MtvR Is a Global Small Noncoding Regulatory RNA in Burkholderia cenocepacia

ABSTRACT Burkholderia cenocepacia J2315 is a highly epidemic and transmissible clinical isolate of the Burkholderia cepacia complex (Bcc), a group of bacteria causing life-threatening respiratory infections among cystic fibrosis patients. This work describes the functional analysis of the 136-nucleotide (nt)-long MtvR small noncoding RNA (sRNA) from the Bcc member B. cenocepacia J2315, with homologues restricted to the genus Burkholderia. Bioinformatic target predictions revealed a total of 309 mRNAs to be putative MtvR targets. The mRNA levels corresponding to 17 of 19 selected genes were found to be affected when MtvR was either overexpressed or silenced. Analysis of the interaction between MtvR and the hfq mRNA, one of its targets, showed that the sRNA binds exclusively to the 5′ untranslated region (UTR) of the hfq mRNA. This interaction resulted in decreased protein synthesis, suggesting a negative regulatory effect of MtvR on the RNA chaperone Hfq. Bacterial strains with MtvR silenced or overexpressed exhibited pleiotropic phenotypes related to growth and survival after several stresses, swimming and swarming motilities, biofilm formation, resistance to antibiotics, and ability to colonize and kill the nematode Caenorhabditis elegans. Together, the results indicate that the MtvR sRNA is a major posttranscriptional regulator in B. cenocepacia.

[1]  Eduardo P C Rocha,et al.  The Genome of Burkholderia cenocepacia J2315, an Epidemic Pathogen of Cystic Fibrosis Patients , 2008, Journal of bacteriology.

[2]  S. Gottesman,et al.  Regulation of Proteolysis of the Stationary-Phase Sigma Factor RpoS , 1998, Journal of bacteriology.

[3]  U. Pennsylvania,et al.  Clinical and Laboratory Standards Institute , 2019, Springer Reference Medizin.

[4]  M. Valvano,et al.  Construction of Aminoglycoside-Sensitive Burkholderia cenocepacia Strains for Use in Studies of Intracellular Bacteria with the Gentamicin Protection Assay , 2010, Applied and Environmental Microbiology.

[5]  S. Sousa,et al.  The Second RNA Chaperone, Hfq2, Is Also Required for Survival under Stress and Full Virulence of Burkholderia cenocepacia J2315 , 2011, Journal of bacteriology.

[6]  C. K. Vanderpool,et al.  Molecular call and response: the physiology of bacterial small RNAs. , 2011, Biochimica et biophysica acta.

[7]  B. Felden,et al.  A Staphylococcus aureus Small RNA Is Required for Bacterial Virulence and Regulates the Expression of an Immune-Evasion Molecule , 2010, PLoS pathogens.

[8]  Michael Zuker,et al.  Mfold web server for nucleic acid folding and hybridization prediction , 2003, Nucleic Acids Res..

[9]  S. Sousa,et al.  Pathogenicity, virulence factors, and strategies to fight against Burkholderia cepacia complex pathogens and related species , 2010, Applied Microbiology and Biotechnology.

[10]  S. Sousa,et al.  Studies on the Involvement of the Exopolysaccharide Produced by Cystic Fibrosis-Associated Isolates of the Burkholderia cepacia Complex in Biofilm Formation and in Persistence of Respiratory Infections , 2004, Journal of Clinical Microbiology.

[11]  T. Coenye,et al.  Molecular Mechanisms of Chlorhexidine Tolerance in Burkholderia cenocepacia Biofilms , 2011, Antimicrobial Agents and Chemotherapy.

[12]  P. Dřevínek,et al.  Burkholderia cenocepacia in cystic fibrosis: epidemiology and molecular mechanisms of virulence. , 2010, Clinical microbiology and infection : the official publication of the European Society of Clinical Microbiology and Infectious Diseases.

[13]  E. Sonnleitner,et al.  The C-terminal domain of Escherichia coli Hfq is required for regulation , 2007, Nucleic acids research.

[14]  I. Zhulin,et al.  Predicted structure and phyletic distribution of the RNA-binding protein Hfq. , 2002, Nucleic acids research.

[15]  S. Sousa,et al.  The hfq gene is required for stress resistance and full virulence of Burkholderia cepacia to the nematode Caenorhabditis elegans. , 2010, Microbiology.

[16]  M. Valvano,et al.  A Decade of Burkholderia cenocepacia Virulence Determinant Research , 2010, Infection and Immunity.

[17]  J. Vogel,et al.  Hfq and its constellation of RNA , 2011, Nature Reviews Microbiology.

[18]  Leo Eberl,et al.  Burkholderia cenocepacia J2315 acyl carrier protein: a potential target for antimicrobials' development? , 2008, Microbial pathogenesis.

[19]  B. Večerek,et al.  Translational autocontrol of the Escherichia coli hfq RNA chaperone gene. , 2005, RNA.

[20]  M. Ferraro Performance standards for antimicrobial susceptibility testing , 2001 .

[21]  Hua Li,et al.  Construction of two mathematical models for prediction of bacterial sRNA targets. , 2008, Biochemical and biophysical research communications.

[22]  R. Giegerich,et al.  Fast and effective prediction of microRNA/target duplexes. , 2004, RNA.

[23]  M. Valvano,et al.  Construction and Evaluation of Plasmid Vectors Optimized for Constitutive and Regulated Gene Expression in Burkholderia cepacia Complex Isolates , 2002, Applied and Environmental Microbiology.

[24]  D A Stahl,et al.  Fluorescent-oligonucleotide probing of whole cells for determinative, phylogenetic, and environmental studies in microbiology , 1990, Journal of bacteriology.

[25]  G. Riccardi,et al.  Molecular approaches to pathogenesis study of Burkholderia cenocepacia, an important cystic fibrosis opportunistic bacterium , 2011, Applied Microbiology and Biotechnology.

[26]  S. Gottesman Micros for microbes: non-coding regulatory RNAs in bacteria. , 2005, Trends in genetics : TIG.

[27]  S. Sousa,et al.  Variation of the antimicrobial susceptibility profiles of Burkholderia cepacia complex clonal isolates obtained from chronically infected cystic fibrosis patients: a five-year survey in the major Portuguese treatment center , 2008, European Journal of Clinical Microbiology & Infectious Diseases.

[28]  Clinical,et al.  Performance Standards for Antimicrobial Susceptibility Testing; Eighteenth Informational Supplement , 2008 .

[29]  J. Leitão,et al.  Experimental identification of small non-coding regulatory RNAs in the opportunistic human pathogen Burkholderia cenocepacia J2315. , 2013, Genomics.

[30]  J. Leitão,et al.  The Novel Cis-Encoded Small RNA h2cR Is a Negative Regulator of hfq2 in Burkholderia cenocepacia , 2012, PloS one.

[31]  Ryan T Gill,et al.  Identification of putative noncoding RNA genes in the Burkholderia cenocepacia J2315 genome. , 2007, FEMS microbiology letters.

[32]  J. Vogel,et al.  Regulatory RNA in bacterial pathogens. , 2010, Cell host & microbe.