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

Surmann K, Michalik S, Hildebrandt P, Gierok P, Depke M, Brinkmann L, Bernhardt J, Salazar MG, Sun Z, Shteynberg D, Kusebauch U, Moritz RL, Wollscheid B, Lalk M, Völker U, Schmidt F - Front Microbiol (2014)

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: Interfaculty Institute for Genetics and Functional Genomics, University Medicine Greifswald Greifswald, Germany.

ABSTRACT
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 × 10(6) 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.

No MeSH data available.


Related in: MedlinePlus

Voronoi treemap analysis of S. aureus proteins. Ratios from intensity values 2.5 h and 6.5 h p.i. compared to the non-adherent control are depicted. All pictures represent data on protein level clustered by pathways. Panels (A,E) serve as a legend showing only the pathways. Data at 2.5 h p.i. are represented in panel (B) (A549 cells), panel (C) (S9 cells), and panel (D) (HEK 293 cells). Data at 6.5 h p.i. are shown in panel (F) (A549 cells), panel (G) (S9 cells), and panel (H) (HEK 293 cells). Blue spots indicate lower levels in the internalized bacteria compared to the non-adherent control; red colors represent higher levels of proteins in response to internalization compared to the non-adherent control. Average values from three independent biological samples are displayed.
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Figure 4: Voronoi treemap analysis of S. aureus proteins. Ratios from intensity values 2.5 h and 6.5 h p.i. compared to the non-adherent control are depicted. All pictures represent data on protein level clustered by pathways. Panels (A,E) serve as a legend showing only the pathways. Data at 2.5 h p.i. are represented in panel (B) (A549 cells), panel (C) (S9 cells), and panel (D) (HEK 293 cells). Data at 6.5 h p.i. are shown in panel (F) (A549 cells), panel (G) (S9 cells), and panel (H) (HEK 293 cells). Blue spots indicate lower levels in the internalized bacteria compared to the non-adherent control; red colors represent higher levels of proteins in response to internalization compared to the non-adherent control. Average values from three independent biological samples are displayed.

Mentions: In order to gain a first inside into the global response of S. aureus to internalization by three different host cell lines a principal component analysis (PCA) plot of three sampling points (non-adherent control, 2.5 h and 6.5 h p.i.) was generated (Figure 3). For this analysis, the median-normalized data of all quantified proteins were taken into account. All samples from internalized bacteria, both at 2.5 h and 6.5 h p.i., were clearly separated from the non-adherent controls by the first component. Remarkably, the protein pattern of S. aureus internalized by HEK 293 kidney cells for 6.5 h was clearly separated in the second component from the other two S. aureus samples internalized by epithelial cell lines for the same time period, which indicates a different behavior of S. aureus inside the HEK 293 cells. The Voronoi treemaps displayed in Figure 4 provide a protein-resolved picture of the proteome adaptation where proteins which are assigned to the same biochemical pathway or adaptation reaction are grouped together (legends in Figures 4A,E). Again the patterns of S. aureus internalized by epithelial cells (S9 or A459) look more similar to each other than to HEK293-internalized S. aureus. Exemplarily one can focus on protein biosynthesis which appears more reduced in level in comparison to the control in S. aureus originating from A549 (Figure 4B) and S9 (Figure 4C) compared to those isolated 2.5 p.i. from HEK 293 cells (Figure 4D). On the contrary, proteins involved in central carbon metabolism tend to be in general more strongly induced in S. aureus from A549 (Figure 4F) and S9 (Figure 4G) cells than in S. aureus from HEK 293 cells 6.5 h p.i. (Figure 4H).


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

Surmann K, Michalik S, Hildebrandt P, Gierok P, Depke M, Brinkmann L, Bernhardt J, Salazar MG, Sun Z, Shteynberg D, Kusebauch U, Moritz RL, Wollscheid B, Lalk M, Völker U, Schmidt F - Front Microbiol (2014)

Voronoi treemap analysis of S. aureus proteins. Ratios from intensity values 2.5 h and 6.5 h p.i. compared to the non-adherent control are depicted. All pictures represent data on protein level clustered by pathways. Panels (A,E) serve as a legend showing only the pathways. Data at 2.5 h p.i. are represented in panel (B) (A549 cells), panel (C) (S9 cells), and panel (D) (HEK 293 cells). Data at 6.5 h p.i. are shown in panel (F) (A549 cells), panel (G) (S9 cells), and panel (H) (HEK 293 cells). Blue spots indicate lower levels in the internalized bacteria compared to the non-adherent control; red colors represent higher levels of proteins in response to internalization compared to the non-adherent control. Average values from three independent biological samples are displayed.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4117987&req=5

Figure 4: Voronoi treemap analysis of S. aureus proteins. Ratios from intensity values 2.5 h and 6.5 h p.i. compared to the non-adherent control are depicted. All pictures represent data on protein level clustered by pathways. Panels (A,E) serve as a legend showing only the pathways. Data at 2.5 h p.i. are represented in panel (B) (A549 cells), panel (C) (S9 cells), and panel (D) (HEK 293 cells). Data at 6.5 h p.i. are shown in panel (F) (A549 cells), panel (G) (S9 cells), and panel (H) (HEK 293 cells). Blue spots indicate lower levels in the internalized bacteria compared to the non-adherent control; red colors represent higher levels of proteins in response to internalization compared to the non-adherent control. Average values from three independent biological samples are displayed.
Mentions: In order to gain a first inside into the global response of S. aureus to internalization by three different host cell lines a principal component analysis (PCA) plot of three sampling points (non-adherent control, 2.5 h and 6.5 h p.i.) was generated (Figure 3). For this analysis, the median-normalized data of all quantified proteins were taken into account. All samples from internalized bacteria, both at 2.5 h and 6.5 h p.i., were clearly separated from the non-adherent controls by the first component. Remarkably, the protein pattern of S. aureus internalized by HEK 293 kidney cells for 6.5 h was clearly separated in the second component from the other two S. aureus samples internalized by epithelial cell lines for the same time period, which indicates a different behavior of S. aureus inside the HEK 293 cells. The Voronoi treemaps displayed in Figure 4 provide a protein-resolved picture of the proteome adaptation where proteins which are assigned to the same biochemical pathway or adaptation reaction are grouped together (legends in Figures 4A,E). Again the patterns of S. aureus internalized by epithelial cells (S9 or A459) look more similar to each other than to HEK293-internalized S. aureus. Exemplarily one can focus on protein biosynthesis which appears more reduced in level in comparison to the control in S. aureus originating from A549 (Figure 4B) and S9 (Figure 4C) compared to those isolated 2.5 p.i. from HEK 293 cells (Figure 4D). On the contrary, proteins involved in central carbon metabolism tend to be in general more strongly induced in S. aureus from A549 (Figure 4F) and S9 (Figure 4G) cells than in S. aureus from HEK 293 cells 6.5 h p.i. (Figure 4H).

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: Interfaculty Institute for Genetics and Functional Genomics, University Medicine Greifswald Greifswald, Germany.

ABSTRACT
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 × 10(6) 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.

No MeSH data available.


Related in: MedlinePlus