Host protection from infection relies on the recognition of pathogens by innate pattern-recognition receptors such as Toll-like receptors (TLRs). Here, we show that the orphan receptor TLR13 in mice recognizes a conserved 23S ribosomal RNA (rRNA) sequence that is the binding site of macrolide, lincosamide, and streptogramin group (MLS) antibiotics (including erythromycin) in bacteria. Notably, 23S rRNA from clinical isolates of erythromycin-resistant Staphylococcus aureus and synthetic oligoribonucleotides carrying methylated adenosine or a guanosine mimicking a MLS resistance-causing modification failed to stimulate TLR13. Thus, our results reveal both a natural TLR13 ligand and specific mechanisms of antibiotic resistance as potent bacterial immune evasion strategy, avoiding recognition via TLR13.

The mammalian target of rapamycin (mTOR) can be viewed as cellular master complex scoring cellular vitality and stress. Whether mTOR controls also innate immune-defenses is currently unknown. Here we demonstrate that TLR activate mTOR via phosphoinositide 3-kinase/Akt. mTOR physically associates with the MyD88 scaffold protein to allow activation of interferon regulatory factor-5 and interferon regulatory factor-7, known as master transcription factors for pro-inflammatory cytokine-and type I IFN-genes. Unexpectedly, inactivation of mTOR did not prevent but increased lethality of endotoxinmediated shock, which correlated with increased levels of IL-1b. Mechanistically, mTOR suppresses caspase-1 activation, thus inhibits release of bioactive IL-1b. We have identified mTOR as indispensable component of PRR signal pathways, which orchestrates the defense program of innate immune cells.Key words: Caspase-1 . IRF . mTOR . TLR Supporting Information available online IntroductionThe phosphoinositide 3-kinase (PI3K) represents a signaling gateway for the activation of various cellular effector functions including cell growth, proliferation, survival and vesicular transport [1,2]. Activated PI3K catalyzes the phosphorylation of membrane-anchored phosphoinositides (PI) and binding of PI-3,4,5-tri-phosphate to both Akt and PI-dependent protein kinase 1, which then drives PI-dependent protein kinase 1 to activate Akt via Thr308 phosphorylation [1]. It is known that inhibition of PI3K interferes with functions of innate immune cells [3,4], yet the molecular basis for this is still unclear.Upon inhibition of Akt, TLR-activated macrophages and DC mimic the phenotype of TLR-stimulated PI3K deficient cells [5]. Therefore, we reasoned that PI3K executes its regulatory function along the Akt pathway. One of the major targets of Akt is the mammalian target of rapamycin (mTOR), known to influence multiple cellular functions including cell cycle control, cellular growth, apoptosis, transcription and translational efficacy [6,7]. Whether and how mTOR signaling becomes integrated into TLR signaling pathways is unknown. Eur. J. Immunol. 2008. 38: 2981-2992 DOI 10.1002 HIGHLIGHTS 2981 FrontlineHere we describe that mTOR signaling is indispensable for the signal pathways of various PRR. First we show that membrane bound TLR directly activate mTOR via the PI3K/Akt axis. Activated mTOR subsequently transcriptionally controls in innate immune cells cytokine and type I IFN production. Essential steps in this transcriptional process include recruitment of activated mTOR to the MyD88 scaffold protein, the site at which interferon regulatory factor (IRF)-5 and IRF-7 become activated in an mTOR-dependent fashion. In addition, mTOR negatively regulates bioactive IL-1b production by inhibiting caspase-1 activation. These data characterize mTOR as transcriptional regulator and controller of acute innate immune reactions. Results TLR activate mTORTo analyze whether TLR signaling drives mTOR activation, we asked whether TLR-mediated activation of bon...

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Gram-negative bacterial infection is a major cause of sepsis and septic shock. An important inducer of infl ammation underlying both syndromes is the cellular recognition of bacterial products through pattern recognition receptors (PRRs), including Toll-like receptors (TLRs). We identifi ed a novel antagonistic mAb (named 1A6) that recognizes the extracellular portion of the TLR4 -MD-2 complex. If applied to mice before infection with clinical isolates of Salmonella enterica or Escherichia coli and subsequent antibiotic therapy, 1A6 prevented otherwise fatal shock, whereas application of 1A6 after infection was ineffective. In contrast, coapplication of 1A6 and an anti-TLR2 mAb up to 4 h after infection with Gram-negative bacteria, in combination with the start of antibiotic therapy (mimicking clinical conditions), provided robust protection. Consistent with our fi ndings in mice, dual blockade of TLR2 and TLR4 inhibited TNF-␣ release from human peripheral blood mononuclear cells upon Gram-negative bacterial infection/antibiotic therapy. Both murine splenocytes and human PBMCs released IFN-␥ in a TLR4-dependent manner, leading to enhanced surface TLR2 expression and sensitivity for TLR2 ligands. Our results implicate TLR2 as an important, TLR4-driven sensor of Gram-negative bacterial infection and provide a rationale for blockade of both TLRs, in addition to antibiotic therapy for the treatment of Gram-negative bacterial infection.