Journal article
Functional Analyses of the RsmY and RsmZ Small Noncoding Regulatory RNAs in Pseudomonas aeruginosa
Journal of bacteriology, Vol.200(11), e00736-17
06/01/2018
DOI: 10.1128/JB.00736-17
PMCID: PMC5952390
PMID: 29463606
Abstract
Pseudomonas aeruginosa
is a Gram-negative opportunistic pathogen with distinct acute and chronic virulence phenotypes. Whereas acute virulence is typically associated with expression of a type III secretion system (T3SS), chronic virulence is characterized by biofilm formation. Many of the phenotypes associated with acute and chronic virulence are inversely regulated by RsmA and RsmF. RsmA and RsmF are both members of the CsrA family of RNA-binding proteins and regulate protein synthesis at the posttranscriptional level. RsmA activity is controlled by two small noncoding regulatory RNAs (RsmY and RsmZ). Bioinformatic analyses suggest that RsmY and RsmZ each have 3 or 4 putative RsmA binding sites. Each predicted binding site contains a GGA sequence presented in the loop portion of a stem-loop structure. RsmY and RsmZ regulate RsmA, and possibly RsmF, by sequestering these proteins from target mRNAs. In this study, we used selective 2′-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP) chemistry to determine the secondary structures of RsmY and RsmZ and functional assays to characterize the contribution of each GGA site to RsmY/RsmZ activity. Our data indicate that RsmA has two preferential binding sites on RsmY and RsmZ, while RsmF has one preferential binding site on RsmY and two sites on RsmZ. Despite RsmF and RsmA sharing a common consensus site, RsmF binding properties are more restrictive than those of RsmA.
IMPORTANCE
CsrA homologs are present in many bacteria. The opportunistic pathogen
Pseudomonas aeruginosa
uses RsmA and RsmF to inversely regulate factors associated with acute and chronic virulence phenotypes. RsmA has an affinity for RsmY and RsmZ higher than that of RsmF. The goal of this study was to understand the differential binding properties of RsmA and RsmF by using the RsmY and RsmZ regulatory small RNAs (sRNAs) as a model. Mutagenesis of the predicted RsmA/RsmF binding sites on RsmY and RsmZ revealed similarities in the sites required to control RsmA and RsmF activity
in vivo
. Whereas binding by RsmA was relatively tolerant of binding site mutations, RsmF was sensitive to disruption to all but two of the sites, further demonstrating that the requirements for RsmF binding activity
in vivo
and
in vitro
are more stringent than those for RsmA.
Details
- Title: Subtitle
- Functional Analyses of the RsmY and RsmZ Small Noncoding Regulatory RNAs in Pseudomonas aeruginosa
- Creators
- Kayley H Janssen - Department of Microbiology and Immunology, University of Iowa, Iowa City, Iowa, USAManisha R Diaz - Department of Microbiology and Immunology, University of Iowa, Iowa City, Iowa, USAMatthew Golden - Department of Microbiology and Immunology, University of Iowa, Iowa City, Iowa, USAJustin W Graham - Marsico Lung Institute, Cystic Fibrosis Research Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USAWes Sanders - Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USAMatthew C Wolfgang - Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USATimothy L Yahr - Department of Microbiology and Immunology, University of Iowa, Iowa City, Iowa, USA
- Resource Type
- Journal article
- Publication Details
- Journal of bacteriology, Vol.200(11), e00736-17
- Publisher
- American Society for Microbiology; 1752 N St., N.W., Washington, DC
- DOI
- 10.1128/JB.00736-17
- PMID
- 29463606
- PMCID
- PMC5952390
- ISSN
- 0021-9193
- eISSN
- 1098-5530
- Grant note
- AI097264 / ; T32GM082729, 5T32AI007511-19 / ;
- Alternative title
- RsmY and RsmZ Noncoding RNAs
- Language
- English
- Date published
- 06/01/2018
- Academic Unit
- Microbiology and Immunology
- Record Identifier
- 9984001141202771
Metrics
19 Record Views