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Functional studies of E . faecalis RNase J2 and its role in virulence and fitness

View Article: PubMed Central - PubMed

ABSTRACT

Post-transcriptional control provides bacterial pathogens a method by which they can rapidly adapt to environmental change. Dual exo- and endonucleolytic activities of RNase J enzymes contribute to Gram-positive RNA processing and decay. First discovered in Bacillus subtilis, RNase J1 plays a key role in mRNA maturation and degradation, while the function of the paralogue RNase J2 is largely unknown. Previously, we discovered that deletion of the Enterococcus faecalis rnjB gene significantly attenuates expression of a major virulence factor involved in enterococcal pathogenesis, the Ebp pili. In this work, we demonstrate that E. faecalis rnjB encodes an active RNase J2, and that the ribonuclease activity of RNase J2 is required for regulation of Ebp pili. To further investigate how rnjB affects E. faecalis gene expression on a global scale, we compared transcriptomes of the E. faecalis strain OG1RF with its isogenic rnjB deletion mutant (ΔrnjB). In addition to Ebp pili regulation, previously demonstrated to have a profound effect on the ability of E. faecalis to form biofilm or establish infection, we identified that rnjB regulates the expression of several other genes involved in bacterial virulence and fitness, including gls24 (a virulence factor important in stress response). We further demonstrated that the E. faecalis RNase J2 deletion mutant is more sensitive to bile salt and greatly attenuated in in vivo organ infection as determined by an IV-sublethal challenge infection mouse model, indicating that E. faecalis RNase J2 plays an important role in E. faecalis virulence.

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IV infection of ICR mice with E. faecalis wild type vs. ΔrnjB.The number of E. faecalis cells recovered from the kidneys and spleens 48hrs after inoculation are determined as CFU per gram of tissue. On both panels shown on left (Kidney) and right (Spleen), circles and squares represent mice infected with OG1RF (n = 16) and ΔrnjB (n = 16), respectively. Horizontal bars represent the geometric mean titers, and the difference of the CFU/gm mean value of wild type and ΔrnjB were determined by unpaired t tests.
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pone.0175212.g007: IV infection of ICR mice with E. faecalis wild type vs. ΔrnjB.The number of E. faecalis cells recovered from the kidneys and spleens 48hrs after inoculation are determined as CFU per gram of tissue. On both panels shown on left (Kidney) and right (Spleen), circles and squares represent mice infected with OG1RF (n = 16) and ΔrnjB (n = 16), respectively. Horizontal bars represent the geometric mean titers, and the difference of the CFU/gm mean value of wild type and ΔrnjB were determined by unpaired t tests.

Mentions: To evaluate the impact of RNase J2 on bacterial virulence in vivo, we utilized a sublethal challenge model in mice [29] to study the capacity of wild type vs. ΔrnjB E. faecalis to establish infection. Two rounds of independent experiments were performed and the combined results are shown in Fig 7. Mice infected with ΔrnjB (n = 16) show a 50-fold reduction in mean CFUs in the spleens (ΔrnjB 1.2 x103 vs. wild type 6.8x104CFU/gm) and a 400-fold reduction kidneys (ΔrnjB 2.2x103 vs. wild type 9.3x105CFU/gm) compared to mice infected with wild type bacteria (n = 16), demonstrating a significant decrease in the ability of the mutant to establish infection (Fig 7). While these results are consistent with the observed inhibition of virulence factor expression in ΔrnjB data presented, the described growth delay of the ΔrnjB is also a potential factor in overall attenuation. Nonetheless, in totality, these observations are further supportive evidence of rnjB playing an important role in Enterococci fitness.


Functional studies of E . faecalis RNase J2 and its role in virulence and fitness
IV infection of ICR mice with E. faecalis wild type vs. ΔrnjB.The number of E. faecalis cells recovered from the kidneys and spleens 48hrs after inoculation are determined as CFU per gram of tissue. On both panels shown on left (Kidney) and right (Spleen), circles and squares represent mice infected with OG1RF (n = 16) and ΔrnjB (n = 16), respectively. Horizontal bars represent the geometric mean titers, and the difference of the CFU/gm mean value of wild type and ΔrnjB were determined by unpaired t tests.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5383250&req=5

pone.0175212.g007: IV infection of ICR mice with E. faecalis wild type vs. ΔrnjB.The number of E. faecalis cells recovered from the kidneys and spleens 48hrs after inoculation are determined as CFU per gram of tissue. On both panels shown on left (Kidney) and right (Spleen), circles and squares represent mice infected with OG1RF (n = 16) and ΔrnjB (n = 16), respectively. Horizontal bars represent the geometric mean titers, and the difference of the CFU/gm mean value of wild type and ΔrnjB were determined by unpaired t tests.
Mentions: To evaluate the impact of RNase J2 on bacterial virulence in vivo, we utilized a sublethal challenge model in mice [29] to study the capacity of wild type vs. ΔrnjB E. faecalis to establish infection. Two rounds of independent experiments were performed and the combined results are shown in Fig 7. Mice infected with ΔrnjB (n = 16) show a 50-fold reduction in mean CFUs in the spleens (ΔrnjB 1.2 x103 vs. wild type 6.8x104CFU/gm) and a 400-fold reduction kidneys (ΔrnjB 2.2x103 vs. wild type 9.3x105CFU/gm) compared to mice infected with wild type bacteria (n = 16), demonstrating a significant decrease in the ability of the mutant to establish infection (Fig 7). While these results are consistent with the observed inhibition of virulence factor expression in ΔrnjB data presented, the described growth delay of the ΔrnjB is also a potential factor in overall attenuation. Nonetheless, in totality, these observations are further supportive evidence of rnjB playing an important role in Enterococci fitness.

View Article: PubMed Central - PubMed

ABSTRACT

Post-transcriptional control provides bacterial pathogens a method by which they can rapidly adapt to environmental change. Dual exo- and endonucleolytic activities of RNase J enzymes contribute to Gram-positive RNA processing and decay. First discovered in Bacillus subtilis, RNase J1 plays a key role in mRNA maturation and degradation, while the function of the paralogue RNase J2 is largely unknown. Previously, we discovered that deletion of the Enterococcus faecalis rnjB gene significantly attenuates expression of a major virulence factor involved in enterococcal pathogenesis, the Ebp pili. In this work, we demonstrate that E. faecalis rnjB encodes an active RNase J2, and that the ribonuclease activity of RNase J2 is required for regulation of Ebp pili. To further investigate how rnjB affects E. faecalis gene expression on a global scale, we compared transcriptomes of the E. faecalis strain OG1RF with its isogenic rnjB deletion mutant (ΔrnjB). In addition to Ebp pili regulation, previously demonstrated to have a profound effect on the ability of E. faecalis to form biofilm or establish infection, we identified that rnjB regulates the expression of several other genes involved in bacterial virulence and fitness, including gls24 (a virulence factor important in stress response). We further demonstrated that the E. faecalis RNase J2 deletion mutant is more sensitive to bile salt and greatly attenuated in in vivo organ infection as determined by an IV-sublethal challenge infection mouse model, indicating that E. faecalis RNase J2 plays an important role in E. faecalis virulence.

No MeSH data available.


Related in: MedlinePlus