Structure of Bacterial Regulatory RNAs Determines Their Performance in Competition for the Chaperone Protein Hfq

Małecka, Ewelina M.; Stróżecka, Joanna; Sobańska, Daria; Olejniczak, Mikołaj

doi:10.1021/bi500741d

Cited by 53 publications

(83 citation statements)

References 60 publications

(269 reference statements)

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“…By contrast, competition for binding to Hfq102 varied far more than predicted by the thermodynamic stabilities of the individual sRNA·Hfq102 complexes as seen previously (35). Although fulllength Hfq102 prioritizes Class II sRNAs, Hfq65 binds most sRNAs equally.…”

Section: Resultsmentioning

confidence: 72%

“…2) and improves binding of structured RNAs to the rim (Table 1), we next asked whether the CTD influences competition between different sRNAs for Hfq. The competition of sRNAs for Hfq has been attributed to a combination of sRNA secondary structure and sequence (12,35,43), and the ability of one sRNA to outcompete another does not necessarily correspond to their relative binding affinities (35,44), consistent with active displacement of bound sRNAs by other sRNAs in solution (37).…”

Section: Resultsmentioning

confidence: 99%

“…3 A-C). The sRNA competitors included DsrA, a Class I sRNA that is a poor competitor (12,35,44), RyhB (Class I), RprA, which has an AAN sequence like Class II sRNAs but exhibits a mixed Class I/Class II phenotype in E. coli (31), and ChiX (Class II). The fraction of 32 P-ChiX bound to either Hfq102 (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…5C, the kinetic advantage of RNAs possessing AAN motifs over RNAs that only bind the proximal surface (Fig. 3) explains the superior competition for Hfq by ChiX sRNA (12,35) and flhA mRNA (56). Our results suggest that removing the CTD nullifies the advantage of distal face binding, collapsing the range of competition between different sRNAs (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…This possibility became more apparent with the recognition that E. coli sRNAs can be divided into two classes based on their interactions with Hfq (31). The common Class I sRNAs typically bind the proximal face and rim of Hfq (14,(32)(33)(34), whereas a smaller number of Class II sRNAs contact the distal face of Hfq via an AAN motif near the 5′ end of the sRNA in addition to the proximal face (31,35). The CTD is not essential for sRNA recognition, because truncated Hfq variants lacking all or part of the CTD bind sRNAs in vitro and in the cell (10, 23-26, 28, 36).…”

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confidence: 99%

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C-terminal domain of the RNA chaperone Hfq drives sRNA competition and release of target RNA

Santiago-Frangos

Kumari

Schu

et al. 2016

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

149

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The bacterial Sm protein and RNA chaperone Hfq stabilizes small noncoding RNAs (sRNAs) and facilitates their annealing to mRNA targets involved in stress tolerance and virulence. Although an arginine patch on the Sm core is needed for Hfq's RNA chaperone activity, the function of Hfq's intrinsically disordered C-terminal domain (CTD) has remained unclear. Here, we use stopped flow spectroscopy to show that the CTD of Escherichia coli Hfq is not needed to accelerate RNA base pairing but is required for the release of dsRNA. The Hfq CTD also mediates competition between sRNAs, offering a kinetic advantage to sRNAs that contact both the proximal and distal faces of the Hfq hexamer. The change in sRNA hierarchy caused by deletion of the Hfq CTD in E. coli alters the sRNA accumulation and the kinetics of sRNA regulation in vivo. We propose that the Hfq CTD displaces sRNAs and annealed sRNA·mRNA complexes from the Sm core, enabling Hfq to chaperone sRNA-mRNA interactions and rapidly cycle between competing targets in the cell.A member of the Sm protein family, Hfq was first identified as a host factor for phage Q beta. Hfq is found in at least 50% of sequenced bacterial genomes (1) and, in many bacteria, contributes to posttranscriptional regulation by small noncoding RNAs (sRNAs). Deletion of Hfq leads to pleiotropic effects, such as altered cellular morphology, slow growth, maladaptation to stress, and avirulence (2-5).Escherichia coli Hfq comprises an Sm domain (amino acids 7-66) that assembles into a stable hexameric ring and an intrinsically disordered C-terminal domain (CTD) that projects from the rim of the hexamer (6-10). The Sm ring binds to both sRNAs and target mRNAs, stabilizing the sRNAs against turnover (11-13) and facilitating base pairing with complementary sequences in the mRNA (7,14,15). The conserved "proximal" face of the Hfq hexamer interacts with uridines at the sRNA 3′ end (9, 16, 17), whereas the "distal" face of Hfq binds AAN triplet repeats (9, 16, 17) often found in the 5′ UTRs of target mRNAs. These sequence-specific interactions recruit sRNAs and mRNAs to Hfq, allowing arginine-rich patches along the rim of Hfq to catalyze base pairing between complementary strands (18).Although the functional importance of the Sm domain is established, the function of the disordered CTD has been unclear. The Hfq CTD varies greatly in length and sequence composition between bacterial families (1, 19), ranging from 7-residue stubs in Bacillaceae (20) to 100-residue tails in Moraxellaceae (21, 22) (Fig. S1). Previous studies reached conflicting conclusions about the importance of the Hfq CTD for sRNA regulation. In early studies, C-terminal deletions of hfq had no obvious phenotype in E. coli (23, 24) and little effect on sRNA binding (23,25). By contrast, later studies found that the CTD was required for in vitro annealing and proper regulation by sRNAs and normal binding to long RNAs, such as the rpoS mRNA (19,26,27). Moreover, Hfq from Pseudomonas aeruginosa and Clostridium difficile, which have much shor...

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Section: Resultsmentioning

confidence: 72%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

See 3 more Smart Citations

C-terminal domain of the RNA chaperone Hfq drives sRNA competition and release of target RNA

Santiago-Frangos

Kumari

Schu

et al. 2016

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

149

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Riboregulation of bacterial and archaeal transposition

Ellis

Haniford

2016

WIREs RNA

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The coexistence of transposons with their hosts depends largely on transposition levels being tightly regulated to limit the mutagenic burden associated with frequent transposition. For 'DNA-based' (class II) bacterial transposons there is growing evidence that regulation through small noncoding RNAs and/or the RNA-binding protein Hfq are prominent mechanisms of defense against transposition. Recent transcriptomics analyses have identified many new cases of antisense RNAs (asRNA) that potentially could regulate the expression of transposon-encoded genes giving the impression that asRNA regulation of DNA-based transposons is much more frequent than previously thought. Hfq is a highly conserved bacterial protein that plays a central role in posttranscriptional gene regulation and stress response pathways in many bacteria. Three different mechanisms for Hfq-directed control of bacterial transposons have been identified to date highlighting the versatility of this protein as a regulator of bacterial transposons. There is also evidence emerging that some DNA-based transposons encode RNAs that could regulate expression of host genes. In the case of IS200, which appears to have lost its ability to transpose, contributing a regulatory RNA to its host could account for the persistence of this mobile element in a wide range of bacterial species. It remains to be seen how prevalent these transposon-encoded RNA regulators are, but given the relatively large amount of intragenic transcription in bacterial genomes, it would not be surprising if new examples are forthcoming.

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Hfq chaperone brings speed dating to bacterial sRNA

2018

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Hfq is a ubiquitous, Sm-like RNA binding protein found in most bacteria and some archaea. Hfq binds small regulatory RNAs (sRNAs), facilitates base pairing between sRNAs and their mRNA targets, and directly binds and regulates translation of certain mRNAs. Because sRNAs regulate many stress response pathways in bacteria, Hfq is essential for adaptation to different environments and growth conditions. The chaperone activities of Hfq arise from multipronged RNA binding by three different surfaces of the Hfq hexamer. The manner in which the structured Sm core of Hfq binds RNA has been well studied, but recent work shows that the intrinsically disordered C-terminal domain of Hfq modulates sRNA binding, creating a kinetic hierarchy of RNA competition for Hfq and ensuring the release of double-stranded sRNA-mRNA complexes. A combination of structural, biophysical, and genetic experiments reveals how Hfq recognizes its RNA substrates and plays matchmaker for sRNAs and mRNAs in the cell. The interplay between structured and disordered domains of Hfq optimizes sRNA-mediated post-transcriptional regulation, and is a common theme in RNA chaperones. This article is categorized under: Regulatory RNAs/RNAi/Riboswitches > Regulatory RNAs RNA Interactions with Proteins and Other Molecules > RNA-Protein Complexes RNA Structure and Dynamics > RNA Structure, Dynamics, and Chemistry.

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Structure of Bacterial Regulatory RNAs Determines Their Performance in Competition for the Chaperone Protein Hfq

Cited by 53 publications

References 60 publications

C-terminal domain of the RNA chaperone Hfq drives sRNA competition and release of target RNA

C-terminal domain of the RNA chaperone Hfq drives sRNA competition and release of target RNA

Riboregulation of bacterial and archaeal transposition

Hfq chaperone brings speed dating to bacterial sRNA

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