Time‐resolved quantitative proteome profiling of host–pathogen interactions: The response of <i>Staphylococcus aureus</i> RN1HG to internalisation by human airway epithelial cells

Schmidt, Frank; Scharf, Sandra S.; Hildebrandt, Petra; Burian, Marc; Bernhardt, Jörg; Dhople, Vishnu M.; Kalinka, Julia; Gutjahr, Melanie; Hammer, Elke; Völker, Uwe

doi:10.1002/pmic.201000045

Cited by 47 publications

(45 citation statements)

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“…In this regard, upregulation of cell wall and techoic acid synthesis genes has been observed in S. aureus following internalization within human lung epithelial cells [35] . Furthermore, in a recent publication reporting the in vivo proteome profile of S. aureus internalized within human bronchial epithelial cells, Schmidt and co-workers [36] showed that, of the 17 potential two-component systems identified in S. aureus , only the response regulator VraR increased in level over time after internalization into host cells. The transcriptional response of S. aureus during intracellular persistence within the mast cell is currently under investigation.…”

Section: Discussionmentioning

confidence: 99%

Staphylococcus aureus Evades the Extracellular Antimicrobial Activity of Mast Cells by Promoting Its Own Uptake

et al. 2011

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In this study, we investigated the interactions of Staphylococcus aureus with mast cells, which are multifunctional sentinels lining the surfaces of the body. We found that bone marrow-derived murine mast cells (BMMC) exerted a powerful phagocytosis-independent antimicrobial activity against S. aureus. Both the release of extracellular traps as well as discharge of antimicrobial compounds were the mechanisms used by the BMMC to kill extracellular S. aureus. This was accompanied by the secretion of mediators such as TNF-α involved in the recruitment of effector cells. Interestingly, S. aureus subverted the extracellular antimicrobial activity of the BMMC by internalizing within these cells. S. aureus was also capable to internalize within human mast cells (HMC-1) and within murine skin mast cells during in vivo infection. Bacteria internalization was, at least in part, mediated by the α5β1 integrins expressed on the surface of the mast cell. In the intracellular milieu, the bacterium survived and persisted by increasing the cell wall thickness and by gaining access into the mast cell cytosol. The expression of α-hemolysin was essential for staphylococci intracellular persistence. By hiding within the long-life mast cells, staphylococci not only avoid clearance but also establish an infection reservoir that could contribute to chronic carriage.

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Section: Discussionmentioning

confidence: 99%

Staphylococcus aureus Evades the Extracellular Antimicrobial Activity of Mast Cells by Promoting Its Own Uptake

et al. 2011

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“…Proteome analysis of internalized bacteria requires enrichment by differential centrifugation, immunomagnetic separation, or fluorescent assisted cell sorting. A first analysis of the complex proteome adaptation of S. aureus to the intracellular milieu of human epithelial cells was performed in 2010 (Schmidt et al, 2010). Human bronchial epithelial S9 cells were infected with S. aureus HG001 pMV158GFP, which expresses green fluorescent protein constitutively, allowing separation of internalized S. aureus from the whole cell lysate by cell sorting prior to LC-MS/MS analysis of the bacterial proteins (Schmidt et al, 2010).…”

Section: Introductionmentioning

confidence: 99%

Comparative proteome analysis reveals conserved and specific adaptation patterns of Staphylococcus aureus after internalization by different types of human non-professional phagocytic host cells

et al. 2014

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Staphylococcus aureus is a human pathogen that can cause a wide range of diseases. Although formerly regarded as extracellular pathogen, it has been shown that S. aureus can also be internalized by host cells and persist within these cells. In the present study, we comparatively analyzed survival and physiological adaptation of S. aureus HG001 after internalization by two human lung epithelial cell lines (S9 and A549), and human embryonic kidney cells (HEK 293). Combining enrichment of bacteria from host-pathogen assays by cell sorting and quantitation of the pathogen's proteome by mass spectrometry we characterized S. aureus adaptation during the initial phase between 2.5 h and 6.5 h post-infection. Starting with about 2 × 106 bacteria, roughly 1450 S. aureus proteins, including virulence factors and metabolic enzymes were identified by spectral comparison and classical database searches. Most of the bacterial adaptation reactions, such as decreased levels of ribosomal proteins and metabolic enzymes or increased amounts of proteins involved in arginine and lysine biosynthesis, enzymes coding for terminal oxidases and stress responsive proteins or activation of the sigma factor SigB were observed after internalization into any of the three cell lines studied. However, differences were noted in central carbon metabolism including regulation of fermentation and threonine degradation. Since these differences coincided with different intracellular growth behavior, complementary profiling of the metabolome of the different non-infected host cell types was performed. This revealed similar levels of intracellular glucose but host cell specific differences in the amounts of amino acids such as glycine, threonine or glutamate. With this comparative study we provide an impression of the common and specific features of the adaptation of S. aureus HG001 to specific host cell environments as a starting point for follow-up studies with different strain isolates and regulatory mutants.

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“…Combination of peptides and concomitant MS measurement allow the relative quantification of an unlabeled to a labeled sample in one measurement, hence avoiding any variability based on slight day-to-day differences in LC-MS/MS analysis. Despite being usually applied to organisms auxotrophic for arginine/ lysine, the technique has also been used successfully for nematodes (Larance et al, 2011) and bacteria such as Staphylococcus aureus (Schmidt et al, 2010), which, although not being auxotrophic, were opportunistic enough to prefer the supplemented amino acids for peptide biosynthesis. Instead of amino acids, isotopically labeled substrates or nutrients can be applied that lead to a defined relative isotope abundance (RIA) in the biomass.…”

Section: Metabolic Labelingmentioning

confidence: 99%

Insights from quantitative metaproteomics and protein-stable isotope probing into microbial ecology

et al. 2013

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The recent development of metaproteomics has enabled the direct identification and quantification of expressed proteins from microbial communities in situ, without the need for microbial enrichment. This became possible by (1) significant increases in quality and quantity of metagenome data and by improvements of (2) accuracy and (3) sensitivity of modern mass spectrometers (MS). The identification of physiologically relevant enzymes can help to understand the role of specific species within a community or an ecological niche. Beside identification, relative and absolute quantitation is also crucial. We will review label-free and label-based methods of quantitation in MS-based proteome analysis and the contribution of quantitative proteome data to microbial ecology. Additionally, approaches of protein-based stable isotope probing (protein-SIP) for deciphering community structures are reviewed. Information on the species-specific metabolic activity can be obtained when substrates or nutrients are labeled with stable isotopes in a protein-SIP approach. The stable isotopes ( 13 C, 15 N, 36 S) are incorporated into proteins and the rate of incorporation can be used for assessing the metabolic activity of the corresponding species. We will focus on the relevance of the metabolic and phylogenetic information retrieved with protein-SIP studies and for detecting and quantifying the carbon flux within microbial consortia. Furthermore, the combination of protein-SIP with established tools in microbial ecology such as other stable isotope probing techniques are discussed.

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Time‐resolved quantitative proteome profiling of host–pathogen interactions: The response of Staphylococcus aureus RN1HG to internalisation by human airway epithelial cells

Cited by 47 publications

References 45 publications

Staphylococcus aureus Evades the Extracellular Antimicrobial Activity of Mast Cells by Promoting Its Own Uptake

Staphylococcus aureus Evades the Extracellular Antimicrobial Activity of Mast Cells by Promoting Its Own Uptake

Comparative proteome analysis reveals conserved and specific adaptation patterns of Staphylococcus aureus after internalization by different types of human non-professional phagocytic host cells

Insights from quantitative metaproteomics and protein-stable isotope probing into microbial ecology

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