| 1. |
- Felletti, Michele, et al.
(författare)
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A nascent polypeptide sequence modulates DnaA translation elongation in response to nutrient availability
- 2021
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Ingår i: eLIFE. - : eLife Sciences Publications Ltd. - 2050-084X. ; 10
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Tidskriftsartikel (refereegranskat)abstract
- The ability to regulate DNA replication initiation in response to changing nutrient conditions is an important feature of most cell types. In bacteria, DNA replication is triggered by the initiator protein DnaA, which has long been suggested to respond to nutritional changes; nevertheless, the underlying mechanisms remain poorly understood. Here, we report a novel mechanism that adjusts DnaA synthesis in response to nutrient availability in Caulobacter crescentus. By performing a detailed biochemical and genetic analysis of the dnaA mRNA, we identified a sequence downstream of the dnaA start codon that inhibits DnaA translation elongation upon carbon exhaustion. Our data show that the corresponding peptide sequence, but not the mRNA secondary structure or the codon choice, is critical for this response, suggesting that specific amino acids in the growing DnaA nascent chain tune translational efficiency. Our study provides new insights into DnaA regulation and highlights the importance of translation elongation as a regulatory target. We propose that translation regulation by nascent chain sequences, like the one described, might constitute a general strategy for modulating the synthesis rate of specific proteins under changing conditions.
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| 2. |
- Rizvanovic, Alisa, et al.
(författare)
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The RNA-binding protein ProQ promotes antibiotic persistence in Salmonella
- 2022
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Ingår i: Molecular and Cellular Biology. - : American Society for Microbiology. - 0270-7306 .- 1098-5549. ; 13:6
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Tidskriftsartikel (refereegranskat)abstract
- Bacterial populations can survive the exposure to antibiotics through transient phenotypic and gene expression changes. These changes can be attributed to a small subpopulation of bacteria, giving rise to antibiotic persistence. Even though this phenomenon has been known for decades, much is still to be learnt about the mechanisms that drive persister formation. The RNA-binding protein ProQ has recently emerged as a global regulator of gene expression. Here, we show that ProQ impacts persister formation in Salmonella. ProQ contributes to growth-arrest in single cells, which are able to survive treatment with high concentrations of different antibiotics. The underlying mechanism for ProQ-dependent persister formation involves activation of the flagellar pathway. Importantly, we show that the ProQ-dependent phenotype is relevant during macrophage infection and allows Salmonella to survive the combined action of host immune defences and antibiotics. Together, our data highlights the importance of ProQ in Salmonella persistence and pathogenesis.
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| 3. |
- Romilly, Cedric, et al.
(författare)
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An RNA pseudoknot is essential for standby-mediated translation of the tisB toxin mRNA in Escherichia coli
- 2020
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Ingår i: Nucleic Acids Research. - : OXFORD UNIV PRESS. - 0305-1048 .- 1362-4962. ; 48:21, s. 12336-12347
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Tidskriftsartikel (refereegranskat)abstract
- In response to DNA damage, Escherichia coli cells activate the expression of the toxin gene tisB of the toxin-antitoxin system tisB-istR1. Of three isoforms, only the processed, highly structured +42 tisB mRNA is active. Translation requires a standby site, composed of two essential elements: a single-stranded region located 100 nucleotides upstream of the sequestered RBS, and a structure near the 5'-end of the active mRNA. Here, we propose that this 5'-structure is an RNA pseudoknot which is required for 30S and protein S1-alone binding to the mRNA. Point mutations that prevent formation of this pseudoknot inhibit formation of translation initiation complexes, impair S1 and 30S binding to the mRNA, and render the tisB mRNA non-toxic in vivo. A set of mutations created in either the left or right arm of stem 2 of the pseudoknot entailed loss of toxicity upon overexpression of the corresponding mRNA variants. Combining the matching right-left arm mutations entirely restored toxicity levels to that of the wild-type, active mRNA. Finally, since many pseudoknots have high affinity for S1, we predicted similar pseudoknots in non-homologous type I toxin-antitoxin systems that exhibit features similar to that of tisB-IstR1, suggesting a shared requirement for standby acting at great distances.
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| 4. |
- Romilly, Cedric, et al.
(författare)
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Small RNAs OmrA and OmrB promote class III flagellar gene expression by inhibiting the synthesis of anti-Sigma factor FlgM
- 2020
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Ingår i: RNA Biology. - : Informa UK Limited. - 1547-6286 .- 1555-8584. ; 17:6, s. 872-880
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Tidskriftsartikel (refereegranskat)abstract
- Bacteria can move by a variety of mechanisms, the best understood being flagella-mediated motility. Flagellar genes are organized in a three-tiered cascade allowing for temporally regulated expression that involves both transcriptional and post-transcriptional control. The class I operon encodes the master regulator FlhDC that drives class II gene transcription. Class II genes include fliA and flgM, which encode the Sigma factor sigma(28), required for class III transcription, and the anti-Sigma factor FlgM, which inhibits sigma(28) activity, respectively. The flhDC mRNA is regulated by several small regulatory RNAs (sRNAs). Two of these, the sequence-related OmrA and OmrB RNAs, inhibit FlhD synthesis. Here, we report on a second layer of sRNA-mediated control downstream of FhlDC in the flagella pathway. By mutational analysis, we confirm that a predicted interaction between the conserved 5MODIFIER LETTER PRIME seed sequences of OmrA/B and the early coding sequence in flgM mRNA reduces FlgM expression. Regulation is dependent on the global RNA-binding protein Hfq. In vitro experiments support a canonical mechanism: binding of OmrA/B prevents ribosome loading and decreases FlgM protein synthesis. Simultaneous inhibition of both FlhD and FlgM synthesis by OmrA/B complicated an assessment of how regulation of FlgM alone impacts class III gene transcription. Using a combinatorial mutation strategy, we were able to uncouple these two targets and demonstrate that OmrA/B-dependent inhibition of FlgM synthesis liberates sigma(28) to ultimately promote higher expression of the class III flagellin gene fliC.
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| 5. |
- Wagner, E. Gerhart H., et al.
(författare)
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The Length of a DNA T-Tract Modulates Expression of a Virulence-Regulating sRNA
- 2020
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Ingår i: Molecular Cell. - : Elsevier BV. - 1097-2765 .- 1097-4164. ; 80:2, s. 175-177
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Eisenbart et al. (2020) find an SSR-associated sRNA, NikS, that is subject to variable repeat-controlled expression. NikS regulates H. pylori virulence by post-transcriptionally repressing pathogenicity factors, including CagA and VacA, via base-pairing to their mRNAs.
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