<scp><i>S</i></scp><i>taphylococcus aureus</i> nitric oxide synthase (sa<scp>NOS</scp>) modulates aerobic respiratory metabolism and cell physiology

Mogen, Austin B.; Carroll, Rachel; James, Kimberly L.; Lima, G. S. C. C.; Silva, Dona; Culver, Jeffrey A.; Petucci, Christopher; Shaw, Lindsey N.; Rice, Kelly C.

doi:10.1111/mmi.13693

Cited by 32 publications

(48 citation statements)

References 119 publications

(169 reference statements)

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“…For cell pellets, an additional tube was used for cell quantification and protein content determination using a Bradford assay with Bio-Rad protein concentration reagent. Samples were processed using LC-MS/MS, as previously described (51). The adenylate energy charge was calculated as (ATP ϩ 0.5ADP)/(AMP ϩ ATP ϩ ADP), as previously described (37).…”

Section: Methodsmentioning

confidence: 99%

Redirection of Metabolism in Response to Fatty Acid Kinase in Staphylococcus aureus

DeMars

Bose

2018

J Bacteriol

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is capable of phosphorylating exogenous fatty acids via the fatty acid kinase FakA for incorporation into the bacterium's membrane. Additionally, FakA plays a significant role in virulence factor regulation and skin infections. We previously showed that a mutant displays altered growth kinetics observed during late-exponential phase of growth. Here, we demonstrate that the absence of FakA leads to key metabolic changes. First, the mutant has an altered acetate metabolism with acetate being consumed at an increased rate than the wild-type strain. Moreover, the growth benefit was diminished with inactivation of the acetate-generating enzyme AckA. Using a mass spectrometry-based approach, we identified altered concentrations of TCA cycle intermediates and both intracellular and extracellular amino acids. Together, these data demonstrate a change in carbohydrate carbon utilization and altered amino acid metabolism in the mutant. Energy status analysis revealed the mutant had a similar ADP/ATP ratio, but reduced adenylate energy charge. The inactivation of changed the NAD+/NADH and NADP+/NADPH ratio, indicating a more oxidized cellular environment. Evidence points towards the global metabolic regulatory proteins CcpA and CodY being important contributors for the altered growth in a mutant. Indeed, it was found that directing amino acids from the urea cycle into the TCA cycle via glutamate dehydrogenase was an essential component of growth after glucose depletion. Together, this data identifies a previously unidentified role for FakA in the global physiology of that links external fatty acid utilization and central metabolism. The fatty acid kinase, FakA, of plays several important roles in the cell. FakA is important for the activation of the SaeRS two-component system and secreted virulence factors like α-hemolysin. However, the contribution of FakA to cellular metabolism has not been explored. Here, we highlight the metabolic consequence of removal of FakA from the cell. The absence of FakA leads to altered acetate metabolism, altered redox balance, as well as a change in intracellular amino acids. Additionally, the use of environmental amino acids sources is affected by FakA. Together, these results demonstrate for the first time that FakA provides a link between the pathways for exogenous fatty acid use, virulence factor regulation, and other metabolic processes.

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Section: Methodsmentioning

confidence: 99%

Redirection of Metabolism in Response to Fatty Acid Kinase in Staphylococcus aureus

DeMars

Bose

2018

J Bacteriol

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“…Targeted and global metabolomics processing was performed by the Southeast Center for Integrated Metabolomics at the University of Florida. Bacterial cell pellets from acute NO stress cultures were lyophilized overnight and processed as previously described (40). Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to quantitate metabolites from cell homogenates and extracellular media.…”

Section: Methodsmentioning

confidence: 99%

“…For the extraction of nucleotides, cell homogenate was spiked with isotopically labeled internal standard and centrifuged, and supernatants were filtered through an AcroPrep Advance 3K Omega filter plate (Pall Corporation) prior to LC-MS/MS analysis. Derivatized organic acids and nucleotides were quantitated as previously described (40). For all classes of metabolites, data for cell samples were normalized to the protein, whereas supernatant concentrations are reported in micromolar units.…”

Section: Methodsmentioning

confidence: 99%

“…In addition, all S. aureus strains produce endogenous NO via oxidation of L-arginine to L-citrulline by a nitric oxide synthase (Nos) (33)(34)(35)(36)(37)(38). Endogenous NO provides protection against oxidative damage by interfering with Fenton chemistry (39), promotes aerobic respiration (40,41), and mediates the switch to nitrate-based respiration during microaerobic growth (42).…”

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confidence: 99%

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The Response of nor and nos Contributes to Staphylococcus aureus Virulence and Metabolism

et al. 2019

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Staphylococcus aureus causes a wide spectrum of disease, with the site and severity of infection dependent on virulence traits encoded within genetically distinct clonal complexes (CCs) and bacterial responses to host innate immunity. The production of nitric oxide (NO) by activated phagocytes is a major host response to which S. aureus metabolically adapts through multiple strategies that are conserved in all CCs, including an S. aureus nitric oxide synthase (Nos). Previous genome analysis of CC30, a lineage associated with chronic endocardial and osteoarticular infections, revealed a putative NO reductase (Nor) not found in other CCs that potentially contributes to NO resistance and clinical outcome. Here, we demonstrate that Nor has true nitric oxide reductase activity, with nor expression enhanced by NO stress and anaerobic growth. Furthermore, we demonstrate that nor is regulated by MgrA and SrrAB, which modulate S. aureus virulence and hypoxic response. Transcriptome analysis of the S. aureus UAMS-1, UAMS-1 Δnor, and UAMS-1 Δnos strains under NO stress and anaerobic growth demonstrates that Nor contributes to nucleotide metabolism and Nos to glycolysis. We demonstrate that Nor and Nos contribute to enhanced survival in the presence of human human polymorphonuclear cells and have organ-specific seeding in a tail vein infection model. Nor contributes to abscess formation in an osteological implant model. We also demonstrate that Nor has a role in S. aureus metabolism and virulence. The regulation overlap between Nor and Nos points to an intriguing link between regulation of intracellular NO, metabolic adaptation, and persistence in the CC30 lineage. IMPORTANCE Staphylococcus aureus can cause disease at most body sites, and illness spans asymptomatic infection to death. The variety of clinical presentations is due to the diversity of strains, which are grouped into distinct clonal complexes (CCs) based on genetic differences. The ability of S. aureus CC30 to cause chronic infections relies on its ability to evade the oxidative/nitrosative defenses of the immune system and survive under different environmental conditions, including differences in oxygen and nitric oxide concentrations. The significance of this work is the exploration of unique genes involved in resisting NO stress and anoxia. A better understanding of the functions that control the response of S. aureus CC30 to NO and oxygen will guide the treatment of severe disease presentations.

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“…NO can also block cysteine oxidation by S -nitrosylating exposed cysteines (46) and activating catalases that reduce the concentration of harmful H 2 O 2 (18). Overall, SaNOS mutants display increased endogenous ROS and superoxide levels (47). Recent studies also find that the production of NO by S. aureus is essential for nasal colonization and skin abscess development, and mutants lacking the enzyme show decreased virulence (48-50).…”

Section: Introductionmentioning

confidence: 99%

The bacillithiol pathway is required for biofilm formation in Staphylococcus aureus

Gulati

Rawat

et al. 2018

Preprint

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word count: 239Abstract 24 Staphylococcus aureus is a major human pathogen that can cause infections that range from 25 superficial skin and mucosal infections to life threatening disseminated infections. S. aureus can 26 attach to medical devices and host tissues and form biofilms that allow the bacteria to evade the 27 host immune system and provide protection from antimicrobial agents. To counter host-generated 28 oxidative and nitrosative stress mechanisms that are part of the normal host responses to invading 29 pathogens, S. aureus utilizes low molecular weight (LMW) thiols, such as bacillithiol (BSH). 30 Additionally, S. aureus synthesizes its own nitric oxide (NO), which combined with its 31 downstream metabolites may also protect the bacteria against specific host responses. We have 32 previously shown that LMW thiols are required for biofilm formation in Mycobacterium 33 smegmatis and Pseudomonas aeruginosa. Our data show that the bshC mutant, which is defective 34 in the last step of the bacillithiol pathway and lacks BSH, is impaired in biofilm formation. We 35 also identify a putative S-nitrosobacillithiol reductase (BSNOR), similar to a S-nitrosomycothiol 36 reductase found in M. smegmatis, and show that the BSNOR mutant has reduced levels of BSH 37 and decreased biofilm formation. Our studies also show that NO plays an important role in biofilm 38 formation and that acidified sodium nitrite severely reduces biofilm thickness. These studies 39 provide insight into the roles of oxidative and nitrosative stress mechanisms on biofilm formation 40 and indicate that bacillithiol and nitric oxide are key players in normal biofilm formation in S. 41 aureus. 42 43 Importance 44 Staphylococcus aureus is the most frequent cause of biofilm-associated infections in hospital 45 settings. The biofilm mode of growth allows the pathogen to escape the host immune response and 46 3is extremely difficult to combat, as biofilms are highly resistant to physical and chemical stressors. 47As outbreaks of antibiotic-resistant bacterial strains become more commonplace, it is essential to 48 understand the pathways involved in biofilm formation in order to target this important virulence 49 factor. Low molecular weight thiols enable S. aureus to combat oxidative and nitrosative stress 50 mechanisms that are used by host cells to defend against infection. Our findings indicate that 51 bacillithiol and nitric oxide, which are produced by S. aureus to combat these host generated 52 stressors, are important for biofilm development, and that disruption of these pathways results in 53 biofilm defects. In the long term, this work may lead to new solutions to eradicate S. aureus 54 biofilms in the clinic. 55 56 Introduction 57 Staphylococcus aureus is a gram-positive, spherical-shaped bacteria that is a major human 58 pathogen capable of causing both superficial as well as life-threatening systemic and chronic 59infections in humans (1, 2). S. aureus is also a commensal microorganism that can reside on the 60 skin, and with...

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Staphylococcus aureus nitric oxide synthase (saNOS) modulates aerobic respiratory metabolism and cell physiology

Cited by 32 publications

References 119 publications

Redirection of Metabolism in Response to Fatty Acid Kinase in Staphylococcus aureus

Redirection of Metabolism in Response to Fatty Acid Kinase in Staphylococcus aureus

The Response of nor and nos Contributes to Staphylococcus aureus Virulence and Metabolism

The bacillithiol pathway is required for biofilm formation in Staphylococcus aureus

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