Identification of two small regulatory RNAs linked to virulence in Brucella abortus 2308

Department of Microbiology and Immunology, East Carolina University School of Medicine, Greenville, NC 27834, USA.
Molecular Microbiology (Impact Factor: 4.42). 06/2012; 85(2):345-60. DOI: 10.1111/j.1365-2958.2012.08117.x
Source: PubMed


Hfq is an RNA-binding protein that functions in post-transcriptional gene regulation by mediating interactions between mRNAs and small regulatory RNAs (sRNAs). Two proteins encoded by BAB1_1794 and BAB2_0612 are highly over-produced in a Brucella abortus hfq mutant compared with the parental strain, and recently, expression of orthologues of these proteins in Agrobacterium tumefaciens was shown to be regulated by two sRNAs, called AbcR1 and AbcR2. Orthologous sRNAs (likewise designated AbcR1 and AbcR2) have been identified in B. abortus 2308. In Brucella, abcR1 and abcR2 single mutants are not defective in their ability to survive in cultured murine macrophages, but an abcR1 abcR2 double mutant exhibits significant attenuation in macrophages. Additionally, the abcR1 abcR2 double mutant displays significant attenuation in a mouse model of chronic Brucella infection. Quantitative proteomics and microarray analyses revealed that the AbcR sRNAs predominantly regulate genes predicted to be involved in amino acid and polyamine transport and metabolism, and Northern blot analyses indicate that the AbcR sRNAs accelerate the degradation of the target mRNAs. In an Escherichia coli two-plasmid reporter system, overexpression of either AbcR1 or AbcR2 was sufficient for regulation of target mRNAs, indicating that the AbcR sRNAs from B. abortus 2308 perform redundant regulatory functions.

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Available from: K. Sayood, Dec 11, 2014
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    • "Previous reports have shown that half of all known sRNAs in Escherichia [80] and Salmonella [81] associate with Hfq. Little is known about sRNAs in Brucella except for abcR1 and abcR2, which were recently reported by Caswell et al [82]. Our work reveals potential target genes for sRNAs regulation in B. melitensis. "
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    ABSTRACT: Brucella melitensis is a facultative intracellular bacterium that replicates within macrophages. The ability of brucellae to survive and multiply in the hostile environment of host macrophages is essential to its virulence. The RNA-binding protein Hfq is a global regulator that is involved in stress resistance and pathogenicity. Here we demonstrate that Hfq is essential for stress adaptation and intracellular survival in B. melitensis. A B. melitensis hfq deletion mutant exhibits reduced survival under environmental stresses and is attenuated in cultured macrophages and mice. Microarray-based transcriptome analyses revealed that 359 genes involved in numerous cellular processes were dysregulated in the hfq mutant. From these same samples the proteins were also prepared for proteomic analysis to directly identify Hfq-regulated proteins. Fifty-five proteins with significantly affected expression were identified in the hfq mutant. Our results demonstrate that Hfq regulates many genes and/or proteins involved in metabolism, virulence, and stress responses, including those potentially involved in the adaptation of Brucella to the oxidative, acid, heat stress, and antibacterial peptides encountered within the host. The dysregulation of such genes and/or proteins could contribute to the attenuated hfq mutant phenotype. These findings highlight the involvement of Hfq as a key regulator of Brucella gene expression and facilitate our understanding of the role of Hfq in environmental stress adaptation and intracellular survival of B. melitensis.
    PLoS ONE 08/2013; 8(8):e71933. DOI:10.1371/journal.pone.0071933 · 3.23 Impact Factor
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    • "Similar approaches also identified several ABC transporter mRNAs as AbcR1 and AbcR2 targets in A. tumefaciens and B. abortus [20], [21]. Preliminary computational predictions have rendered long and partially overlapping lists of target candidates for AbcR1 and AbcR2 in S. meliloti, with 35–45% of the top scoring hits corresponding to ABC transporter genes [11]. "
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    ABSTRACT: The legume symbiont Sinorhizobium meliloti expresses a plethora of small noncoding RNAs (sRNAs) whose function is mostly unknown. Here, we have functionally characterized two tandemly encoded S. meliloti Rm1021 sRNAs that are similar in sequence and structure. Homologous sRNAs (designated AbcR1 and AbcR2) have been shown to regulate several ABC transporters in the related α-proteobacteria Agrobacterium tumefaciens and Brucella abortus. In Rm1021, AbcR1 and AbcR2 exhibit divergent unlinked regulation and are stabilized by the RNA chaperone Hfq. AbcR1 is transcribed in actively dividing bacteria, either in culture, rhizosphere or within the invasion zone of mature alfalfa nodules. Conversely, AbcR2 expression is induced upon entry into stationary phase and under abiotic stress. Only deletion of AbcR1 resulted into a discrete growth delay in rich medium, but both are dispensable for symbiosis. Periplasmic proteome profiling revealed down-regulation of the branched-chain amino acid binding protein LivK by AbcR1, but not by AbcR2. A double-plasmid reporter assay confirmed the predicted specific targeting of the 5'-untranslated region of the livK mRNA by AbcR1 in vivo. Our findings provide evidences of independent regulatory functions of these sRNAs, probably to fine-tune nutrient uptake in free-living and undifferentiated symbiotic rhizobia.
    PLoS ONE 07/2013; 8(7):e68147. DOI:10.1371/journal.pone.0068147 · 3.23 Impact Factor
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    • "Small non-coding RNAs (ncRNAs) are transcripts that function in a bacterial cell structurally as RNA molecules rather than as templates for translation into polypeptides (Majdalani et al., 2005). ncRNAs play important regulatory roles in bacterial stress responses to diverse environmental signals, such as changes in temperature, osmolarity, iron and oxidative stress (Gottesman, 2005; Gottesman et al., 2006; Massé et al., 2007; Romby et al., 2006; Silvaggi et al., 2006), and have key roles in the regulation of virulence factors in a variety of pathogens (Altier et al., 2000; Caswell et al., 2012; De Lay & Gottesman, 2012; Hébrard et al., 2012; Kreikemeyer et al., 2001; Le Rhun & Charpentier, 2012; Lenz et al., 2005, 2004; Mangold et al., 2004; Sonnleitner et al., 2012), including Pseudomonas aeruginosa (Burrowes et al., 2005; Heurlier et al., 2004). A single ncRNA species can directly regulate multiple genes, leading to pleiotropic effects (Gottesman et al., 2006). "
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    ABSTRACT: Bacteria contain small non-coding RNAs (ncRNAs) that are responsible for altering transcription, translation, or mRNA stability. ncRNAs are important because they regulate virulence factors and susceptibility to various stresses. Here, the regulation of a recently described ncRNA of Pseudomonas syringae pv. tomato DC3000, P16 was investigated. We determined that RpoS regulates the expression of P16. We found that deletion of P16 results in increased sensitivity to hydrogen peroxide compared to the wild-type strain, suggesting that P16 plays a role in the bacteria's susceptibility to oxidative stress. Additionally the P16 mutant displayed enhanced resistance to heat stress. Our findings provide new information on the regulation and role of this ncRNA in P. syringae.
    Microbiology 12/2012; 159(Pt 2). DOI:10.1099/mic.0.063826-0 · 2.56 Impact Factor
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