| 1. |
- Darfeuille, Fabien, et al.
(författare)
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An antisense RNA inhibits translation by competing with standby ribosomes
- 2007
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Ingår i: Molecular Cell. - : Elsevier BV. - 1097-2765 .- 1097-4164. ; 26:3, s. 381-392
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Tidskriftsartikel (refereegranskat)abstract
- Most antisense RNAs in bacteria inhibit translation by competing with ribosomes for translation initiation regions (TIRs) on nascent mRNA. We propose a mechanism by which an antisense RNA inhibits translation without binding directly to a TIR. The tisAB locus encodes an SOS-induced toxin, and IstR-1 is the antisense RNA that counteracts toxicity. We show that full-length tisAB mRNA (+1) is translationally inactive and endonucleolytic processing produces an active mRNA (+42). IstR-1 binding inhibits translation of this mRNA, and subsequent RNase III cleavage generates a truncated, inactive mRNA (+106). In vitro translation, toeprinting, and structure mapping suggest that active, but not inactive, tisAB mRNAs contain an upstream ribosome loading or “standby” site. Standby binding is required for initiation at the highly structured tisB TIR. This may involve ribosome sliding to a transiently open tisB TIR. IstR-1 competes with ribosomes by base pairing to the standby site located 100 nucleotides upstream.
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| 2. |
- Rizvanovic, Alisa
(författare)
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Adapting to succeed : Post-transcriptional gene regulation in Salmonella
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Salmonella are zoonotic pathogens of worldwide economic and health importance. Both during life outside and inside the host, these pathogens are subject to continuously changing environmental conditions, such as temperature changes, acid stress, nutrient limitations, and others. In order to thrive and survive, Salmonella must respond to these changes by adapting their physiology and morphology through changes in gene expression. RNA-binding proteins (RBPs) often work in concert with small RNAs (sRNAs) to control gene expression at the post-transcriptional level. Their mode of action includes regulation of RNA translation and/or stability, either positive or negative. Recently, ProQ was discovered to be a global RBP with a large repertoire of mRNA and sRNA targets in Salmonella. However, many details regarding ProQ biology are not fully understood, including the requirements for RNA-binding, mechanisms of gene regulation, and ProQ-dependent phenotypic changes. The main purpose of this doctoral thesis was to characterize the RBP ProQ and its regulatory role in Salmonella.First, we developed a method based on saturation mutagenesis coupled to phenotypic sorting and high-throughput sequencing to chart the functionally important regions in ProQ. Our results reveal that both the N-terminal and C-terminal domains are important for ProQ’s gene regulatory function, but the underlying mechanisms differ. Second, we show that ProQ is important for flagellar-mediated motility in Salmonella. More specifically, we show that ProQ and an associated sRNA promotes flagellar gene expression and motility by affecting translation of the master flagellar regulator FlhDC. Finally, we reveal that ProQ induces persister formation in Salmonella and enables a subpopulation of cells to survive high doses of different types of antibiotics through growth arrest.In conclusion, the findings presented herein provide new insights into the role of ProQ as a global post-transcriptional regulator of gene expression in Salmonella. Together, these findings contribute to our understanding of how Salmonella shapes its lifestyle and induces pathogenesis.
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| 3. |
- Udekwu, Klas I, et al.
(författare)
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Hfq-dependent regulation of OmpA synthesis is mediated by an an-tisense RNA.
- 2005
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Ingår i: Genes & Development. - : Cold Spring Harbor Laboratory. - 0890-9369 .- 1549-5477. ; 19:19, s. 2355-2366
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Tidskriftsartikel (refereegranskat)abstract
- This paper shows that the small RNA MicA (previously SraD) is an antisense regulator of ompA in Escherichia coli. MicA accumulates upon entry into stationary phase and down-regulates the level of ompA mRNA. Regulation of ompA (outer membrane protein A), previously attributed to Hfq/mRNA binding, is lost upon deletion of the micA gene, whereas overexpression of MicA inhibits the synthesis of OmpA. In vitro, MicA binds to the ompA mRNA leader. Enzymatic and chemical probing was used to map the structures of MicA, the ompA mRNA leader, and the complex formed upon binding. MicA binding generates a footprint across the ompA Shine-Dalgarno sequence, consistent with a 12 + 4 base-pair interaction, which is additionally supported by the effect of mutations in vivo and by bioinformatics analysis of enterobacterial micA/ompA homolog sequences. MicA is conserved in many enterobacteria, as is its ompA target site. In vitro toeprinting confirmed that binding of MicA specifically interferes with ribosome binding. We propose that MicA, when present at high levels, blocks ribosome binding at the ompA translation start site, which—in line with previous work—secondarily facilitates RNase E cleavage and subsequent mRNA decay. MicA requires the presence of the Hfq protein, although the mechanistic basis for this remains unclear.
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| 4. |
- Vo, Duc Duy, et al.
(författare)
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Building of neomycin-nucleobase-amino acid conjugates for the inhibition of oncogenic miRNAs biogenesis
- 2018
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Ingår i: Organic and biomolecular chemistry. - : Royal Society of Chemistry (RSC). - 1477-0520 .- 1477-0539. ; 16:34, s. 6262-6274
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Tidskriftsartikel (refereegranskat)abstract
- MicroRNAs (miRNAs) are a recently discovered category of small RNA molecules that regulate gene expression at the post-transcriptional level. Accumulating evidence indicates that miRNAs are aberrantly expressed in a variety of human cancers, thus being oncogenic. The inhibition of oncogenic miRNAs (defined as the blocking of miRNAs' production or function) would find application in the therapy of different types of cancer in which these miRNAs are implicated. In this work, we describe the design and synthesis of new small-molecule RNA ligands with the aim of inhibiting Dicer-mediated processing of oncogenic miRNAs. One of the synthesized compound (4b) composed of the aminoglycoside neomycin conjugated to an artificial nucleobase and to amino acid histidine is able to selectively decrease miR-372 levels in gastric adenocarcinoma (AGS) cells and to restore the expression of the target LATS2 protein. This activity led to the inhibition of proliferation of these cells. The study of the interactions of 4b with pre-miR-372 allowed for the elucidation of the molecular mechanism of the conjugate, thus leading to new perspectives for the design of future inhibitors.
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| 5. |
- Wagner, Gerhart E. H., et al.
(författare)
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Small regulatory RNAs in bacteria
- 2006
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Ingår i: Small RNAs. - Berlin, Heidelberg : Springer Verlag Berlin Heidelberg. - 9783540281306 ; , s. 1-29
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Bokkapitel (övrigt vetenskapligt/konstnärligt)abstract
- In recent years, small regulatory RNAs have been discovered at a staggering rate both in prokaryotes and eukaryotes. By now it is clear that post-transcriptional regulation of gene expression mediated by such RNAs is the rule rather than—as previously believed—the exception. In this chapter, we focus on small RNAs (sRNAs) encoded by bacterial chromosomes. The strategies for their discovery, their biological roles, and their mechanisms of action are discussed. Even though the number of well-characterized sRNAs in, for example, the best studied model enterobacterium Escherichia coli, is still small, the emerging pattern suggests that antisense mechanisms predominate. In terms of their roles in bacterial physiology, most of these RNAs appear to be involved in stress response regulation. Some other examples indicate functions in regulation of virulence. Two aspects of sRNA-mediated control arising from recent observations are addressed as well. Firstly, some sRNAs need proteins (notably Hfq) as helpers in their antisense activities—at this point the reason for this requirement is not understood. Secondly, only limited sequence complementarity is generally observed in antisense–target RNA pairs. This raises the fundamental question of how specific recognition is accomplished, and what the structure/sequence determinants for rapid and productive interaction are.
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