Analysis of Transcription of the Staphylococcus aureus Aerobic Class Ib and Anaerobic Class III Ribonucleotide Reductase Genes in Response to Oxygen

Abstract

Staphylococcus aureus is a gram-positive facultative aerobe that can grow in the absence of oxygen by fermentation or by using an alternative electron acceptor. To investigate the mechanism by which S. aureus is able to adapt to changes in oxygen concentration, we analyzed the transcriptional regulation of genes that encode the aerobic class Ib and anaerobic class III ribonucleotide reductase (RNR) systems that are responsible for the synthesis of deoxyribonucleotides needed for DNA synthesis. The S. aureus class Ib RNR nrdIEF and class III RNR nrdDG genes and their regulatory regions were cloned and sequenced. Inactivation of the nrdDG genes showed that the class III RNR is essential for anaerobic growth. Inhibition of aerobic growth by hydroxyurea showed that the class Ib RNR is an oxygen-dependent enzyme. Northern blot analysis and primer extension analysis demonstrated that transcription of class III nrdDG genes is regulated by oxygen concentration and was at least 10-fold higher under anaerobic than under aerobic conditions. In contrast, no significant effect of oxygen concentration was found on the transcription of class Ib nrdIEF genes. Disruption or deletion of S. aureus nrdDG genes caused up to a fivefold increase in nrdDG and nrdIEF transcription under anaerobic conditions but not under aerobic conditions. Similarly, hydroxyurea, an inhibitor of the class I RNRs, resulted in increased transcription of class Ib and class III RNR genes under aerobic conditions. These findings establish that transcription of class Ib and class III RNR genes is upregulated under conditions that cause the depletion of deoxyribonucleotide. Promoter analysis of class Ib and class III RNR operons identified several inverted-repeat elements that may account for the transcriptional response of the nrdIEF and nrdDG genes to oxygen.Staphylococcus aureus is a gram-positive facultative aerobe and a major human pathogen (33,39). In common with other facultative aerobes, S. aureus can grow in the absence of oxygen either by fermentation or by using an alternative terminal electron acceptor, such as nitrate. Several studies suggest that oxygen plays a role in the pathogenesis of S. aureus, in both its capacity to produce virulence factors and its ability to persist and grow in different and often hostile environmental niches (5,6,26,37,40,53). The ability of S. aureus to adapt to extreme changes in external oxygen concentration implies the existence of one or more oxygen-sensing systems that regulate the expression of genes in the transition from aerobic to anaerobic growth. While considerable progress has been made in characterizing global regulators of anaerobic gene expression, for example, the FNR and Arc families of proteins (3, 14, 17, 42), relatively little is known about the regulatory systems that operate in S. aureus during anaerobiosis. Knowledge of these signal transduction systems is crucial for understanding how, in S. aureus, oxygen brings about changes in the expression of virulence genes. Several recent in vitro studie...

Alternative oxygen‐dependent and oxygen‐independent ribonucleotide reductases in Streptomyces: cross‐regulation and physiological role in response to oxygen limitation

“…Quantification of S. coelicolor nrd transcripts in total RNA was performed by Northern blot analysis as described (Masalha et al, 2001). RNA (5 mg) was electrophoresed in an agarose/ formaldehyde gel and transferred to NytranN nylon membranes (Schleicher & Schuell).…”

“…In some cases the rationale for having more than one RNR system is evident. In facultative aerobes such as Staphylococcus aureus , a class Ib RNR is used exclusively for aerobic growth whereas a class III RNR is limited to anaerobic growth (Masalha et al ., 2001). In other bacteria, such as Pseudomonas aeruginosa , which contains genes coding for all three kinds of RNR, the situation is more complex (Jordan et al ., 1999).…”

Corynebacterium ammoniagenes class Ib ribonucleotide reductase: transcriptional regulation of an atypical genomic organization in the nrd cluster

“…Staphylococcus epidermidis also has two copies of the nrdEF genes, one of these with the nrdI-nrdE-nrdF structure and the other one, as in B. subtilis of phage origin, with the nrdI-nrdE-nrdF structure. Staphylococcus aureus shows the nrdI-nrdE-nrdF structure (Masalha et al, 2001). Streptococcus mutans and Streptococcus pneumoniae show the nrdH-nrdE-nrdF structure plus an nrdI gene located elsewhere.…”

“…Recent studies demonstrated a potential role for the class Ib enzyme under some special growth conditions (Masalha et al, 2001;Monje-Casa et al, 2001). A distinguishing feature of all known class Ib operons is that they code for a IP: 34.217.195.47 On: Fri, 11 May 2018 14:02:28 small protein, NrdI, of unknown function but seemingly essential for cell growth (I. Sala, unpublished data), and often for a specific external electron donor (NrdH).…”

Effects of iron nano-particle’s on expression of tetracycline resistance encoding genes in Staphylococcus aureus by Real Time-PCR

“…The biochemical analysis showed that it is often positive for catalase and nitrate reduction and is a facultative anaerobe that can grow without the need for oxygen (Masalha et al, 2001). Pathogenic strains often promote infections by producing virulence factors such as potent protein toxins, and the expression of cell-surface proteins that bind and inactivate antibodies.…”