Hope the following abstract will give the lead:

Regulation of Superoxide Dismutase (sod) Genes by SarA in Staphylococcus aureus▿†

Anand Ballal and Adhar C. Manna*, Journal of Bacteriology, Accepted manuscript posted online 13 March 2009, doi:10.1128/JB.01496-08J. Bacteriol. May 2009 vol. 191no. 10 3301-3310

(http://jb.asm.org/content/191/10/3301.full)

The scavenging of reactive oxygen species (ROS) within cells is regulated by several interacting factors, including transcriptional regulators. Involvement ofsarA family genes in the regulation of proteins involved in the scavenging of ROS is largely unknown. In this report, we show that under aerobic conditions, the levels of sodM and sodA transcription, in particular the sodM transcript, are markedly enhanced in the sarA mutant among the tested sarA family mutants. Increased levels of sod expression returned to near the parental level in a single-copy sarAcomplemented strain. Under microaerophilc conditions, transcription of both sodMand sodA was considerably enhanced in the sarA mutant compared to the wild-type strain. Various genotypic, phenotypic, and DNA binding studies confirmed the involvement of SarA in the regulation of sod transcripts in different strains ofStaphylococcus aureus. The sodA mutant was sensitive to an oxidative stress-inducing agent, methyl viologen, but the sarA sodA double mutant was more resistant to the same stressor than the single sodA mutant. These results suggest that overexpression of SodM, which occurs in the sarA background, can rescue the methyl viologen-sensitive phenotype observed in the absence of the sodA gene. Analysis with various oxidative stress-inducing agents indicates that SarA may play a greater role in modulating oxidative stress resistance in S. aureus. This is the first report that demonstrates the direct involvement of a regulatory protein (SarA) in control of sod expression in S. aureus.

Staphylococcus aureus is a major human pathogen that has colonized more than one-third of the world's population. It causes a variety of infections in immunocompromised hosts as well as healthy individuals. Most S. aureusinfections begin as minor colonization of soft tissue or skin, from which the organism can spread to the bloodstream and subsequently disseminate into various tissues. Once inside tissues, S. aureus produces a large number of factors, including adhesins, enzymes, toxins, capsular polysaccharides, etc., which facilitate tissue colonization, tissue destruction, and immune evasion. The expression of many of these determinants is coordinately controlled by regulatory systems, such as agr, saeRS, vraRS, tcaRA, sarA, and nine other sarA paralogs (4,6-10, 17, 24, 27, 30, 32-36, 39, 42, 52).

The sarA locus, an important transcriptional regulatory system, comprises a major open reading frame, sarA, driven by three distinct promoters, resulting in three overlapping transcripts with a common 3′ end (1). Inactivation of the sarA locus upregulates fibronectin and fibrinogen binding protein synthesis, hemolysins (α-, β-, and δ-hemolysins), enterotoxins, toxic shock syndrome toxin 1 toxin 1, genes involved in biofilm formation (e.g., icaRA and bap), and capsule and downregulates proteases, protein A, and collagen binding protein synthesis (3, 6, 10, 53). The SarA protein has also been shown to bind to several regulatory loci (e.g., agr, sarS,rot, sarV, and sarT) to modulate target gene transcription directly and indirectly (8,9, 34, 37, 45, 46). Transcriptional profiling studies showed that mutation of thesarA gene led to altered expression of roughly 120 genes (76 upregulated and 44 downregulated) (13). Using a combination of different techniques and genome sequence information, an additional nine SarA paralogs (SarR to -V, SarX, SarZ, Rot, and MgrA) have been identified. These SarA paralogs have been shown to regulate expression of target genes, including those involved in virulence, regulation, biofilm formation, autolysis, antibiotic resistance, and metabolic processes (3, 8, 13, 24, 30, 32-37, 39, 42, 46, 52).

Reactive oxygen species (ROS), such as superoxide anion (O2−), hydrogen peroxide (H2O2), and hydroxyl radicals (OH), are generated as the by-products of cellular metabolism and have been implicated in different human diseases. ROS are also important for normal cellular processes such as apoptosis, cell proliferation and differentiation, and transcriptional regulation (41, 43). They are also involved in bacterial and viral killing by neutrophils and other phagocyte cells (2, 40). In bacteria, during aerobic growth, ROS arise as intracellular natural products from incomplete reduction of oxygen during respiration and are highly toxic to nucleic acids, proteins, and cell membrane fatty acids (23, 38). In most cases, the harmful ROS are sensed directly by transcriptional regulators, which upon oxidation increase expression of genes involved in the detoxification of ROS (19, 41). In S. aureus four main iron-dependent global regulators (Fur, PerR, Zur, and MntR) are known to be involved in resistance to oxidative stress (20, 22, 28,57). Among these, PerR predominantly protects S. aureus cells against oxidative stress challenge mediated by H2O2. The genes katA (catalase), ahpCF(alkylhydroperoxide reductase), mrgA (Dps-like protein), bcp (bacterioferitin coregulatory protein), and trxB (thioredoxin reductase) are members of the PerR regulons that are activated due to perR deletion (12, 20). Another set of enzymes, superoxide dismutases (SODs), are responsible for converting superoxides to hydrogen peroxide and oxygen. Thereafter, catalase converts hydrogen peroxide to water and oxygen.

In several bacterial species, SOD has been shown to be important for defense against killing by the phagocytes of vertebrate hosts. Inactivation of the sod genes in Shigella flexneri and Escherichia coli K-12 results in enhanced sensitivity to killing by serum and neutrophils (15, 38). The sod gene inactivation inStreptococcus pneumoniae, Campylobacter coli, and Haemophilus influenzaeresults in attenuation of virulence, reduced colonization of the chicken stomach, decreased survival in the spleen or liver of mice, and the inability to colonize the rat nasopharynx (56). In S. aureus, there are conflicting reports regarding the roles of SODs in virulence and staphylococcal disease. An earlier study suggested no correlation between SOD activity and lethality in a mouse model of infection (31). An isogenic sodA or sodM mutant of strain 8325-4 showed no effect on virulence in a mouse abscess model (11, 47); however, SOD activity was found to be significantly higher in S. aureus strains isolated from patients with staphylococcal disease (25). It has also been reported that isogenic sodA, sodM, and sodA sodMmutants of SH1000, an rsbU-complemented derivative of 8325-4, had reduced virulence compared to the wild-type strain in a mouse abscess model of infection (26). Therefore, these results suggest that enzymatic superoxide anion scavenging by SODs is important for the survival of the pathogen.

 S. aureus has two SOD genes, sodA and sodM (11, 55). In Staphylococcus, SodA is the major SOD enzyme, and in fact, coagulase-negative staphylococci such asStaphylococcus epidermidis lack the sodM gene, suggesting that SodM may have a unique role in S. aureus (56). The regulation of the sod genes in S. aureus is not well understood. Therefore, we explored the possible regulatory role played by staphylococcus-specific sarA family genes in regulation of the sod genes in S. aureus. In this study, we report that among the SarA family, SarA is involved in negative regulation of both sodM and sodA genes. SarA specifically binds to thesodA and sodM promoter regions, thus suggesting a direct mode of regulation. Analysis of various single and double sod mutants with sarA in the presence or absence of different oxidative stress-responsive agents indicates that SarA may play an important role in modulating oxidative stress resistance in S. aureus.

Dr. Mirza Arshad Ali Beg

Many thanks, it's really useful.