The binding of Class II sRNA MgrR to two different sites on matchmaker protein Hfq enables efficient competition for Hfq and annealing to regulated mRNAs

Kwiatkowska, J.; Wróblewska, Zuzanna; Johnson, Kenneth A.; Olejniczak, Mikołaj

doi:10.1261/rna.067777.118

Cited by 20 publications

(21 citation statements)

References 56 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…An interaction between RelA-like protein and RNA was documented previously for RelQ (40). The authors showed that the small alarmone synthetase RelQ from the Gram- (17,24,(45)(46)(47). Also, ChiX, MgrR and DsrA that lack the GGAG sequence were reported to be better competitors for binding Hfq than RyhB, McaS or CyaR sRNAs (46).…”

Section: Rela Ggag Srna Binding and (P)ppgpp Productions Are Mutuallymentioning

confidence: 66%

“…The authors showed that the small alarmone synthetase RelQ from the Gram- (17,24,(45)(46)(47). Also, ChiX, MgrR and DsrA that lack the GGAG sequence were reported to be better competitors for binding Hfq than RyhB, McaS or CyaR sRNAs (46). Interestingly, both MgrR and ChiX carry in addition to poly(U) tail three and four ARN motifs, respectively (25,45).…”

Section: Rela Ggag Srna Binding and (P)ppgpp Productions Are Mutuallymentioning

confidence: 99%

See 1 more Smart Citation

RNA binding of Hfq monomers promotes RelA-mediated hexamerization in a limiting Hfq environment

Basu¹,

Elgrably‐Weiss²,

Hassouna³

et al. 2020

Preprint

View full text Add to dashboard Cite

The RNA chaperone Hfq acting as a hexamer, is a known mediator of post-transcriptional regulation expediting basepairing between small RNAs (sRNAs) and their target mRNAs. However, the intricate details associated with Hfq-RNA biogenesis are still unclear. Previously, we reported that the stringent response regulator, RelA is a functional partner of Hfq that facilitates Hfq-mediated sRNA-mRNA regulation in vivo and induces Hfq hexamerization in vitro. Here, for the first time we show that RelA-mediated Hfq hexamerization requires an initial binding of RNA, preferably sRNA to Hfq monomers. By interacting with a Shine-Dalgarno-like sequence (GGAG) in the sRNA, RelA stabilizes the initially unstable complex of RNA bound-Hfq monomer, enabling the attachment of more Hfq subunits to form a functional hexamer. Overall, our study showing that RNA binding to Hfq monomers is at the heart of RelA-mediated Hfq hexamerization, challenges the previous concept that only Hfq hexamers can bind RNA.

show abstract

Section: Rela Ggag Srna Binding and (P)ppgpp Productions Are Mutuallymentioning

confidence: 66%

Section: Rela Ggag Srna Binding and (P)ppgpp Productions Are Mutuallymentioning

confidence: 99%

RNA binding of Hfq monomers promotes RelA-mediated hexamerization in a limiting Hfq environment

Basu¹,

Elgrably‐Weiss²,

Hassouna³

et al. 2020

Preprint

View full text Add to dashboard Cite

show abstract

“…1E, F). This approach was used before to show that ChiX and MgrR sRNAs bind both the distal face and the proximal face of Hfq [38, 39], while RybB sRNA binds only to the proximal face of Hfq, in agreement with previous studies [29, 31]. Our data showed that both AgvB and 3′ETS leuZ easily induce the dissociation of A 27 -Hfq complexes, with low IC 50 values (3.2 ± 1.4 nM and 2.8 ± 0.35 nM, respectively, Fig.…”

Section: Resultsmentioning

confidence: 99%

“…4D). In this way GcvB resembles mRNAs targeted by class II sRNAs, such as eptB or chiP , for which the rim mutant also did not stimulate annealing (Kwiatkowska et al, 2018). Like class II mRNAs, the sequence of GcvB is rich in uridines which mediate the interactions with the Hfq rim [28, 29, 46].…”

Section: Discussionmentioning

confidence: 99%

Bacterial chaperone protein Hfq facilitates the annealing of sponge RNAs to small regulatory RNAs

Małecka

Sobańska

Olejniczak

2021

Preprint

View full text Add to dashboard Cite

Bacterial small RNAs (sRNAs) in association with the chaperone protein Hfq regulate the expression of many target mRNAs. Since sRNAs’ action is crucial to engender a response to changing environmental conditions, their activity needs to be regulated. One such mechanism occurs at posttranscriptional level and involves sponge RNAs (or anti-sRNAs) which sequester sRNAs affecting their regulatory output. Both types of RNAs were identified on Hfq, but it is not known how Hfq interacts with RNA sponges and stimulates their base-pairing with sRNAs. Here, we used biochemical methods to demonstrate that anti-sRNAs resemble sRNAs by their structure and their modes of Hfq binding. Hfq facilitates sponge RNA annealing to sRNA, and each surface of the protein plays a particular role in the process. Moreover, we found that the efficiency of sponge RNA interactions with sRNAs can be improved, therefore, we propose that natural RNA sponges might not sequester sRNAs optimally.

show abstract

“…8B) (9,12). While Hfq can bind A-rich sequences, for example ARN motifs, in some of its RNA ligands, these sequences are located either in mRNA 5ʹ-UTRs (35)(36)(37) or far upstream of intrinsic terminators in a subgroup of Hfq-specific sRNAs, called Class II sRNAs (40)(41)(42). However, in the dominant subgroup of Hfq-binding sRNAs, called Class I sRNAs, the Hfq-binding module consists of a U-rich motif just upstream of the intrinsic terminator, which is followed by a long 3ʹ oligoUtail (39,54).…”

Section: A Terminator Adjacent A-rich Motif Serves As a Negative Detementioning

confidence: 99%

Determinants of RNA recognition by the FinO domain of theEscherichia coliProQ protein

Stein

Kwiatkowska

Basczok

et al. 2020

Preprint

Self Cite

View full text Add to dashboard Cite

The regulation of gene expression by small RNAs in Escherichia coli depends on RNA binding proteins Hfq and ProQ, which bind mostly distinct RNA pools. To understand how ProQ discriminates between RNA substrates, we compared its binding to six different RNA molecules. Full-length ProQ bound all six RNAs similarly, while the isolated N-terminal FinO domain (NTD) of ProQ specifically recognized RNAs with Rho-independent terminators. Analysis of malM 3ʹ-UTR mutants showed that tight RNA binding by the ProQ NTD required a terminator hairpin of at least two base pairs preceding an 3ʹ oligoU tail of at least four uridine residues. Substitution of an A-rich sequence on the 5ʹ side of the terminator to uridines strengthened the binding of several ProQ-specific RNAs to the Hfq protein, but not to the ProQ NTD. Substitution of the motif in the malM-3ʹ and cspE-3ʹ RNAs also conferred the ability to bind Hfq in E. coli cells, as measured using a three-hybrid assay. In summary, these data suggest that the ProQ NTD specifically recognizes 3ʹ intrinsic terminators of RNA substrates, and that the discrimination between RNA ligands by E. coli ProQ and Hfq depends both on positive determinants for binding to ProQ and negative determinants against binding to Hfq.RNA-binding proteins are important contributors to major life processes, including the regulation of gene expression by RNAs (1). In many bacteria, the prominent role played by trans-encoded base-pairing small RNAs (sRNAs) in regulating gene expression requires a matchmaker protein called Hfq, which assists sRNA in pairing to complementary sequences in target mRNAs and affects sRNA stability (2-6). Global identification of new RNA/protein interactions has been enabled by deep-sequencing approaches (7), such as Grad-seq, which uses glycerol gradients to identify novel RNA/protein complexes (8), CLIP-seq, which uses crosslinking to define protein binding sites in the transcriptome (9), and RIL-seq which identifies protein-dependent RNA-RNA interactions (10). Recent studies using these approaches showed that another protein, named ProQ, is a global RNA binding protein in Escherichia coli and Salmonella enterica (11-13), and is involved in sRNA interactions with other RNA molecules (12,14,15). The pool of RNA ligands of ProQ is mostly distinct from that of Hfq and contains more mRNAs than sRNAs (11,12).While ProQ was originally discovered in a search for genes that affect proline transport (16), this protein contributes to several physiological processes in E.coli and S. enterica (17)including DNA metabolism (13,14), bacterial virulence (15), and adaptation to osmotic stress (12) and resource limitation (18). ProQ is active as a monomer (19), and it belongs to the FinO-domain family with representatives in numerous γ-proteobacteria (17,20). Other members of this family include the F-like plasmid FinO protein (21,22), Legionella pneumophila RocC protein (23), and S. enterica pCol1B9 plasmid-encoded FopA protein (J.Vogel, personal communication), which each interact with few RNAs...

show abstract

The binding of Class II sRNA MgrR to two different sites on matchmaker protein Hfq enables efficient competition for Hfq and annealing to regulated mRNAs

Cited by 20 publications

References 56 publications

RNA binding of Hfq monomers promotes RelA-mediated hexamerization in a limiting Hfq environment

RNA binding of Hfq monomers promotes RelA-mediated hexamerization in a limiting Hfq environment

Bacterial chaperone protein Hfq facilitates the annealing of sponge RNAs to small regulatory RNAs

Determinants of RNA recognition by the FinO domain of theEscherichia coliProQ protein

Contact Info

Product

Resources

About