Functional effects of variants of the RNA chaperone Hfq

Sonnleitner, Elisabeth; Napetschnig, Johanna; Afonyushkin, Taras; Ecker, Karin; Večerek, Branislav; Moll, Isabella; Kaberdin, Vladimir R.; Bläsi, Udo

doi:10.1016/j.bbrc.2004.08.190

Cited by 37 publications

(44 citation statements)

References 37 publications

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“…4) suggests that the CTD is more important for competition against other RNAs than the intrinsic stability of the Hfq-sRNA binding. The interplay of autoinhibition, sRNA competition, and sRNA accumulation could be the source of the conflicting results on the importance of the CTD (23,24,26,27).…”

Section: Discussionmentioning

confidence: 99%

“…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).…”

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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.

139

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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: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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.

139

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“…DsrA also plays a regulatory role in acid resistance (41). The regulatory effects of DsrA are mediated by specific RNA-to-RNA pairing interactions, while its stability and activity require the recruitment of Hfq (8,46,59,60).…”

Section: Deletion Of Lsrr and Lsrk Does Not Affect Growth Or Motilitymentioning

confidence: 99%

Quorum Sensing in Escherichia coli Is Signaled by AI-2/LsrR: Effects on Small RNA and Biofilm Architecture

et al. 2007

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The regulatory network for the uptake of Escherichia coli autoinducer 2 (AI-2) is comprised of a transporter complex, LsrABCD; its repressor, LsrR; and a cognate signal kinase, LsrK. This network is an integral part of the AI-2 quorum-sensing (QS) system. Because LsrR and LsrK directly regulate AI-2 uptake, we hypothesized that they might play a wider role in regulating other QS-related cellular functions. In this study, we characterized physiological changes due to the genomic deletion of lsrR and lsrK. We discovered that many genes were coregulated by lsrK and lsrR but in a distinctly different manner than that for the lsr operon (where LsrR serves as a repressor that is derepressed by the binding of phospho-AI-2 to the LsrR protein). An extended model for AI-2 signaling that is consistent with all current data on AI-2, LuxS, and the LuxS regulon is proposed. Additionally, we found that both the quantity and architecture of biofilms were regulated by this distinct mechanism, as lsrK and lsrR knockouts behaved identically. Similar biofilm architectures probably resulted from the concerted response of a set of genes including flu and wza, the expression of which is influenced by lsrRK. We also found for the first time that the generation of several small RNAs (including DsrA, which was previously linked to QS systems in Vibrio harveyi) was affected by LsrR. Our results suggest that AI-2 is indeed a QS signal in E. coli, especially when it acts through the transcriptional regulator LsrR.Bacteria communicate with each other through small "hormone-like" organic molecules referred to as autoinducers. Autoinducer-based bacterial cell-to-cell communication, enabling population-based multicellularity, has been termed quorum sensing (QS) (27). Cellular functions controlled by QS are varied and reflect the needs of a particular bacterial species for inhabiting a given niche (10,38,65).QS among Escherichia coli and Salmonella strains has been a topic of great interest, and different intercellular signaling systems have been identified: that mediated by the LuxR homolog SdiA; the LuxS/autoinducer 2 (AI-2) system, an AI-3 system, and a signaling system mediated by indole (2,19,36,57,61,68). Among these systems, the LuxS/AI-2 system possesses the unique feature of endowing cell population-dependent behavior while interacting with central metabolism through the intracellular activated methyl cycle (20,21,45,65,73). Therefore, it has the potential to influence both gene regulation and bacterial fitness.AI-2's function has been studied using luxS mutants and by adding either conditioned medium or in vitro-synthesized AI-2 to bacterial cultures. It is noteworthy that the luxS transcription profile is not synchronous with the accumulation profile of extracellular AI-2 in bacterial supernatants (5,31,75). In E.coli, extracellular AI-2 activity peaks during the mid-to lateexponential phase and rapidly decreases during entry into the stationary phase. A corresponding decrease in LuxS protein levels is not observed (31,75). The disap...

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“…Second, mutational analyses of Eco-Hfq indicate that additional conserved residues on the proximal face are involved in RNA binding. Specifically, Hfq point mutations at Lys3 or Arg17 result in a protein that cannot support Qb replication, and residue Arg16 has been implicated in binding of the regulatory DsrA RNA (Brescia et al 2003;Sonnleitner et al 2004;Sun and Wartell 2006). In addition, data from mutational analyses indicate the presence of another binding site on the distal face of the Eco-Hfq hexamer that specifically recognizes poly(A) RNA (Mikulecky et al 2004;Sun and Wartell 2006).…”

Section: Expression Isolation and Crystallization Of Mja-hfqmentioning

confidence: 99%

An Hfq-like protein in archaea: Crystal structure and functional characterization of the Sm protein from Methanococcus jannaschii

Nielsen¹,

Bøggild²,

Andersen³

et al. 2007

RNA

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The Sm and Sm-like proteins are conserved in all three domains of life and have emerged as important players in many different RNA-processing reactions. Their proposed role is to mediate RNA-RNA and/or RNA-protein interactions. In marked contrast to eukaryotes, bacteria appear to contain only one distinct Sm-like protein belonging to the Hfq family of proteins. Similarly, there are generally only one or two subtypes of Sm-related proteins in archaea, but at least one archaeon, Methanococcus jannaschii, encodes a protein that is related to Hfq. This archaeon does not contain any gene encoding a conventional archaeal Sm-type protein, suggesting that Hfq proteins and archaeal Sm-homologs can complement each other functionally. Here, we report the functional characterization of M. jannaschii Hfq and its crystal structure at 2.5 Å resolution. The protein forms a hexameric ring. The monomer fold, as well as the overall structure of the complex is similar to that found for the bacterial Hfq proteins. However, clear differences are seen in the charge distribution on the distal face of the ring, which is unusually negative in M. jannaschii Hfq. Moreover, owing to a very short N-terminal a-helix, the overall diameter of the archaeal Hfq hexamer is significantly smaller than its bacterial counterparts. Functional analysis reveals that Escherichia coli and M. jannaschii Hfqs display very similar biochemical and biological properties. It thus appears that the archaeal and bacterial Hfq proteins are largely functionally interchangeable.

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Functional effects of variants of the RNA chaperone Hfq

Cited by 37 publications

References 37 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

Quorum Sensing in Escherichia coli Is Signaled by AI-2/LsrR: Effects on Small RNA and Biofilm Architecture

An Hfq-like protein in archaea: Crystal structure and functional characterization of the Sm protein from Methanococcus jannaschii

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