Expression of lysostaphin in HeLa cells protects from host cell killing by intracellular Staphylococcus aureus

Abstract

The Staphylococcus aureus-specific cell wall endopeptidase lysostaphin was used as a model for an intracellular acting bactericidal antibiotic. HeLa cells were transfected with an expression vector directing the heterologous expression of lysostaphin in the cytoplasm. Expression, subcellular localization and enzymatic activity of lysostaphin were investigated by immunoblotting, fluorescent microscopy and agar diffusion assays. Both transiently and stably transfected HeLa cells showed a strong expression of active lysostaphin. After infection with S. aureus, the intracellular number of S. aureus and the host cell viability were determined. This staphylolytic activity resulted in a strong reduction of intracellular S. aureus in a time- and dose-dependent manner. Furthermore, host cells expressing lysostaphin became protected from S. aureus-induced cell death. Our data demonstrate the potential of intracellularly acting cell-wall active drugs or antibiotics that kill S. aureus without causing harm to the infected host cells.

In vitro endothelial cell damage is positively correlated with enhanced virulence and poor vancomycin responsiveness in experimental endocarditis due to methicillin-resistant Staphylococcus aureus

“…The RNAIII effector molecule of the agr locus, which encodes δ-toxin, has been reported to be a key factor in S. aureus -induced cytotoxicity (Shompole et al , 2003). δ-toxin has been shown to possess cytolytic properties (reviewed in (Verdon et al , 2009)), and has been suggested to mediate phagosomal escape by S. aureus (Giese et al , 2011, Shompole et al , 2003), a proposed prerequisite for induction of host cell death (Klein et al , 2006). We found that agr null mutants caused approximately 20% less EC damage than did their respective parental strains.…”

Staphylococcus aureus host cell invasion and post-invasion events

“…CFT-1, LCFSN RN6390 IF (Jarry and Cheung, 2006) Reduced proportion of LAMP-positive phagosomes in CFT-1 at 2 and 4 h, as opposed to LCFSN HeLa ATCC29213 Lysostaphin expression in host cells (Klein et al, 2006) Reduced whereas multiple virulence factors are down-regulated. When investigating phagolysosomal localisation of S. aureus by IF for lysosomal markers such as LAMP-1, LAMP-2, etc., ring-like structures can be detected surrounding the staphylococci.…”

“…It has been suggested as early as 1998 that S. aureus escapes the phagosomal compartment in non-professional phagocytes (Bayles et al, 1998). Phagosomal escape of S. aureus seems to be a prerequisite for induction of host cell death, since the glycyl-glycyl endopeptidase lysostaphin expressed in the cytoplasm of HeLa cells protected HeLa from induction of host cell death by internalized S. aureus ATCC29213 (Klein et al, 2006) and since only invasive and haemolytic strains induce apoptosis in human endothelial cells (Haslinger-Löffler et al, 2005).…”