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  • Article  (46)
  • Vogel, J.  (46)
  • Gene Expression
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  • Article  (46)
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  • 1
    Language: English
    In: Proceedings of the National Academy of Sciences of the United States of America, 11 October 2016, Vol.113(41), pp.11591-11596
    Description: The functional annotation of transcriptomes and identification of noncoding RNA (ncRNA) classes has been greatly facilitated by the advent of next-generation RNA sequencing which, by reading the nucleotide order of transcripts, theoretically allows the rapid profiling of all transcripts in a cell. However, primary sequence per se is a poor predictor of function, as ncRNAs dramatically vary in length and structure and often lack identifiable motifs. Therefore, to visualize an informative RNA landscape of organisms with potentially new RNA biology that are emerging from microbiome and environmental studies requires the use of more functionally relevant criteria. One such criterion is the association of RNAs with functionally important cognate RNA-binding proteins. Here we analyze the full ensemble of cellular RNAs using gradient profiling by sequencing (Grad-seq) in the bacterial pathogen Salmonella enterica, partitioning its coding and noncoding transcripts based on their network of RNA-protein interactions. In addition to capturing established RNA classes based on their biochemical profiles, the Grad-seq approach enabled the discovery of an overlooked large collective of structured small RNAs that form stable complexes with the conserved protein ProQ. We show that ProQ is an abundant RNA-binding protein with a wide range of ligands and a global influence on Salmonella gene expression. Given its generic ability to chart a functional RNA landscape irrespective of transcript length and sequence diversity, Grad-seq promises to define functional RNA classes and major RNA-binding proteins in both model species and genetically intractable organisms.
    Keywords: Hfq ; Proq ; RNA–Protein Interaction ; Noncoding RNA ; Small RNA ; Bacterial Proteins -- Metabolism ; High-Throughput Nucleotide Sequencing -- Methods ; RNA, Bacterial -- Metabolism ; RNA-Binding Proteins -- Metabolism ; Salmonella Enterica -- Metabolism
    ISSN: 00278424
    E-ISSN: 1091-6490
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  • 2
    In: EMBO Journal, 03 July 2013, Vol.32(13), pp.1802-1804
    Description: CRISPR systems not only defend bacteria from foreign DNA but also contribute to pathogenicity, by regulating endogenous gene expression to evade host innate immune responses.
    Keywords: Animals–Immunology ; Female–Pathogenicity ; Gammaproteobacteria–Immunology ; Gammaproteobacteria–Immunology ; Immune Evasion–Immunology ; Immunity, Innate–Immunology ; Germany ; Prokaryotes ; Gene Expression ; Eukaryotes ; Bacteria ; Molecular Biology;
    ISSN: 0261-4189
    E-ISSN: 1460-2075
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  • 3
    Language: English
    In: Proceedings of the National Academy of Sciences of the United States of America, 10 September 2013, Vol.110(37), pp.E3487-96
    Description: Small RNAs (sRNAs) constitute a large and heterogeneous class of bacterial gene expression regulators. Much like eukaryotic microRNAs, these sRNAs typically target multiple mRNAs through short seed pairing, thereby acting as global posttranscriptional regulators. In some bacteria, evidence for hundreds to possibly more than 1,000 different sRNAs has been obtained by transcriptome sequencing. However, the experimental identification of possible targets and, therefore, their confirmation as functional regulators of gene expression has remained laborious. Here, we present a strategy that integrates phylogenetic information to predict sRNA targets at the genomic scale and reconstructs regulatory networks upon functional enrichment and network analysis (CopraRNA, for Comparative Prediction Algorithm for sRNA Targets). Furthermore, CopraRNA precisely predicts the sRNA domains for target recognition and interaction. When applied to several model sRNAs, CopraRNA revealed additional targets and functions for the sRNAs CyaR, FnrS, RybB, RyhB, SgrS, and Spot42. Moreover, the mRNAs gdhA, lrp, marA, nagZ, ptsI, sdhA, and yobF-cspC were suggested as regulatory hubs targeted by up to seven different sRNAs. The verification of many previously undetected targets by CopraRNA, even for extensively investigated sRNAs, demonstrates its advantages and shows that CopraRNA-based analyses can compete with experimental target prediction approaches. A Web interface allows high-confidence target prediction and efficient classification of bacterial sRNAs.
    Keywords: E. Coli ; RNA–RNA Interaction ; Regulatory RNA ; RNA, Bacterial -- Genetics
    ISSN: 00278424
    E-ISSN: 1091-6490
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  • 4
    Language: English
    In: Nature, 28 January 2016, Vol.529(7587), pp.496-501
    Description: Bacteria express many small RNAs for which the regulatory roles in pathogenesis have remained poorly understood due to a paucity of robust phenotypes in standard virulence assays. Here we use a generic 'dual RNA-seq' approach to profile RNA expression simultaneously in pathogen and host during Salmonella enterica serovar Typhimurium infection and reveal the molecular impact of bacterial riboregulators. We identify a PhoP-activated small RNA, PinT, which upon bacterial internalization temporally controls the expression of both invasion-associated effectors and virulence genes required for intracellular survival. This riboregulatory activity causes pervasive changes in coding and noncoding transcripts of the host. Interspecies correlation analysis links PinT to host cell JAK-STAT signalling, and we identify infection-specific alterations in multiple long noncoding RNAs. Our study provides a paradigm for a sensitive RNA-based analysis of intracellular bacterial pathogens and their hosts without physical separation, as well as a new discovery route for hidden functions of pathogen genes.
    Keywords: Gene Expression Regulation -- Genetics ; Host-Pathogen Interactions -- Genetics ; RNA, Bacterial -- Genetics ; RNA, Untranslated -- Genetics ; Salmonella Typhimurium -- Genetics
    ISSN: 00280836
    E-ISSN: 1476-4687
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  • 5
    In: EMBO Journal, 13 November 2013, Vol.32(22), pp.2963-2979
    Description: Small RNAs use a diversity of well‐characterized mechanisms to repress mRNAs, but how they activate gene expression at the mRNA level remains not well understood. The predominant activation mechanism of Hfq‐associated small RNAs has been translational control whereby base pairing with the target prevents the formation of an intrinsic inhibitory structure in the mRNA and promotes translation initiation. Here, we report a translation‐independent mechanism whereby the small RNA RydC selectively activates the longer of two isoforms of mRNA (encoding cyclopropane fatty acid synthase) in . Target activation is achieved through seed pairing of the pseudoknot‐exposed, conserved 5′ end of RydC to an upstream region of the mRNA. The seed pairing stabilizes the messenger, likely by interfering directly with RNase E‐mediated decay in the 5′ untranslated region. Intriguingly, this mechanism is generic such that the activation is equally achieved by seed pairing of unrelated small RNAs, suggesting that this mechanism may be utilized in the design of RNA‐controlled synthetic circuits. Physiologically, RydC is the first small RNA known to regulate membrane stability. The small RNA RydC stabilizes target mRNAs in a translation‐independent manner through base pairing to the 5′UTR, blocking RNase E access. Cyclopropane fatty acid synthase is a target for RydC, providing the first link between sRNA regulation and membrane biosynthesis in bacteria.
    Keywords: Fatty Acid Synthesis ; Hfq ; Mrna Activation ; Noncoding Rna ; Small Rna
    ISSN: 0261-4189
    E-ISSN: 1460-2075
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  • 6
    Language: English
    In: Nature, November 2018, Vol.563(7729), pp.121-125
    Description: Many evolutionarily distant pathogenic organisms have evolved similar survival strategies to evade the immune responses of their hosts. These include antigenic variation, through which an infecting organism prevents clearance by periodically altering the identity of proteins that are visible to the immune system of the host. Antigenic variation requires large reservoirs of immunologically diverse antigen genes, which are often generated through homologous recombination, as well as mechanisms to ensure the expression of one or very few antigens at any given time. Both homologous recombination and gene expression are affected by three-dimensional genome architecture and local DNA accessibility. Factors that link three-dimensional genome architecture, local chromatin conformation and antigenic variation have, to our knowledge, not yet been identified in any organism. One of the major obstacles to studying the role of genome architecture in antigenic variation has been the highly repetitive nature and heterozygosity of antigen-gene arrays, which has precluded complete genome assembly in many pathogens. Here we report the de novo haplotype-specific assembly and scaffolding of the long antigen-gene arrays of the model protozoan parasite Trypanosoma brucei, using long-read sequencing technology and conserved features of chromosome folding. Genome-wide chromosome conformation capture (Hi-C) reveals a distinct partitioning of the genome, with antigen-encoding subtelomeric regions that are folded into distinct, highly compact compartments. In addition, we performed a range of analyses-Hi-C, fluorescence in situ hybridization, assays for transposase-accessible chromatin using sequencing and single-cell RNA sequencing-that showed that deletion of the histone variants H3.V and H4.V increases antigen-gene clustering, DNA accessibility across sites of antigen expression and switching of the expressed antigen isoform, via homologous recombination. Our analyses identify histone variants as a molecular link between global genome architecture, local chromatin conformation and antigenic variation.
    Keywords: Antigenic Variation -- Genetics ; Chromatin -- Genetics ; DNA, Protozoan -- Metabolism ; Genome -- Genetics ; Trypanosoma Brucei Brucei -- Genetics
    ISSN: 00280836
    E-ISSN: 1476-4687
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  • 7
    In: Nature, 2010, Vol.464(7286), p.250
    Description: Genome sequencing of Helicobacter pylori has revealed the potential proteins and genetic diversity of this prevalent human pathogen, yet little is known about its transcriptional organization and noncoding RNA output. Massively parallel cDNA sequencing (RNA-seq) has been revolutionizing global transcriptomic analysis. Here, using a novel differential approach (dRNA-seq) selective for the 5' end of primary transcripts, we present a genome-wide map of H. pylori transcriptional start sites and operons. We discovered hundreds of transcriptional start sites within operons, and opposite to annotated genes, indicating that complexity of gene expression from the small H. pylori genome is increased by uncoupling of polycistrons and by genome-wide antisense transcription. We also discovered an unexpected number of approximately 60 small RNAs including the epsilon-subdivision counterpart of the regulatory 6S RNA and associated RNA products, and potential regulators of cis- and trans-encoded target messenger RNAs. Our approach establishes a paradigm for mapping and annotating the primary transcriptomes of many living species.
    Keywords: Gene Expression Profiling ; Genome, Bacterial -- Genetics ; Helicobacter Infections -- Microbiology ; Helicobacter Pylori -- Genetics ; RNA, Bacterial -- Genetics;
    ISSN: 0028-0836
    E-ISSN: 14764687
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  • 8
    Language: English
    In: Journal of bacteriology, 15 September 2016, Vol.198(18), pp.2410-8
    Description: Bacteroides is a major component of the human gut microbiota which has a broad impact on the development and physiology of its host and a potential role in a wide range of disease syndromes. The predominance of this genus is due in large part to expansion of paralogous gene clusters, termed polysaccharide utilization loci (PULs), dedicated to the uptake and catabolism of host-derived and dietary polysaccharides. The nutritive value and availability of polysaccharides in the gut vary greatly; thus, their utilization is hierarchical and strictly controlled. A typical PUL includes regulatory genes that induce PUL expression in response to the presence of specific glycan substrates. However, the existence of additional regulatory mechanisms has been predicted to explain phenomena such as hierarchical control and catabolite repression. In this report, a previously unknown layer of regulatory control was discovered in Bacteroides fragilis Exploratory transcriptome sequencing (RNA-seq) analysis revealed the presence of cis-encoded antisense small RNAs (sRNAs) associated with 15 (30%) of the B. fragilis PULs. A model system using the Don (degradation of N-glycans) PUL showed that the donS sRNA negatively regulated Don expression at the transcriptional level, resulting in a decrease in N-glycan utilization. Additional studies performed with other Bacteroides species indicated that this regulatory mechanism is highly conserved and, interestingly, that the regulated PULs appear to be closely linked to the utilization of host-derived glycans rather than dietary plant polysaccharides. The findings described here demonstrate a global control mechanism underlying known PUL regulatory circuits and provide insight into regulation of Bacteroides physiology. The human gut is colonized by a dense microbiota which is essential to the health and normal development of the host. A key to gut homeostasis is the preservation of a stable, diverse microbiota. Bacteroides is a dominant genus in the gut, and the ability of Bacteroides species to efficiently compete for a wide range of glycan energy sources is a crucial advantage for colonization. Glycan utilization is mediated by a large number of polysaccharide utilization loci (PULs) which are regulated by substrate induction. In this report, a novel family of antisense sRNAs is described whose members repress gene expression in a distinct subset of PULs. This repression downregulates PUL expression in the presence of energy sources that are more readily utilized such as glucose, thereby allowing efficient glycan utilization.
    Keywords: Bacteroides -- Metabolism ; Gene Expression Regulation, Bacterial -- Physiology ; Polysaccharides -- Metabolism ; RNA, Bacterial -- Metabolism ; RNA, Small Interfering -- Metabolism
    ISSN: 00219193
    E-ISSN: 1098-5530
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  • 9
    In: PLoS ONE, 2015, Vol.10(11)
    Description: Bacillus amyloliquefaciens subsp. plantarum FZB42 is a representative of Gram-positive plant-growth-promoting rhizobacteria (PGPR) that inhabit plant root environments. In order to better understand the molecular mechanisms of bacteria-plant symbiosis, we have systematically analyzed the primary transcriptome of strain FZB42 grown under rhizosphere-mimicking conditions using differential RNA sequencing (dRNA-seq). Our analysis revealed 4,877 transcription start sites for protein-coding genes, identified genes differentially expressed under different growth conditions, and corrected many previously mis-annotated genes. We also identified a large number of riboswitches and cis- encoded antisense RNAs, as well as trans- encoded small noncoding RNAs that may play important roles in the gene regulation of Bacillus . Overall, our analyses provided a landscape of Bacillus primary transcriptome and improved the knowledge of rhizobacteria-host interactions.
    Keywords: Research Article
    E-ISSN: 1932-6203
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  • 10
    Language: English
    In: Proceedings of the National Academy of Sciences of the United States of America, 15 May 2012, Vol.109(20), pp.E1277-86
    Description: More than 50 y of research have provided great insight into the physiology, metabolism, and molecular biology of Salmonella enterica serovar Typhimurium (S. Typhimurium), but important gaps in our knowledge remain. It is clear that a precise choreography of gene expression is required for Salmonella infection, but basic genetic information such as the global locations of transcription start sites (TSSs) has been lacking. We combined three RNA-sequencing techniques and two sequencing platforms to generate a robust picture of transcription in S. Typhimurium. Differential RNA sequencing identified 1,873 TSSs on the chromosome of S. Typhimurium SL1344 and 13% of these TSSs initiated antisense transcripts. Unique findings include the TSSs of the virulence regulators phoP, slyA, and invF. Chromatin immunoprecipitation revealed that RNA polymerase was bound to 70% of the TSSs, and two-thirds of these TSSs were associated with σ(70) (including phoP, slyA, and invF) from which we identified the -10 and -35 motifs of σ(70)-dependent S. Typhimurium gene promoters. Overall, we corrected the location of important genes and discovered 18 times more promoters than identified previously. S. Typhimurium expresses 140 small regulatory RNAs (sRNAs) at early stationary phase, including 60 newly identified sRNAs. Almost half of the experimentally verified sRNAs were found to be unique to the Salmonella genus, and 〈20% were found throughout the Enterobacteriaceae. This description of the transcriptional map of SL1344 advances our understanding of S. Typhimurium, arguably the most important bacterial infection model.
    Keywords: Gene Expression Regulation, Bacterial -- Genetics ; RNA, Small Untranslated -- Genetics ; Regulatory Sequences, Ribonucleic Acid -- Genetics ; Salmonella Typhimurium -- Genetics ; Transcription, Genetic -- Genetics
    ISSN: 00278424
    E-ISSN: 1091-6490
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