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Loss of rpoE Encoding the δ-Factor of RNA Polymerase Impacts Pathophysiology of the Streptococcus pyogenes M1T1
Joseph S Rom1, Yoann Le Breton1, Emrul Islam1
1Department of Cell Biology and Molecular Genetics, Maryland Pathogen Research Institute, University of Maryland, College Park, MD 20742, USA.
Abstract:
Streptococcus pyogenes, also known as the Group A Streptococcus (GAS), is a Gram-positive bacterial pathogen of major clinical significance. Despite remaining relatively susceptible to conventional antimicrobial therapeutics, GAS still causes millions of infections and hundreds of thousands of deaths each year worldwide. Thus, a need for prophylactic and therapeutic interventions for GAS is in great demand. In this study, we investigated the importance of the gene encoding the delta (δ) subunit of the GAS RNA polymerase, rpoE, for its impact on virulence during skin and soft-tissue infection. A defined 5448 mutant with an insertionally-inactivated rpoE gene was defective for survival in whole human blood and was attenuated for both disseminated lethality and lesion size upon mono-culture infection in mouse soft tissue. Furthermore, the mutant had reduced competitive fitness when co-infected with wild type (WT) 5448 in the mouse model. We were unable to attribute this attenuation to any observable growth defect, although colony size and the ability to grow at higher temperatures were both affected when grown with nutrient-rich THY media. RNA-seq of GAS grown in THY to late log phase found that mutation of rpoE significantly impacted (>2-fold) the expression of 429 total genes (205 upregulated, 224 downregulated), including multiple virulence and “housekeeping” genes. The arc operon encoding the arginine deiminase (ADI) pathway was the most upregulated in the rpoE mutant and this could be confirmed phenotypically. Taken together, these findings demonstrate that the delta (δ) subunit of RNA polymerase is vital in GAS gene expression and virulence.
Insights
The delta (δ) subunit of Streptococcus pyogenes RNA polymerase, encoded by rpoE, is crucial for bacterial virulence. Disrupting rpoE impairs GAS survival, reduces lethality in mice, and alters gene expression, highlighting its role in pathogenesis.
Area of Science:
- Microbiology
- Bacterial Pathogenesis
- Molecular Biology
Background:
- Streptococcus pyogenes (Group A Streptococcus, GAS) is a significant bacterial pathogen causing millions of infections globally.
- Despite susceptibility to antibiotics, GAS infections remain a major cause of mortality, necessitating new interventions.
- The delta (δ) subunit of GAS RNA polymerase is a potential target for understanding and combating GAS virulence.
Purpose of the Study:
- To investigate the role of the rpoE gene, encoding the delta (δ) subunit of GAS RNA polymerase, in bacterial virulence.
- To assess the impact of rpoE inactivation on GAS survival, lethality, and infection progression in a mouse model.
- To analyze global gene expression changes in response to rpoE mutation.
Main Methods:
- Construction of a defined rpoE insertion mutant (5448) in Streptococcus pyogenes.
- Assessment of mutant survival in whole human blood and virulence in a mouse skin and soft-tissue infection model.
- RNA sequencing (RNA-seq) to analyze global gene expression changes in the rpoE mutant compared to wild type.
Main Results:
- The rpoE mutant showed defective survival in human blood and was attenuated in disseminated lethality and lesion size in mice.
- The mutant exhibited reduced competitive fitness when co-infected with wild-type GAS.
- RNA-seq revealed significant differential expression of 429 genes, including upregulation of the arginine deiminase (arc) operon, in the rpoE mutant.
Conclusions:
- The delta (δ) subunit of RNA polymerase (rpoE) is essential for Streptococcus pyogenes virulence.
- rpoE plays a critical role in regulating GAS gene expression, impacting both virulence and housekeeping genes.
- Targeting the rpoE gene or its product could offer novel strategies for GAS infection control.
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