA Transcriptome of the Oral Microbiome during Periodontitis Progression

2015 ◽  
Vol 81 (19) ◽  
pp. 6688-6699 ◽  
Author(s):  
Ana E. Duran-Pinedo ◽  
Susan Yost ◽  
Jorge Frias-Lopez
Keyword(s):  
Posttranscriptional Regulation ◽  
Protein Targeting ◽  
Signal Recognition Particle ◽  
Regulatory Elements ◽  
Plasmid Copy Number ◽  
Oral Microbiome ◽  
Sequencing Analysis ◽  
Plasmid Copy ◽  
Protein Coding ◽  
Small Noncoding Rnas

ABSTRACTThe oral microbiome is one of the most complex microbial communities in the human body, and due to circumstances not completely understood, the healthy microbial community becomes dysbiotic, giving rise to periodontitis, a polymicrobial inflammatory disease. We previously reported the results of community-wide gene expression changes in the oral microbiome during periodontitis progression and identified signatures associated with increasing severity of the disease. Small noncoding RNAs (sRNAs) are key players in posttranscriptional regulation, especially in fast-changing environments such as the oral cavity. Here, we expanded our analysis to the study of the sRNA metatranscriptome during periodontitis progression on the same samples for which mRNA expression changes were analyzed. We observed differential expression of 12,097 sRNAs, identifying a total of 20 Rfam sRNA families as being overrepresented in progression and 23 at baseline. Gene ontology activities regulated by the differentially expressed (DE) sRNAs included amino acid metabolism, ethanolamine catabolism, signal recognition particle-dependent cotranslational protein targeting to membrane, intron splicing, carbohydrate metabolism, control of plasmid copy number, and response to stress. In integrating patterns of expression of protein coding transcripts and sRNAs, we found that functional activities of genes that correlated positively with profiles of expression of DE sRNAs were involved in pathogenesis, proteolysis, ferrous iron transport, and oligopeptide transport. These findings represent the first integrated sequencing analysis of the community-wide sRNA transcriptome of the oral microbiome during periodontitis progression and show that sRNAs are key regulatory elements of the dysbiotic process leading to disease.

RNA polymerase I-mediated protein-coding gene expression in Trypanosoma brucei

1992 ◽  
Vol 8 (12) ◽  
pp. 414-418 ◽  
Author(s):  
H-M. Chung ◽  
M.G-S. Lee ◽  
L.H.T. Van der Ploeg
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Trypanosoma Brucei ◽  
Rna Polymerase I ◽  
Protein Coding ◽  
Polymerase I

scholarly journals Plasmid co-infection: linking biological mechanisms to ecological and evolutionary dynamics

2021 ◽  
Vol 377 (1842) ◽  
Author(s):  
Claudia Igler ◽  
Jana S. Huisman ◽  
Berit Siedentop ◽  
Sebastian Bonhoeffer ◽  
Sonja Lehtinen
Keyword(s):  
Evolutionary Dynamics ◽  
Theoretical Work ◽  
Plasmid Copy Number ◽  
Biological Processes ◽  
Ecological Interactions ◽  
Biological Mechanisms ◽  
Plasmid Copy ◽  
Host Interactions ◽  
Evolutionary Epidemiology ◽  
The Impact

As infectious agents of bacteria and vehicles of horizontal gene transfer, plasmids play a key role in bacterial ecology and evolution. Plasmid dynamics are shaped not only by plasmid–host interactions but also by ecological interactions between plasmid variants. These interactions are complex: plasmids can co-infect the same cell and the consequences for the co-resident plasmid can be either beneficial or detrimental. Many of the biological processes that govern plasmid co-infection—from systems that exclude infection by other plasmids to interactions in the regulation of plasmid copy number—are well characterized at a mechanistic level. Modelling plays a central role in translating such mechanistic insights into predictions about plasmid dynamics and the impact of these dynamics on bacterial evolution. Theoretical work in evolutionary epidemiology has shown that formulating models of co-infection is not trivial, as some modelling choices can introduce unintended ecological assumptions. Here, we review how the biological processes that govern co-infection can be represented in a mathematical model, discuss potential modelling pitfalls, and analyse this model to provide general insights into how co-infection impacts ecological and evolutionary outcomes. In particular, we demonstrate how beneficial and detrimental effects of co-infection give rise to frequency-dependent selection on plasmid variants. This article is part of the theme issue ‘The secret lives of microbial mobile genetic elements’.

scholarly journals Improved Plasmid-Based Inducible and Constitutive Gene Expression in Corynebacterium glutamicum

2021 ◽  
Vol 9 (1) ◽  
pp. 204
Author(s):  
Nadja A. Henke ◽  
Irene Krahn ◽  
Volker F. Wendisch
Keyword(s):  
Gene Expression ◽  
Corynebacterium Glutamicum ◽  
Xanthomonas Campestris ◽  
Heterologous Gene Expression ◽  
Constitutive Expression ◽  
Xylose Isomerase ◽  
Plasmid Copy Number ◽  
Expression Vectors ◽  
Plasmid Copy ◽  
Synthetic Promoters

Corynebacterium glutamicum has been safely used in white biotechnology for the last 60 years and the portfolio of new pathways and products is increasing rapidly. Hence, expression vectors play a central role in discovering endogenous gene functions and in establishing heterologous gene expression. In this work, new expression vectors were designed based on two strategies: (i) a library screening of constitutive native and synthetic promoters and (ii) an increase of the plasmid copy number. Both strategies were combined and resulted in a very strong expression and overproduction of the fluorescence protein GfpUV. As a second test case, the improved vector for constitutive expression was used to overexpress the endogenous xylulokinase gene xylB in a synthetic operon with xylose isomerase gene xylA from Xanthomonas campestris. The xylose isomerase activity in crude extracts was increased by about three-fold as compared to that of the parental vector. In terms of application, the improved vector for constitutive xylA and xylB expression was used for production of the N-methylated amino acid sarcosine from monomethylamine, acetate, and xylose. As a consequence, the volumetric productivity of sarcosine production was 50% higher as compared to that of the strain carrying the parental vector.

scholarly journals Impact of RNA degradation on measurements of gene expression

2014 ◽  
Author(s):  
Irene Gallego Romero ◽  
Athma A. Pai ◽  
Jenny Tung ◽  
Yoav Gilad
Keyword(s):  
Gene Expression ◽  
Expression Profiling ◽  
Rna Extraction ◽  
Rna Degradation ◽  
Significant Loss ◽  
Whole Genome ◽  
Expression Levels ◽  
Rna Quality ◽  
The Impact ◽  
Gene Expression Levels

The use of low quality RNA samples in whole-genome gene expression profiling remains controversial. It is unclear if transcript degradation in low quality RNA samples occurs uniformly, in which case the effects of degradation can be normalized, or whether different transcripts are degraded at different rates, potentially biasing measurements of expression levels. This concern has rendered the use of low quality RNA samples in whole-genome expression profiling problematic. Yet, low quality samples are at times the sole means of addressing specific questions – e.g., samples collected in the course of fieldwork. We sought to quantify the impact of variation in RNA quality on estimates of gene expression levels based on RNA-seq data. To do so, we collected expression data from tissue samples that were allowed to decay for varying amounts of time prior to RNA extraction. The RNA samples we collected spanned the entire range of RNA Integrity Number (RIN) values (a quality metric commonly used to assess RNA quality). We observed widespread effects of RNA quality on measurements of gene expression levels, as well as a slight but significant loss of library complexity in more degraded samples. While standard normalizations failed to account for the effects of degradation, we found that a simple linear model that controls for the effects of RIN can correct for the majority of these effects. We conclude that in instances where RIN and the effect of interest are not associated, this approach can help recover biologically meaningful signals in data from degraded RNA samples.

scholarly journals Functional and transcriptional profiling of non-coding RNAs in yeast reveal context-dependent phenotypes and widespread in trans effects on the protein regulatory network

2020 ◽  
Author(s):  
Laura Natalia Balarezo-Cisneros ◽  
Steven Parker ◽  
Marcin G Fraczek ◽  
Soukaina Timouma ◽  
Ping Wang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factors ◽  
Environmental Conditions ◽  
Mode Of Action ◽  
Protein Coding ◽  
Protein Coding Genes ◽  
Genome Wide ◽  
In Trans ◽  
Non Coding Rnas ◽  
The Impact

AbstractNon-coding RNAs (ncRNAs), including the more recently identified Stable Unannotated Transcripts (SUTs) and Cryptic Unstable Transcripts (CUTs), are increasingly being shown to play pivotal roles in the transcriptional and post-transcriptional regulation of genes in eukaryotes. Here, we carried out a large-scale screening of ncRNAs in Saccharomyces cerevisiae, and provide evidence for SUT and CUT function. Phenotypic data on 372 ncRNA deletion strains in 23 different growth conditions were collected, identifying ncRNAs responsible for significant cellular fitness changes. Transcriptome profiles were assembled for 18 haploid ncRNA deletion mutants and 2 essential ncRNA heterozygous deletants. Guided by the resulting RNA-seq data we analysed the genome-wide dysregulation of protein coding genes and non-coding transcripts. Novel functional ncRNAs, SUT125, SUT126, SUT035 and SUT532 that act in trans by modulating transcription factors were identified. Furthermore, we described the impact of SUTs and CUTs in modulating coding gene expression in response of different environmental conditions, regulating important biological process such as respiration (SUT125, SUT126, SUT035, SUT432), steroid biosynthesis (CUT494, SUT530, SUT468) or rRNA processing (SUT075 and snR30). Overall, this data captures and integrates the regulatory and phenotypic network of ncRNAs and protein coding genes, providing genome-wide evidence of the impact of ncRNAs on cellular homeostasis.Author SummaryThe yeast genome contains 25% of non-coding RNA molecules (ncRNAs), which do not translate into proteins but are involved in regulation of gene expression. ncRNAs can affect nearby genes by physically interfering with their transcription (cis mode of action), or they interact with DNA, proteins or others RNAs to regulate the expression of distant genes (trans mode of action). Examples of cis-acting ncRNAs have been broadly described, however genome-wide studies to identify functional trans-acting ncRNAs involved in global gene regulation are still lacking. Here, we used the ncRNA yeast deletion collection to score their impact on cellular function in different environmental conditions. A group of 20 ncRNAs mutants with broad fitness diversity were selected to investigate their effect on the protein and ncRNA expression network. We showed a high correlation between altered phenotypes and global transcriptional changes, in an environmental dependent manner. We confirmed the widespread trans acting expressional regulation of ncRNAs in the genome and their role in affecting transcription factors. These findings support the notion of the involvement on ncRNAs in fine tuning the cellular expression via regulations of TFs, as an advantageous RNA-mediated mechanism that can be fast and cost-effective for the cells.

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