The δ Subunit of RNA Polymerase Guides Promoter Selectivity and Virulence in Staphylococcus aureus

Weiss, Amy L.; Ibarra, J. Antonio; Paoletti, Jessica; Carroll, Rachel; Shaw, Lindsey N.

doi:10.1128/iai.01508-14

Cited by 34 publications

(55 citation statements)

References 51 publications

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“…Analysis of RNA-seq data from the USA300 Houston strain demonstrate that the ctpA 2 gene is monocistronic and therefore can be disrupted without causing polar effects on downstream genes (Fig. S2) (Weiss et al, 2014). The mutant generated did not exhibit a growth defect under any laboratory growth conditions tested (data not shown).…”

Section: Analysis Of Ctpa Gene Expressionmentioning

confidence: 99%

The lone S41 family C-terminal processing protease in Staphylococcus aureus is localized to the cell wall and contributes to virulence

et al. 2014

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Staphylococcus aureus is a versatile pathogen of humans and a continued public health concern due to the rise and spread of multidrug-resistant strains. As part of an ongoing investigation into the pathogenic mechanisms of this organism we previously demonstrated that an intracellular Nterminal processing protease is required for S. aureus virulence. Following on from this, here we examine the role of CtpA, the lone C-terminal processing protease of S. aureus. CtpA, a member of the S41 family, is a serine protease whose homologues in Gram-negative bacteria have been implicated in a range of biological functions, including pathogenesis. We demonstrate that S. aureus CtpA is localized to the bacterial cell wall and expression of the ctpA gene is maximal upon exposure to conditions encountered during infection. Disruption of the ctpA gene leads to decreased heat tolerance and increased sensitivity when exposed to components of the host immune system. Finally we demonstrate that the ctpA " mutant strain is attenuated for virulence in a murine model of infection. Our results represent the first characterization of a C-terminal processing protease in a pathogenic Gram-positive bacterium and show that it plays a critical role during infection.

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Section: Analysis Of Ctpa Gene Expressionmentioning

confidence: 99%

The lone S41 family C-terminal processing protease in Staphylococcus aureus is localized to the cell wall and contributes to virulence

et al. 2014

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“…Since our RNAP composition studies showed that rpoZ deletion also results in ␦ depletion from RNAP, we next assessed whether removal of ␦ from the transcription complex was the driving force behind the observed changes. This is particularly important, as our previous studies have shown that ␦ is a key factor for maintaining transcriptional specificity within the S. aureus cell (8). Upon comparing RNA sequencing-derived transcriptional changes of the rpoE and rpoZ mutants, we found an overlapping regulon of only 24 genes (Fig.…”

Section: Resultsmentioning

confidence: 91%

“…Deletion of these subunits does not result in lethality for the cell, and thus their diminutive size and nonessential nature have resulted in their being classified as the "small accessory subunits" (reviewed in reference 6). Nevertheless, for the ␦ factor it has been shown that, in various species, deletion is accompanied by a deregulation of the transcriptional process, leading to decreased fitness and impaired virulence in pathogenic organisms (7,8). While there is an extensive history of research for ␦, has only recently been described as an RNAP subunit, with only a single study thus far performed in Bacillus subtilis (5), which suggested a role in phage immunity.…”

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The ω Subunit Governs RNA Polymerase Stability and Transcriptional Specificity in Staphylococcus aureus

et al. 2017

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Staphylococcus aureus is a major human pathogen that causes infection in a wide variety of sites within the human body. Its ability to adapt to the human host and to produce a successful infection requires precise orchestration of gene expression. While DNA-dependent RNA polymerase (RNAP) is generally well characterized, the roles of several small accessory subunits within the complex have yet to be fully explored. This is particularly true for the omega ( or RpoZ) subunit, which has been extensively studied in Gram-negative bacteria but largely neglected in Grampositive counterparts. In Escherichia coli, it has been shown that ppGpp binding, and thus control of the stringent response, is facilitated by . Interestingly, key residues that facilitate ppGpp binding by are not conserved in S. aureus, and consequently, survival under starvation conditions is unaffected by rpoZ deletion. Further to this, -lacking strains of S. aureus display structural changes in the RNAP complex, which result from increased degradation and misfolding of the ␤= subunit, alterations in ␦ and factor abundance, and a general dissociation of RNAP in the absence of . Through RNA sequencing analysis we detected a variety of transcriptional changes in the rpoZ-deficient strain, presumably as a response to the negative effects of depletion on the transcription machinery. These transcriptional changes translated to an impaired ability of the rpoZ mutant to resist stress and to fully form a biofilm. Collectively, our data underline, for the first time, the importance of for RNAP stability, function, and cellular physiology in S. aureus. IMPORTANCEIn order for bacteria to adjust to changing environments, such as within the host, the transcriptional process must be tightly controlled. Transcription is carried out by DNA-dependent RNA polymerase (RNAP). In addition to its major subunits (␣ 2 ␤␤=) a fifth, smaller subunit, , is present in all forms of life. Although this small subunit is well studied in eukaryotes and Gram-negative bacteria, only limited information is available for Gram-positive and pathogenic species. In this study, we investigated the structural and functional importance of , revealing key roles in subunit folding/stability, complex assembly, and maintenance of transcriptional integrity. Collectively, our data underline, for the first time, the importance of for RNAP function and cellular harmony in S. aureus.KEYWORDS RNA polymerase subunit omega, RpoZ, Staphylococcus aureus, gene regulation T ranscription in all forms of life is a tightly controlled process, necessitated by the essentiality of correct temporal and spatial expression of genes for survival. All transcriptional activity within a cell is maintained by the DNA-dependent RNA polymerase (RNAP). This multiprotein complex is structurally and functionally similar in distant forms of life, displaying only minor variations in composition, e.g., the presence/ absence of certain subunits (1, 2). In bacteria RNAP consists of four main subunits, i.e.,

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“…This was explained by ␦-mediated changes in the requirement of iNTP by RNAP to stabilize the open complex formation at the promoters where the stability of the open complex formation is rate-limiting for transcription. Transcriptome analysis of S. aureus strains with and without ␦ show that the protein is not only involved in up-regulation of certain genes but is also involved in down-regulation of the genes that encode virulence factors (13).…”

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Bacillus subtilis δ Factor Functions as a Transcriptional Regulator by Facilitating the Open Complex Formation

Prajapati

Sengupta

Rudra

et al. 2016

Journal of Biological Chemistry

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Most bacterial RNA polymerases (RNAP) contain five conserved subunits, viz. 2␣, ␤, ␤, and . However, in many Grampositive bacteria, especially in fermicutes, RNAP is associated with an additional factor, called ␦. For over three decades since its identification, it had been thought that ␦ functioned as a subunit of RNAP to enhance the level of transcripts by recycling RNAP. In support of the previous observations, we also find that ␦ is involved in recycling of RNAP by releasing the RNA from the ternary complex. We further show that ␦ binds to RNA and is able to recycle RNAP when the length of the nascent RNA reaches a critical length. However, in this work we decipher a new function of ␦. Performing biochemical and mutational analysis, we show that Bacillus subtilis ␦ binds to DNA immediately upstream of the promoter element at A-rich sequences on the abrB and rrnB1 promoters and facilitates open complex formation. As a result, ␦ facilitates RNAP to initiate transcription in the second scale, compared with minute scale in the absence of ␦. Using transcription assay, we show that ␦-mediated recycling of RNAP cannot be the sole reason for the enhancement of transcript yield. Our observation that ␦ does not bind to RNAP holo enzyme but is required to bind to DNA upstream of the ؊35 promoter element for transcription activation suggests that ␦ functions as a transcriptional regulator.Transcription is the first step in gene regulation in bacteria in which RNA polymerase (RNAP) 3 together with different factors and transcriptional regulators control the gene expression. Bacterial RNAP core enzyme contains five conserved subunits: 2 ␣, ␤, ␤Ј, and . A specificity factor associates with RNAP core enzyme to form RNAP holo enzyme that is able to recognize and initiate transcription at promoters.In certain Gram-positive bacteria, including Bacillus subtilis and Staphylococcus aureus, an additional factor, called ␦, is associated with RNAP. The ␦ factor was first identified in 1975 during the purification of RNAP from phage (SP01)-infected B. subtilis (1). The protein copurified with RNAP, and therefore it was thought that ␦ functions as a subunit of RNAP. Attempts were made to characterize the functional role of the protein in transcription. Several reports suggested that ␦ was involved in promoter selection (2-5) and functioned together with A as an initiation subunit of RNAP (6, 7) or as an allosteric modulator of RNAP conformation in both initiation and the RNAP core recycling phase (5). Other reports showed that ␦ and A bind to RNAP core with negative cooperativity (8, 9), and ␦ has no effect on transcription initiation, the rate of elongation, or termination (5). Using in vitro transcription assays, several groups showed that ␦ enhances the production of transcripts from certain promoters. This increase in transcript yield in the presence of ␦ is attributed to the recycling of RNAP possibly by ␦-mediated release of RNAP from the elongation complex following transcription termination or by inhibiting the formation of s...

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The δ Subunit of RNA Polymerase Guides Promoter Selectivity and Virulence in Staphylococcus aureus

Cited by 34 publications

References 51 publications

The lone S41 family C-terminal processing protease in Staphylococcus aureus is localized to the cell wall and contributes to virulence

The lone S41 family C-terminal processing protease in Staphylococcus aureus is localized to the cell wall and contributes to virulence

The ω Subunit Governs RNA Polymerase Stability and Transcriptional Specificity in Staphylococcus aureus

Bacillus subtilis δ Factor Functions as a Transcriptional Regulator by Facilitating the Open Complex Formation

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