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Positive regulation by small RNAs and the role of Hfq.

ABSTRACT: Bacterial small noncoding RNAs carry out both positive and negative regulation of gene expression by pairing with mRNAs; in Escherichia coli, this regulation often requires the RNA chaperone Hfq. Three small regulatory RNAs (sRNAs), DsrA, RprA, and ArcZ, positively regulate translation of the sigma factor RpoS, each pairing with the 5' leader to open up an inhibitory hairpin. In vitro, rpoS interaction with sRNAs depends upon an (AAN)(4) Hfq-binding site upstream of the pairing region. Here we show that both Hfq and this Hfq binding site are required for RprA or ArcZ to act in vivo and to form a stable complex with rpoS mRNA in vitro; both were partially dispensable for DsrA at 37 degrees C. ArcZ sRNA is processed from 121 nt to a stable 56 nt species that contains the pairing region; only the 56 nt ArcZ makes a strong Hfq-dependent complex with rpoS. For each of these sRNAs, the stability of the sRNA*mRNA complexes, rather than their rate of formation, best predicted in vivo activity. These studies demonstrate that binding of Hfq to the rpoS mRNA is critical for sRNA regulation under normal conditions, but if the stability of the sRNA*mRNA complex is sufficiently high, the requirement for Hfq can be bypassed.

SUBMITTER: Soper T

PROVIDER: S-EPMC2906882 | biostudies-other | 2010 May

REPOSITORIES: biostudies-other

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Positive regulation by small RNAs and the role of Hfq.

Soper Toby T Mandin Pierre P Majdalani Nadim N Gottesman Susan S Woodson Sarah A SA

Proceedings of the National Academy of Sciences of the United States of America 20100510 21

Bacterial small noncoding RNAs carry out both positive and negative regulation of gene expression by pairing with mRNAs; in Escherichia coli, this regulation often requires the RNA chaperone Hfq. Three small regulatory RNAs (sRNAs), DsrA, RprA, and ArcZ, positively regulate translation of the sigma factor RpoS, each pairing with the 5' leader to open up an inhibitory hairpin. In vitro, rpoS interaction with sRNAs depends upon an (AAN)(4) Hfq-binding site upstream of the pairing region. Here we s ...[more]

PMID: 20457943

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Project description:BackgroundErwinia amylovora is a phytopathogenic bacterium and causal agent of fire blight disease in apples and pears. Although many virulence factors have been characterized, the coordination of expression of these virulence factors in E. amylovora is still not clear. Regulatory small RNAs (sRNAs) are important post-transcriptional regulatory components in bacteria. A large number of sRNAs require the RNA chaperone Hfq for both stability and functional activation. In E. amylovora, Hfq was identified as a major regulator of virulence and various virulence traits. However, information is still lacking about Hfq-dependent sRNAs on a genome scale, including the virulence regulatory functions of these sRNAs in E. amylovora.ResultsUsing both an RNA-seq analysis and a Rho-independent terminator search, 40 candidate Hfq-dependent sRNAs were identified in E. amylovora. The expression and sizes of 12 sRNAs and the sequence boundaries of seven sRNAs were confirmed by Northern blot and 5' RACE assay respectively. Sequence conservation analysis identified sRNAs conserved only in the Erwinia genus as well as E. amylovora species-specific sRNAs. In addition, a dynamic re-patterning of expression of Hfq-dependent sRNAs was observed at 6 and 12 hours after induction in Hrp-inducing minimal medium. Furthermore, sRNAs that control virulence traits were characterized, among which ArcZ positively controls the type III secretion system (T3SS), amylovoran exopolysaccahride production, biofilm formation, and motility, and negatively modulates attachment while RmaA (Hrs6) and OmrAB both negatively regulate amylovoran production and positively regulate motility.ConclusionsThis study has significantly enhanced our understanding of the Hfq-dependent sRNAs in E. amylovora at the genome level. The identification of multiple virulence-regulating sRNAs also suggests that post-transcriptional regulation by sRNAs may play a role in the deployment of virulence factors needed during varying stages of pathogenesis during host invasion by E. amylovora.

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Positive regulation by small RNAs and the role of Hfq.

Publications

Positive regulation by small RNAs and the role of Hfq.

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