A conserved RNA seed‐pairing domain directs small RNA‐mediated stress resistance in enterobacteria

Peschek, Nikolai; Hoyos, Mona; Herzog, Rudolf; Förstner, Konrad U.; Papenfort, Kai

doi:10.15252/embj.2019101650

Cited by 26 publications

(38 citation statements)

References 46 publications

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“…1D). At low cell densities, the top five most abundant sRNAs were the yet-uncharacterized Vcr090 sRNA (23), the highly conserved Spot 42 (38), MicV (26), FarS [previously identified as Vcr076 (23)], and VqmR (23). At high cell densities, the relative levels of Vcr090, MicV, and Spot 42 decreased, while VqmR became the most abundant sRNA, followed by FarS.…”

Section: Resultsmentioning

confidence: 99%

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Switching fatty acid metabolism by an RNA-controlled feed forward loop

Huber

Fröhlich

Radmer

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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Hfq (host factor for phage Q beta) is key for posttranscriptional gene regulation in many bacteria. Hfq’s function is to stabilize sRNAs and to facilitate base-pairing withtrans-encoded target mRNAs. Loss of Hfq typically results in pleiotropic phenotypes, and, in the major human pathogenVibrio cholerae, Hfq inactivation has been linked to reduced virulence, failure to produce biofilms, and impaired intercellular communication. However, the RNA ligands of Hfq inV. choleraeare currently unknown. Here, we used RIP-seq (RNA immunoprecipitation followed by high-throughput sequencing) analysis to identify Hfq-bound RNAs inV. cholerae. Our work revealed 603 coding and 85 noncoding transcripts associated with Hfq, including 44 sRNAs originating from the 3′ end of mRNAs. Detailed investigation of one of these latter transcripts, named FarS (fatty acid regulated sRNA), showed that this sRNA is produced by RNase E-mediated maturation of thefabB3′UTR, and, together with Hfq, inhibits the expression of two paralogousfadEmRNAs. ThefabBandfadEgenes are antagonistically regulated by the major fatty acid transcription factor, FadR, and we show that, together, FadR, FarS, and FadE constitute a mixed feed-forward loop regulating the transition between fatty acid biosynthesis and degradation inV. cholerae. Our results provide the molecular basis for studies on Hfq inV. choleraeand highlight the importance of a previously unrecognized sRNA for fatty acid metabolism in this major human pathogen.

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Section: Resultsmentioning

confidence: 99%

“…In addition, V. cholerae sRNAs controlling cellcell communication, e.g. Qrr1-4 (21) and VqmR (22)(23)(24), as well as sRNAs responding to cell-envelope damage (25,26), contribute to virulence gene expression.…”

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confidence: 99%

Switching fatty acid metabolism by an RNA-controlled feed forward loop

Huber

Fröhlich

Radmer

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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“…VrrA also represses the major OMP OmpT, the biofilm matrix protein RbmC and the ribosome-binding protein Vrp, involved in starvation survival [ 118 , 119 , 120 ]. The second sRNA, MicV, shares the same seed-pairing domain as VrrA and more than 23 genes are targeted by both sRNAs including the genes encoding OMP OmpA and OmpT, suggesting a redundant function of VrrA and MicV [ 121 ].…”

Section: Srnas Regulating Host-pathogen Interactionsmentioning

confidence: 99%

Interplay between Regulatory RNAs and Signal Transduction Systems during Bacterial Infection

Piattelli

Peltier

Soutourina

2020

Genes

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The ability of pathogenic bacteria to stably infect the host depends on their capacity to respond and adapt to the host environment and on the efficiency of their defensive mechanisms. Bacterial envelope provides a physical barrier protecting against environmental threats. It also constitutes an important sensory interface where numerous sensing systems are located. Signal transduction systems include Two-Component Systems (TCSs) and alternative sigma factors. These systems are able to sense and respond to the ever-changing environment inside the host, altering the bacterial transcriptome to mitigate the impact of the stress. The regulatory networks associated with signal transduction systems comprise small regulatory RNAs (sRNAs) that can be directly involved in the expression of virulence factors. The aim of this review is to describe the importance of TCS- and alternative sigma factor-associated sRNAs in human pathogens during infection. The currently available genome-wide approaches for studies of TCS-regulated sRNAs will be discussed. The differences in the signal transduction mediated by TCSs between bacteria and higher eukaryotes and the specificity of regulatory RNAs for their targets make them appealing targets for discovery of new strategies to fight against multi-resistant bacteria.

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“…Shaker-mill homogenization as a form of mechanical lysis has been proven time and again as a successful method for disrupting tissues, microorganisms, and biologic samples for downstream molecular analysis; however, these downstream applications often require the use of additional puri cation, isolation, or extraction procedures before those analyses can be completed [9,10,11,12].…”

Section: Introductionmentioning

confidence: 99%

“…While, shaker-mill homogenization procedures have not yet been applied directly to diagnostic technologies, its ability to lyse organisms and tissues far tougher than a virus have been well categorized [9,10,11,12]. Building off the increased need for novel viral diagnostic methodologies, which reduce the resources and time required for accurate viral pathogen detection, we felt it was time to examine the capabilities of shaker-mill homogenization for viral lysis to be used for downstream detection.…”

Section: Introductionmentioning

confidence: 99%

A Novel Two-Step, Direct-to-PCR Method for Virus Detection off Swabs using Human Coronavirus 229E

Morehouse

Proctor

Ryan

et al. 2020

Preprint

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Abstract Background Currently, one of the most reliable methods for viral infection detection are polymerase chain reaction (PCR) based assays. This process is time and resource heavy, requiring multiple steps of lysis, extraction, purification, and amplification procedures. Herein, we have developed a method to detect virus off swabs using solely shaker-mill based mechanical lysis and the transfer of the viral lysate directly to a PCR assay for virus detection, bypassing the substantial reagent and time investments required for extraction and purification steps.Methods Using Human Coronavirus 229E (HCoV-229E) as a model system, we spiked swabs in vitro for proof-of-concept testing. Swabs were spiked in serial dilutions from 1.2x106 to 1.2x101 copies/mL and then placed in 2mL tubes with viral transport media (VTM) to mimic the specimen collection procedures in the clinic prior to processing via shaker-mill homogenization. After homogenization, 1 µL of lysate was processed using RT-qPCR for amplification of the nucleocapsid (N) gene, qualifying viral detection. Results HCoV-229E in vitro spiked swabs were processed in a novel two-step, direct-to-PCR methodology for viral detection. After running 54 swabs, we confidently determined our limit of detection to be 1.2x103 viral copies/mL with 96.30% sensitivity. Conclusion We have proven that the shaker-mill homogenization-based two-step, direct-to-PCR procedures provides sufficient viral lysis off swabs, where the resulting lysate can be used directly in PCR for the detection of HCoV-229E. This finding allows for reductions in the time and resources required for PCR based virus detection in comparison to the traditional extraction-to-PCR methodology.

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A conserved RNA seed‐pairing domain directs small RNA‐mediated stress resistance in enterobacteria

Cited by 26 publications

References 46 publications

Switching fatty acid metabolism by an RNA-controlled feed forward loop

Switching fatty acid metabolism by an RNA-controlled feed forward loop

Interplay between Regulatory RNAs and Signal Transduction Systems during Bacterial Infection

A Novel Two-Step, Direct-to-PCR Method for Virus Detection off Swabs using Human Coronavirus 229E

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