Pathogen activation of innate immune pattern recognition receptors (PRRs) such as Toll-like receptors (TLRs) stimulates cellular signaling pathways. This often leads to outcomes that contribute to pathogen clearance. Alternatively, activation of specific PRR pathways can aid pathogen survival. The human pathogen Staphylococcus aureus is a case in point, employing strategies to escape innate immune recognition and killing by the host. As for other bacteria, PRR-stimulated type I interferon (IFN-I) induction has been proposed as one such immune escape pathway that may favor S. aureus. Cell wall components of S. aureus elicit TLR2-dependent cellular responses, but the exact signaling pathways activated by S. aureus–TLR2 engagement and the consequences of their activation for the host and bacterium are not fully known. We previously showed that TLR2 activates both a cytoplasmic and an endosome-dependent signaling pathway, the latter leading to IFN-I production. Here, we demonstrate that S. aureus infection of human monocytes activates a TLR2-dependent endosomal signaling pathway, leading to IFN-I induction. We mapped the signaling components of this pathway and identified roles in IFN-I stimulation for the Toll-interleukin-1 receptor (TIR) adaptor Myd88 adaptor-like (Mal), TNF receptor-associated factor 6 (TRAF6), and IκB kinase (IKK)-related kinases, but not for TRIF-related adaptor molecule (TRAM) and TRAF3. Importantly, monocyte TLR2-dependent endosomal signaling enabled immune escape for S. aureus, because this pathway, but not IFN-I per se, contributed to intracellular bacterial survival. These results reveal a TLR2-dependent mechanism in human monocytes whereby S. aureus manipulates innate immune signaling for its survival in cells.

Methicillin-resistant Staphylococcus aureus (MRSA) has been classified as a high priority pathogen by the World Health Organization underlining the high demand for new therapeutics to treat infections. Human group IIA secreted phospholipase A2 (hGIIA) is among the most potent bactericidal proteins against Gram-positive bacteria, including S. aureus. To determine hGIIA-resistance mechanisms of MRSA we screened the Nebraska Transposon Mutant Library using a sublethal concentration of recombinant hGIIA. We identified and confirmed the role of lspA, encoding the lipoprotein signal peptidase LspA, as a new hGIIA resistance gene in both in vitro assays and an infection model in hGIIA-transgenic mice. Increased susceptibility of the lspA mutant was associated with faster and increased cell wall penetration of hGIIA. Moreover, lspA deletion also increased susceptibility to daptomycin, a last-resort antibiotic to treat MRSA infections. Exposure of MRSA wild-type to the LspA-specific inhibitors globomycin and myxovirescin A1 induced a lspA mutant phenotype with regard to hGIIA and daptomycin killing. Analysis of >26,000 S. aureus genomes showed that LspA is highly sequence-conserved, suggesting that LspA inhibition could be applied universally. The role of LspA in hGIIA resistance was not restricted to MRSA since Streptococcus mutans and Enterococcus faecalis were also more hGIIA-susceptible after lspA deletion or LspA inhibition, respectively. Overall, our data suggest that pharmacological blocking of LspA may disarm Gram-positive pathogens, including MRSA, to enhance clearance by innate host defense molecules and clinically-applied antibiotics.