Small RNAs and RNA chaperone Hfq in enterohemorrhagic E. coli
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ABSTRACT:
PROVIDER: PRJNA197285 | ENA |
REPOSITORIES: ENA
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Project description:Bacterial small regulatory RNAs (sRNAs) regulate gene expression by base-pairing to their target mRNAs. In Escherichia coli and many other bacteria, this process is dependent on the RNA chaperone Hfq, which binds sRNAs and mRNAs on different faces. YhbS (renamed here as HqbA), a putative Gcn5-related N-acetyltransferase (GNAT), was identified as a novel silencer of sRNA signaling in a genomic library screen. Here, we studied how HqbA regulates sRNA signaling and determined its physiological roles in modulating Hfq activity. Using fluorescent reporter assays, we found that HqbA overproduction suppresses all tested Hfq-dependent sRNA signaling. Chromosomal HqbA suppresses the signaling of the ChiX sRNA when the C-terminus of Hfq was deleted. Direct interaction between HqbA and Hfq was demonstrated both in vivo and in vitro, and mutants that blocked interaction also interfered with HqbA suppression of Hfq. However, an acetylation-deficient HqbA mutant still disrupted sRNA signaling, suggesting that HqbA is bifunctional, with separate roles for regulating via Hfq interaction and via acetylation of undefined substrates. Gel shift assays indicated that HqbA strongly reduced the interaction between the Hfq distal face and low-affinity RNAs, but not high-affinity RNAs. Hfq-IP RNA-Seq in WT and chromosomal hqbA mutants led to the identification of two tRNA precursors, metZWV and proM, that were enriched in Hfq binding in the absence of HqbA interaction. Our results suggest that HqbA provides a level of quality control for Hfq by competing with low-affinity RNA binders.
2023-11-13 | GSE235194 | GEO
Project description:The RNA chaperone Hfq is regarded as a critical effector promoting interaction between small regulatory RNAs (sRNAs) and cognate target mRNAs. While much of this interpretation is based on in vitro assays, no in vivo evidence actually exists to support this model. Here we report that Hfq is typically unnecessary for binding of various sRNA-mRNA complexes in vivo. Data obtained from pull-downs of MS2-tagged RyhB, RybB and DsrA sRNAs followed by RNAseq identification of target mRNAs suggest that absence of Hfq is not essential for sRNA-mRNA complex formation in vivo.
2021-09-22 | GSE113587 | GEO
Project description:The RNA chaperone Hfq is regarded as a critical effector promoting interaction between small regulatory RNAs (sRNAs) and cognate target mRNAs. While much of this interpretation is based on in vitro assays, no in vivo evidence actually exists to support this model. Here we report that Hfq is typically unnecessary for binding of various sRNA-mRNA complexes in vivo. Data obtained from pull-downs of MS2-tagged RyhB, RybB and DsrA sRNAs followed by RNAseq identification of target mRNAs suggest that absence of Hfq is not essential for sRNA-mRNA complex formation in vivo.
2021-09-22 | GSE113591 | GEO
Project description:The RNA chaperone Hfq is regarded as a critical effector promoting interaction between small regulatory RNAs (sRNAs) and cognate target mRNAs. While much of this interpretation is based on in vitro assays, no in vivo evidence actually exists to support this model. Here we report that Hfq is typically unnecessary for binding of various sRNA-mRNA complexes in vivo. Data obtained from pull-downs of MS2-tagged RyhB, RybB and DsrA sRNAs followed by RNAseq identification of target mRNAs suggest that absence of Hfq is not essential for sRNA-mRNA complex formation in vivo.
2021-09-22 | GSE113589 | GEO
Project description:Hfq, a bacterial RNA chaperone, stabilizes small regulatory RNAs (sRNAs) and facilitates sRNA base-pairing with target mRNAs. Hfq has a conserved N-terminal domain and a poorly conserved disordered C-terminal domain (CTD). In a transcriptome-wide examination of the effects of a chromosomal CTD deletion (Hfq1-65), the Escherichia coli mutant was most defective for the accumulation of sRNAs that bind the proximal and distal faces of Hfq (Class II sRNAs), but other sRNAs also were affected. There were only modest effects on the levels of mRNAs, suggesting little disruption of sRNA-dependent regulation. However, cells expressing Hfq lacking the CTD deletion in combination with a weak distal face mutation were defective for the function of the Class II sRNA ChiX and repression of mutS, both dependent upon distal face RNA binding. Loss of the region between amino acids 66-72 was critical for this defect. The CTD region beyond amino acid 72 was not necessary for distal face-dependent regulation, but was needed for functions associated with the Hfq rim, seen most clearly in combination with a rim mutant. Our results suggest that the C-terminus collaborates in various ways with different binding faces of Hfq, leading to distinct outcomes for individual sRNAs.
2021-09-17 | GSE139988 | GEO
Project description:In many bacteria, the base pairing between most small regulatory RNAs (sRNAs) and their targets is facilitated by the Hfq RNA chaperone. However, recent studies have shown FinO-domain proteins also bind sRNAs. To compare the contributions of Hfq and the FinO-domain ProQ protein in Escherichia coli, we carried out RIL-seq, which allows global identification of two RNAs bound to the same protein. We detected hundreds of RNA pairs on ProQ. Intriguingly, 33% of the ProQ-bound RNA pairs are also found associated with Hfq, suggesting overlapping, complementary or competing roles for the two proteins.
2019-12-08 | GSE131520 | GEO
Project description:cDNA microarray analysis to identify genes regulated by the RNA chaperone, Hfq. Four experiments were performed: 1/ Hfq+ vs Hfq- strains. 269 significantly differentially regulated genes were identified by SAM (Statistical Analysis of Microarrays), of which 120 changed more than 1.5 fold (48 increased and 72 decreased in hfq-). Amongst other genes, these experiments identified significant regulation of the sigma E and sigma 32 regulons. However, only genes induced by sigma E were similarly induced in hfq-; 8 operons repressed by sigma E were not repressed in hfq-. 2/ wt vs delta rseA. RseA is the antisigma factor for sigmaE. This comparison results in elevated steady-state levels of sigma E, and confirmed induction and repression of target regulon members. 3/ hfq+ vs hfq+ rpoE overexpression. RpoE encoding sigma E was overexpressed in an hfq+ background, confirming normal regulation of the sigma E regulon. 4/ hfq+ vs hfq- rpoE overexpression. Sigma E was overexpressed in an hfq- background. This demonstrated that 8 operons normally repressed by sigma E require hfq for this repression. The simple conclusion is that sigma E regulates small RNAs that, together with Hfq, bind target mRNAs and results in their rapid degradation. This study is detailed in Guisbert et al 2007 (J Bacteriol, 189:1963-73) Keywords: Genetic modification
2007-01-01 | GSE6444 | GEO
Project description:We report the identification of new noncoding RNAs in Brucella suis 1330 and that are associated to the chaperone protein Hfq Coimmunoprecipitation using Flag-tagged Hfq as bait
2015-11-25 | E-GEOD-73621 | biostudies-arrayexpress
Project description:We report the identification of new noncoding RNAs in Brucella suis 1330 and that are associated to the chaperone protein Hfq
2015-11-25 | GSE73621 | GEO
Project description:The alpha-proteobacterium Caulobacter crescentus thrives in oligotrophic environments and is able to optimally exploit minimal resources by entertaining an intricate network of gene expression control mechanisms. Numerous transcriptional activators and repressors have been reported to contribute to these processes, but only few studies have focused on regulation at the post-transcriptional level in C. crescentus. Small RNAs (sRNAs) are a prominent class of regulators of bacterial gene expression, and most sRNAs characterized today engage in direct base-pairing interactions to modulate translation and/or stability of target mRNAs. In many cases, the ubiquitous RNA chaperone, Hfq, contributes to the establishment of RNA-RNA interactions. Although the deletion of the hfq gene is associated with a severe loss of fitness in C. crescentus, the RNA ligands of the chaperone have remained largely unexplored. Here we report on the identification of coding and non-coding transcripts associated with Hfq in C. crescentus, and demonstrate Hfq-dependent post-transcriptional regulation in this organism. We show that the conserved, Hfq-bound sRNA RusT is transcriptionally controlled by the conserved NtrYX two-component system and induced in response to iron starvation. By combining RusT pulse expression with whole-genome transcriptome analysis we determine 16 candidate target transcripts, more than half of which encode outer membrane transporters. We hence suggest RusT to support remodeling of the C. crescentus cell surface when iron supplies are limiting.
2023-11-16 | GSE247928 | GEO