Oxidation of bacillithiol during killing of Staphylococcus aureus USA300 inside neutrophil phagosomes

Targeting immune evasion tactics of pathogenic bacteria may hold the key to treating recalcitrant bacterial infections. Staphylococcus aureus produces bacillithiol (BSH), its major low-molecular-weight thiol, which is thought to protect this opportunistic human pathogen against the bombardment of oxidants inside neutrophil phagosomes. Here, we show that BSH was oxidized when human neutrophils phagocytosed S. aureus, but provided limited protection to the bacteria. We used mass spectrometry to measure the oxidation of BSH upon exposure of S. aureus USA300 to either a bolus of hypochlorous acid (HOCl) or a flux generated by the neutrophil enzyme myeloperoxidase. Oxidation of BSH and loss of bacterial viability were strongly correlated (r = 0.99, p < 0.001). BSH was fully oxidized after exposure of S. aureus to lethal doses of HOCl. However, there was no relationship between the initial BSH levels and the dose of HOCl required for bacterial killing. In contrast to the HOCl systems, only 50% of total BSH was oxidized when neutrophils killed the majority of phagocytosed bacteria. Oxidation of BSH was decreased upon inhibition of myeloperoxidase, implicating HOCl in phagosomal BSH oxidation. A BSH-deficient S. aureus USA300 mutant was slightly more susceptible to treatment with either HOCl or ammonia chloramine, or to killing within neutrophil phagosomes. Collectively, our data show that myeloperoxidase-derived oxidants react with S. aureus inside neutrophil phagosomes, leading to partial BSH oxidation, and contribute to bacterial killing. However, BSH offers only limited protection against the neutrophil's multifaceted killing mechanisms

MerA functions as a hypothiocyanous acid reductase and defense mechanism in Staphylococcus aureus

The major pathogen Staphylococcus aureus has to cope with host-derived oxidative stress to cause infections in humans. Here, we report that S. aureus tolerates high concentrations of hypothiocyanous acid (HOSCN), a key antimicrobial oxidant produced in the respiratory tract. We discovered that the flavoprotein disulfide reductase (FDR) MerA protects S. aureus from this oxidant by functioning as a HOSCN reductase, with its deletion sensitizing bacteria to HOSCN. Crystal structures of homodimeric MerA (2.4 Å) with a Cys43–Cys48 intramolecular disulfide, and reduced MerACys43S (1.6 Å) showed the FAD cofactor close to the active site, supporting that MerA functions as a group I FDR. MerA is controlled by the redox-sensitive repressor HypR, which we show to be oxidized to intermolecular disulfides under HOSCN stress, resulting in its inactivation and derepression of merA transcription to promote HOSCN tolerance. Our study highlights the HOSCN tolerance of S. aureus and characterizes the structure and function of MerA as a major HOSCN defense mechanism. Crippling the capacity to respond to HOSCN may be a novel strategy for treating S. aureus infections

MerA functions as a hypothiocyanous acid reductase and defense mechanism in Staphylococcus aureus

The major pathogen Staphylococcus aureus has to cope with host-derived oxidative stress to cause infections in humans. Here, we report that S. aureus tolerates high concentrations of hypothiocyanous acid (HOSCN), a key antimicrobial oxidant produced in the respiratory tract. We discovered that the flavoprotein disulfide reductase (FDR) MerA protects S. aureus from this oxidant by functioning as a HOSCN reductase, with its deletion sensitizing bacteria to HOSCN. Crystal structures of homodimeric MerA (2.4 Å) with a Cys$_{43}$–Cys$_{48}$ intramolecular disulfide, and reduced MerACys$_{43}$S (1.6 Å) showed the FAD cofactor close to the active site, supporting that MerA functions as a group I FDR. MerA is controlled by the redox-sensitive repressor HypR, which we show to be oxidized to intermolecular disulfides under HOSCN stress, resulting in its inactivation and derepression of merA transcription to promote HOSCN tolerance. Our study highlights the HOSCN tolerance of S. aureus and characterizes the structure and function of MerA as a major HOSCN defense mechanism. Crippling the capacity to respond to HOSCN may be a novel strategy for treating S. aureus infections

Clinical relevance of host immunity in breast cancer: from TILs to the clinic.

The clinical relevance of the host immune system in breast cancer has long been unexplored. Studies developed over the past decade have highlighted the biological heterogeneity of breast cancer, prompting researchers to investigate whether the role of the immune system in this malignancy is similar across different molecular subtypes of the disease. The presence of high levels of lymphocytic infiltration has been consistently associated with a more-favourable prognosis in patients with early stage triple-negative and HER2-positive breast cancer. These infiltrates seem to reflect favourable host antitumour immune responses, suggesting that immune activation is important for improving survival outcomes. In this Review, we discuss the composition of the immune infiltrates observed in breast cancers, as well as data supporting the clinical relevance of host antitumour immunity, as represented by lymphocytic infiltration, and how this biomarker could be used in the clinical setting. We also discuss the rationale for enhancing immunity in breast cancer, including early data on the efficacy of T-cell checkpoint inhibition in this setting.SCOPUS: re.jinfo:eu-repo/semantics/publishe

Toward Functional Carboxylate-Bridged Diiron Protein Mimics: Achieving Stability and Conformational Flexibility Using a Macrocylic Ligand

A dinucleating macrocycle, H[subscript 2]PIM, containing phenoxylimine metal-binding units has been prepared. Reaction of H[subscript 2]PIM with [Fe[subscript 2](Mes)[subscript 4]] (Mes = 2,4,6-trimethylphenyl) and sterically hindered carboxylic acids, Ph[subscript 3]CCO[subscript 2]H or ArTolCO[subscript 2]H (2,6-bis(p-tolyl)benzoic acid), afforded complexes [Fe[subscript 2](PIM)(Ph[subscript 3]CCO[subscript 2])[subscript 2]] (1) and [Fe[subscript 2](PIM)(Ar[subscript Tol]CO[subscript 2])[subscript 2]] (2), respectively. X-ray diffraction studies revealed that these diiron(II) complexes closely mimic the active site structures of the hydroxylase components of bacterial multicomponent monooxygenases (BMMs), particularly the syn disposition of the nitrogen donor atoms and the bridging μ-η[superscript 1]η[superscript 2] and μ-η[superscript 1]η[superscript 1] modes of the carboxylate ligands at the diiron(II) centers. Cyclic voltammograms of 1 and 2 displayed quasi-reversible redox couples at +16 and +108 mV vs ferrocene/ferrocenium, respectively. Treatment of 2 with silver perchlorate afforded a silver(I)/iron(III) heterodimetallic complex, [Fe[subscript 2](μ-OH)[subscript 2](ClO[subscript 4])[subscript 2](PIM)(Ar[superscript Tol]CO[subscript 2])Ag] (3), which was structurally and spectroscopically characterized. Complexes 1 and 2 both react rapidly with dioxygen. Oxygenation of 1 afforded a (μ-hydroxo)diiron(III) complex [Fe[subscript 2](μ-OH)(PIM)(Ph[subscript 3]CCO[subscript 2])[subscript 3]] (4), a hexa(μ-hydroxo)tetrairon(III) complex [Fe[subscript 4](μ-OH)[subscript 6](PIM)[subscript 2](Ph[subscript 3]CCO[subscript 2])[subscript 2]] (5), and an unidentified iron(III) species. Oxygenation of 2 exclusively formed di(carboxylato)diiron(III) compounds, a testimony to the role of the macrocylic ligand in preserving the dinuclear iron center under oxidizing conditions. X-ray crystallographic and [superscript 57]Fe Mössbauer spectroscopic investigations indicated that 2 reacts with dioxygen to give a mixture of (μ-oxo)diiron(III) [Fe[subscript 2](μ-O)(PIM)(Ar[superscript Tol]CO[subscript 2])[subscript 2]] (6) and di(μ-hydroxo)diiron(III) [Fe[subscript 2](μ-OH)[subscript 2](PIM)(Ar[superscript Tol]CO[subscript 2])[subscript 2]] (7) units in the same crystal lattice. Compounds 6 and 7 spontaneously convert to a tetrairon(III) complex, [Fe[subscript 4](μ-OH)[subscript 6](PIM)[subscript 2](Ar[superscript Tol]CO[subscript 2])[subscript 2]] (8), when treated with excess H[subscript 2]O.National Institute of General Medical Sciences (U.S.) (Grant GM032134