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

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 × 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.

A proteomic perspective of the interplay of Staphylococcus aureus and human alveolar epithelial cells during infection

“…Intracellular survival and persistence is accompanied by a reduction of bacterial growth reflected by the decreased numbers of ribosomes. These findings confirm results of previous studies [26,57].…”

“…Further confirmation for a microaerobic environment inside host cells is provided by the switch from the main oxidase Qox to the alternative oxidase Cyd, which is preferentially used by S. aureus under microaerobic conditions [61]. This observation is supported by a previous proteome study where the bacterial adaptation was only investigated 2.5 h and 6.5 h after infection [57].…”

“…This confirms our recently published study, where the bacterial response was compared between 2.5 h and 6.5 h p.i. [36].…”

“…Extra-and intracellular metabolites of non-infected A549 cells treated with sterile infection mix were analyzed as described previously [36]. In brief, extracellular samples were analyzed from cell culture supernatant by 1 H-NMR using a Bruker AVANCE-II 600 NMR spectrometer, operated by TOPSPIN 2.1 software (Bruker Biospin GmbH, Rheinstetten, Germany).…”

Staphylococcus aureus Infection Reduces Nutrition Uptake and Nucleotide Biosynthesis in a Human Airway Epithelial Cell Line

“…Within the last two decades, however, S. aureus has also been recognized as an invasive pathogen with an intracellular lifestyle [5,6]. It was shown by proteomic and transcriptomic studies that intracellular S. aureus undergoes changes in expression of metabolic genes, nutrient transporters and virulence factors to adapt to the intracellular environment [7,8]. …”