A small RNA regulates the expression of genes involved in iron metabolism in
            <i>Escherichia</i>
            <i>coli</i>

Massé, Éric; Gottesman, Susan

doi:10.1073/pnas.032066599

Cited by 1,036 publications

(1,210 citation statements)

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“…induction of iro mRNAs as well as RyhB and IsrE sRNAs (Masse and Gottesman, 2002)) and magnesium limitation (mgtB) were observed ( Fig. 2.22a).…”

Section: Salmonella Encounters Various Stresses Inside Its Host Cell mentioning

confidence: 89%

“…Overall, intracellular sRNA expression levels matched well to the findings obtained from the pilot study (chapter 2). For example, an activation of the FURregulated sRNA RyhB (Masse and Gottesman, 2002;Padalon-Brauch et al, 2008) shows again how intracellular Salmonella encounter iron limitation and induction of OmrA/B indicates the presence of bacterial surface stress (Beisel and Storz, 2010). In addition, sRNAs known to be controlled by the master regulators of virulence followed the expected kinetics (see Fig.…”

Section: Stnc440 Expression Is Rapidly Induced After Invasion Of Varimentioning

confidence: 96%

“…2.24a,b). To confirm Dual RNA-seq-based gene expression profiling of intracellular Salmonella by unrelated methods, four of the induced sRNAs were selected for independent qRT-PCR experiments: The wellcharacterized Salmonella sRNAs RyhB (Masse and Gottesman, 2002) and RybB (Balbontin et al, 2010;Papenfort et al, 2006) were chosen as representative for a highly (>100 fold) or slightly (~2 fold) induced sRNA. In addition, STnc440 -a previously validated (Sittka et al, 2008) but poorly characterized sRNA -was included as was a computationally predicted candidate sRNA, STnc510.…”

Section: Srna Expression Profile Of Intracellular Salmonellamentioning

confidence: 99%

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Dual RNA-seq of pathogen and host

2012

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SummaryThe infection of a eukaryotic host cell by a bacterial pathogen is one of the most intimate examples of cross-kingdom interactions in biology. Infection processes are highly relevant from both a basic research as well as a clinical point of view. Sophisticated mechanisms have evolved in the pathogen to manipulate the host response and vice versa host cells have developed a wide range of anti-microbial defense strategies to combat bacterial invasion and clear infections.However, it is this diversity and complexity that makes infection research so challenging to technically address as common approaches have either been optimized for bacterial or eukaryotic organisms. Instead, methods are required that are able to deal with the often dramatic discrepancy between host and pathogen with respect to various cellular properties and processes. One class of cellular macromolecules that exemplify this host-pathogen heterogeneity is given by their transcriptomes: Bacterial transcripts differ from their eukaryotic counterparts in many aspects that involve both quantitative and qualitative traits. The entity of RNA transcripts present in a cell is of paramount interest as it reflects the cell's physiological state under the given condition. Genome-wide transcriptomic techniques such as RNA-seq have therefore been used for single-organism analyses for several years, but their applicability has been limited for infection studies.The present work describes the establishment of a novel transcriptomic approach for infection biology which we have termed "Dual RNA-seq". Using this technology, it was intended to shed light particularly on the contribution of non-protein-encoding transcripts to virulence, as these classes have mostly evaded previous infection studies due to the lack of suitable methods. The performance of Dual RNA-seq was evaluated in an in vitro infection model based on the important facultative intracellular pathogen Salmonella enterica serovar Typhimurium and different human cell lines. Dual RNA-seq was found to be capable of capturing all major bacterial and human transcript classes and proved reproducible. During the course of these experiments, a previously largely uncharacterized bacterial small non-coding RNA (sRNA), referred to as STnc440, was identified as one of the most strongly induced genes in intracellular Salmonella.Interestingly, while inhibition of STnc440 expression has been previously shown to cause a virulence defect in different animal models of Salmonellosis, the underlying molecular mechanisms have remained obscure. Here, classical genetics, transcriptomics and biochemical assays proposed a complex model of Salmonella gene expression control that is orchestrated by this sRNA. In particular, STnc440 was found to be involved in the regulation of multiple bacterial target mRNAs by direct base pair interaction with consequences for Salmonella virulence and

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“…induction of iro mRNAs as well as RyhB and IsrE sRNAs (Masse and Gottesman, 2002)) and magnesium limitation (mgtB) were observed ( Fig. 2.22a).…”

Section: Salmonella Encounters Various Stresses Inside Its Host Cell mentioning

confidence: 89%

Section: Stnc440 Expression Is Rapidly Induced After Invasion Of Varimentioning

confidence: 96%

Section: Srna Expression Profile Of Intracellular Salmonellamentioning

confidence: 99%

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Dual RNA-seq of pathogen and host

2012

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“…Escherichia coli Hfq mutants show disrupted signaling in stress response pathways 7,8 , arising from the need for Hfq to mediate base-pairing between regulatory ncRNAs and their mRNA targets. Examples of these partnerships include DsrA-rpoS 7,9,10 , OxyS-fhlA 11,12 , OxyS-rpoS 13 , RprA-rpoS 14 , RyhBsodB [15][16][17] .…”

Section: Introductionmentioning

confidence: 99%

Escherichia coli Hfq has distinct interaction surfaces for DsrA, rpoS and poly(A) RNAs

Mikulecky

Kaw

Brescia

et al. 2004

Nat Struct Mol Biol

226

395

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The bacterial Sm-like protein Hfq facilitates RNA-RNA interactions involved in posttranscriptional regulation of the stress response. Specifically, Hfq helps pair noncoding RNAs (ncRNAs) with complementary regions of target mRNAs. To probe the mechanism of this pairing, we generated a series of Hfq mutants and measured their affinity for RNAs like those with which Hfq must associate in vivo. We tested the mutants' DsrA-dependent activation of rpoS, and their ability to stabilize DsrA ncRNA against degradation in vivo. Our results suggest that Hfq has two independent RNA-binding surfaces. In addition to a well-known site around the core of the Hfq hexamer, we observe interactions with the distal face of Hfq, a new locus with which mRNAs and poly(A) sequences associate. Our model explains how Hfq can simultaneously bind a ncRNA and its mRNA target to facilitate the strand displacement reaction required for Hfq-dependent translational regulation.Hfq protein from Escherichia coli was first described in connection with Qβ-phage replication 1,2 . Hfq has recently emerged as a central player in post-transcriptional gene regulation as mediated by bacterial ncRNAs [3][4][5][6] . Escherichia coli Hfq mutants show disrupted signaling in stress response pathways 7,8 , arising from the need for Hfq to mediate base-pairing between regulatory ncRNAs and their mRNA targets. Examples of these partnerships include DsrA-rpoS 7,9,10 , OxyS-fhlA 11,12 , OxyS-rpoS 13 , RprA-rpoS 14 , RyhBsodB [15][16][17] .Complexes between ncRNAs and their mRNA targets function in several ways. Most commonly, complexed structures lead to translational activation or repression by remodeling mRNA regulatory regions containing the ribosome-binding site (RBS) and/or start codon. Alternatively, the interaction can enhance decay of the target mRNA16 or simply block translation11. Clearly, Hfq facilitates base-pairing between ncRNAs and their targets, but how it does so is poorly understood. How the chaperone function relates to other Hfq activities such as the control of poly(A) tail elongation19 , 20 and regulation of mRNA stability21 , 22 is also unknown. COMPETING INTERESTS STATEMENTThe authors declare that they have no competing financial interests. NIH Public Access Author ManuscriptNat Struct Mol Biol. Author manuscript; available in PMC 2011 April 5. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptHfq shares sequence similarity to the eukaryotic Lsm proteins [23][24][25][26][27] We addressed these questions through a mutational analysis of Hfq, probing in vitro binding to several model RNAs that represent species with which Hfq must interact. Hfq mutants were assayed in vivo using a reporter assay and RNA lifetime experiments. Together, the results support a model wherein at least two independent RNA-binding sites exist on the Hfq hexamer, and juxtaposition of bound RNAs facilitates base-pairing. RESULTS Hfq mutagenesisTo identify amino acids essential for RNA binding, we constructed a series of E. coli Hfq misse...

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“…RyhB is a small regulatory RNA that modulates multiple mRNAs involved in iron metabolism in Escherichia coli (Massé and Gottesman 2002). Fe is an important nutrient involved in several cellular processes, including respiration, the citric acid cycle, DNA synthesis, elimination of H 2 O 2 and O 2 − , photosynthesis and others (for review, see Andrews 1998).…”

mentioning

confidence: 99%

Coupled degradation of a small regulatory RNA and its mRNA targets in Escherichia coli

Massé¹,

Escorcia²,

Gottesman³

2003

Genes Dev.

Self Cite

656

836

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RyhB is a small antisense regulatory RNA that is repressed by the Fur repressor and negatively regulates at least six mRNAs encoding Fe-binding or Fe-storage proteins in Escherichia coli. When Fe is limiting, RyhB levels rise, and target mRNAs are rapidly degraded. RyhB is very stable when measured after treatment of cells with the transcription inhibitor rifampicin, but is unstable when overall mRNA transcription continues. We propose that RyhB turnover is coupled to and dependent on pairing with the target mRNAs. Degradation of both mRNA targets and RyhB is dependent on RNase E and is slowed in degradosome mutants. RyhB requires the RNA chaperone Hfq. In the absence of Hfq, RyhB is unstable, even when general transcription is inhibited; degradation is dependent upon RNase E. Hfq and RNase E bind similar sites on the RNA; pairing may allow loss of Hfq and access by RNase E. Two other Hfq-dependent small RNAs, DsrA and OxyS, are also stable when overall transcription is off, and unstable when it is not, suggesting that they, too, are degraded when their target mRNAs are available for pairing. Thus, this large class of regulatory RNAs share an unexpected intrinsic mechanism for shutting off their action.

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A small RNA regulates the expression of genes involved in iron metabolism in Escherichia coli

Cited by 1,036 publications

References 40 publications

Dual RNA-seq of pathogen and host

Dual RNA-seq of pathogen and host

Escherichia coli Hfq has distinct interaction surfaces for DsrA, rpoS and poly(A) RNAs

Coupled degradation of a small regulatory RNA and its mRNA targets in Escherichia coli

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