Predicted structure and phyletic distribution of the RNA-binding protein Hfq

Sun, Xiao‐Man; Zhulin, Igor B.; Wartell, Roger M.

doi:10.1093/nar/gkf508

Cited by 191 publications

(190 citation statements)

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“…the welldocumented Hfq, have a role in the regulation of the expression of genes involved in virulence (Brennan & Link, 2007;Chao & Vogel, 2010; Christiansen et al, 2004;Vogel & Luisi, 2011). Of note, enterococci and streptococci do not harbour hfq-like genes (Sun et al, 2002).The aim of the present study was to further investigate the role of CspR in the virulence of E. faecalis. Interestingly, we observed that the addition of anti-CspR immune rabbit serum protected larvae of G. mellonella against killing by E. faecalis.…”

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Cold-shock RNA-binding protein CspR is also exposed to the surface of Enterococcus faecalis

et al. 2013

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CspR has been characterized recently as a cold-shock RNA-binding protein in Enterococcus faecalis, a natural member of the gastro-intestinal tract capable of switching from a commensal relationship with the host to an important nosocomial pathogen. In addition to its involvement in the cold-shock response, CspR also plays a role in the long-term survival and virulence of E. faecalis. In the present study, we demonstrated that anti-CspR immune rabbit serum protected larvae of Galleria mellonella against a lethal challenge of the WT strain. These results suggested that CspR might have a surface location. This hypothesis was verified by Western blot that showed detection of CspR in the total as well as in the surface protein fraction. In addition, identification of surface polypeptides by proteolytic shaving of intact bacterial cells followed by liquid chromatography-MS-MS revealed that cold-shock proteins (EF1367, EF2939 and CspR) were present on the cell surface. Lastly, anti-CspR immune rabbit serum was used for immunolabelling and detected with colloidal gold-labelled goat anti-rabbit IgG in order to determine the immunolocalization of CspR on E. faecalis WT strain. Electron microscopy images confirmed that the cold-shock protein RNA-binding protein CspR was present in both cytoplasmic and surface parts of the cell. These data strongly suggest that CspR, in addition to being located intracellularly, is also present in the extracellular protein fraction of the cells and has important functions in the infection process of Galleria larvae. INTRODUCTIONEnterococcus faecalis, a natural member of the intestinal tract, is versatile and can switch from a commensal relationship with the host to a leading cause of nosocomial infections (Murray, 1990;Sreeja et al., 2012; Wisplinghoff et al., 2004). This bacterium, well known to be able to cope with hostile environments (Ogier & Serror, 2008) and used as an indicator of faecal contamination, is responsible for serious infections such as endocarditis or surgical wound infection, especially in immunocompromised patients Shepard & Gilmore, 2002). Therefore, the virulence of E. faecalis has been intensively studied over the past 20 years and~12 putative virulence genes have been reported, including several transcriptional and post-transcriptional regulators. The recently identified cold-shock RNA-binding protein CspR is part of this last category of virulence-associated factors (Fox et al., 2009;Hancock & Gilmore, 2002;Lebreton et al., 2009;Michaux et al., 2011 Michaux et al., , 2012Qin et al., 2001; Shankar et al., 2001). In some Gram-positive bacteria such as Staphylococcus aureus and Bacillus subtilis, cold-shock polypeptides have been shown to be involved in several aspects of bacterial life, i.e. survival under starvation and low-temperature conditions or resistance to anti-microbial agents (Duval et al., 2010;Graumann & Marahiel, 1999, 1996 Graumann et al., 1997). In addition, the cold-shock polypeptides usually present the conserved RNA-binding motifs RNP-1 and RNP-...

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Section: Introductionmentioning

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Cold-shock RNA-binding protein CspR is also exposed to the surface of Enterococcus faecalis

et al. 2013

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“…Hfq shares sequence similarity to the eukaryotic Lsm proteins [23][24][25][26][27] We addressed these questions through a mutational analysis of Hfq, probing in vitro binding to several model RNAs that represent species with which Hfq must interact. Hfq mutants were assayed in vivo using a reporter assay and RNA lifetime experiments.…”

Section: Nih-pa Author Manuscriptmentioning

confidence: 99%

“…Hfq shares sequence similarity to the eukaryotic Lsm proteins [23][24][25][26][27] . In the Conserved Domain Database 28 Hfq is listed under the Sm and Sm-like protein family as well as among the eubacterial Hfqs.…”

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Escherichia coli Hfq has distinct interaction surfaces for DsrA, rpoS and poly(A) RNAs

Mikulecky

Kaw

Brescia

et al. 2004

Nat Struct Mol Biol

226

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The bacterial Sm-like protein Hfq facilitates RNA-RNA interactions involved in posttranscriptional regulation of the stress response. Specifically, Hfq helps pair noncoding RNAs (ncRNAs) with complementary regions of target mRNAs. To probe the mechanism of this pairing, we generated a series of Hfq mutants and measured their affinity for RNAs like those with which Hfq must associate in vivo. We tested the mutants' DsrA-dependent activation of rpoS, and their ability to stabilize DsrA ncRNA against degradation in vivo. Our results suggest that Hfq has two independent RNA-binding surfaces. In addition to a well-known site around the core of the Hfq hexamer, we observe interactions with the distal face of Hfq, a new locus with which mRNAs and poly(A) sequences associate. Our model explains how Hfq can simultaneously bind a ncRNA and its mRNA target to facilitate the strand displacement reaction required for Hfq-dependent translational regulation.Hfq protein from Escherichia coli was first described in connection with Qβ-phage replication 1,2 . Hfq has recently emerged as a central player in post-transcriptional gene regulation as mediated by bacterial ncRNAs [3][4][5][6] . Escherichia coli Hfq mutants show disrupted signaling in stress response pathways 7,8 , arising from the need for Hfq to mediate base-pairing between regulatory ncRNAs and their mRNA targets. Examples of these partnerships include DsrA-rpoS 7,9,10 , OxyS-fhlA 11,12 , OxyS-rpoS 13 , RprA-rpoS 14 , RyhBsodB [15][16][17] .Complexes between ncRNAs and their mRNA targets function in several ways. Most commonly, complexed structures lead to translational activation or repression by remodeling mRNA regulatory regions containing the ribosome-binding site (RBS) and/or start codon. Alternatively, the interaction can enhance decay of the target mRNA16 or simply block translation11. Clearly, Hfq facilitates base-pairing between ncRNAs and their targets, but how it does so is poorly understood. How the chaperone function relates to other Hfq activities such as the control of poly(A) tail elongation19 , 20 and regulation of mRNA stability21 , 22 is also unknown. COMPETING INTERESTS STATEMENTThe authors declare that they have no competing financial interests. NIH Public Access Author ManuscriptNat Struct Mol Biol. Author manuscript; available in PMC 2011 April 5. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptHfq shares sequence similarity to the eukaryotic Lsm proteins [23][24][25][26][27] We addressed these questions through a mutational analysis of Hfq, probing in vitro binding to several model RNAs that represent species with which Hfq must interact. Hfq mutants were assayed in vivo using a reporter assay and RNA lifetime experiments. Together, the results support a model wherein at least two independent RNA-binding sites exist on the Hfq hexamer, and juxtaposition of bound RNAs facilitates base-pairing. RESULTS Hfq mutagenesisTo identify amino acids essential for RNA binding, we constructed a series of E. coli Hfq misse...

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“…In 2002, a standard BLAST search identified Hfq in about half of the completed or nearly completed bacterial genomes (44). However, this standard BLAST search did not identify Hfq in cyanobacteria.…”

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Hfq Is Required for Optimal Nitrate Assimilation in the Cyanobacterium Anabaena sp. Strain PCC 7120

Puerta-Fernández

Vioque

2011

Journal of Bacteriology

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Hfq is an RNA binding protein involved in posttranscriptional regulation of gene expression in bacteria. It acts by binding to regulatory small RNAs (sRNAs), which confer specificity for the regulation. Recently, orthologues of the Hfq protein were annotated in cyanobacterial genomes, although its capacity to regulate gene expression by interacting with sRNAs has not been yet demonstrated. Anabaena sp. strain PCC 7120 is a filamentous cyanobacterium that, in the absence of combined nitrogen, is able to fix atmospheric nitrogen by differentiating specialized cells called heterocysts. We have generated an hfq knockout mutant of Anabaena sp. PCC 7120. Deletion of this gene results in differentiation of heterocysts in the presence of nitrate, suggesting a defect in nitrate assimilation. We show that hfq mutant cells are affected in transport and use of nitrate and nitrite. An analysis of the expression of several genes in the nir operon, encoding different elements of the nitrate assimilation pathway, demonstrates a downregulation of their transcription in mutant cells. We also observed that genes ntcB and cnaT, involved in the regulation of the nir operon, show a lower expression in cells lacking Hfq. Finally, when hfq was reintroduced in the mutant, heterocyst differentiation was no longer observed in the presence of nitrate. Therefore, our results indicate that the RNA chaperone Hfq is involved in the regulation of the nir operon, although the mechanism for this regulation is still unknown.

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Predicted structure and phyletic distribution of the RNA-binding protein Hfq

Cited by 191 publications

References 48 publications

Cold-shock RNA-binding protein CspR is also exposed to the surface of Enterococcus faecalis

Cold-shock RNA-binding protein CspR is also exposed to the surface of Enterococcus faecalis

Escherichia coli Hfq has distinct interaction surfaces for DsrA, rpoS and poly(A) RNAs

Hfq Is Required for Optimal Nitrate Assimilation in the Cyanobacterium Anabaena sp. Strain PCC 7120

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