Abstract
ABSTRACTThe alternative sigma factor σ54 has been shown to regulate the expression of a wide array of virulence-associated genes, as well as central metabolism, in bacterial pathogens. In Gram-positive organisms, the σ54 is commonly associated with carbon metabolism. In this study, we show that the Enterococcus faecalis alternative sigma factor σ54 (RpoN) and its cognate enhancer binding protein MptR are essential for mannose utilization and are primary contributors to glucose uptake through the Mpt phosphotransferase system. To gain further insight into how RpoN contributes to global transcriptional changes, we performed microarray transcriptional analysis of strain V583 and an isogenic rpoN mutant grown in a chemically defined medium with glucose as the sole carbon source. Transcripts of 340 genes were differentially affected in the rpoN mutant; the predicted functions of these genes mainly related to nutrient acquisition. These differentially expressed genes included those with predicted catabolite-responsive element (cre) sites, consistent with loss of repression by the major carbon catabolite repressor CcpA. To determine if the inability to efficiently metabolize glucose/mannose affected infection outcome, we utilized two distinct infection models. We found that the rpoN mutant is significantly attenuated in both rabbit endocarditis and murine catheter-associated urinary tract infection (CAUTI). Here, we examined a ccpA mutant in the CAUTI model and showed that the absence of carbon catabolite control also significantly attenuates bacterial tissue burden in this model. Our data highlight the contribution of central carbon metabolism to growth of E. faecalis at various sites of infection.
Highlights
The alternative sigma factor s 54 has been shown to regulate the expression of a wide array of virulence-associated genes, as well as central metabolism, in bacterial pathogens
As RpoN requires a bacterial enhancer binding protein (bEBP) for open complex formation in order for transcription to proceed, we evaluated the contribution of the four bEBPs in E. faecalis V583 by constructing deletion mutants for lpoR, mphR, mpoR, and mptR
It is of note that we identified a three-gene operon, which encodes the Lpt phosphotransferase systems (PTS) complex that was downregulated in the rpoN mutant that contained a cre site, indicating that this operon may be regulated by catabolite control protein A (CcpA) for expression
Summary
The alternative sigma factor s 54 has been shown to regulate the expression of a wide array of virulence-associated genes, as well as central metabolism, in bacterial pathogens. Transcripts of 340 genes were differentially affected in the rpoN mutant; the predicted functions of these genes mainly related to nutrient acquisition These differentially expressed genes included those with predicted catabolite-responsive element (cre) sites, consistent with loss of repression by the major carbon catabolite repressor CcpA. We found that the rpoN mutant is significantly attenuated in both rabbit endocarditis and murine catheter-associated urinary tract infection (CAUTI). Enterococci are the second leading cause of hospital-acquired infections They form biofilms at surgical sites and are often associated with infections of the urinary tract following catheterization. Urinary tract infections (CAUTI) [1] Both disease manifestations for endocarditis and CAUTI are thought to be biofilm mediated, and enterococci rank as the second leading cause of CAUTI in U.S hospitals [1]. While several studies have examined transcriptional profiles of enterococci grown in urine, serum, and abscesses [3,4,5], little is known of preferred nutrients and how E. faecalis acquires them in the host
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