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Salmonella Typhimurium (S. Typhimurium) is a Gram-negative, facultative intracellular bacterium that exploits the host’s type I interferon (IFN-I) response to induce cell death in macrophages. We have previously demonstrated that activation of the IFN-I receptor (Ifnar1) results in recruitment of RIP1 and subsequent formation of a RIP1/RIP3 complex, leading to a specific form of programmed cell death, termed necroptosis, in S. Typhimurium-infected macrophages (Robinson et al. 2012). Despite our detailed knowledge on IFN-I/RIP1/RIP3-dependent necroptosis execution, the IFN-I-mediated pathways that determine whether infected macrophages will undergo necroptosis remain elusive. This work therefore sought to identify the IFN-I-mediated events that sensitize S. Typhimurium-infected macrophages to cell death. Here, we demonstrate that S. Typhimurium infection causes mitochondrial damage and impairs the host´s anti-oxidative stress response through upregulation of the mitochondrial phosphatase Pgam5 downstream of IFN-I/RIP3. Pgam5 subsequently interacts with the transcription factor Nrf2, which sequesters Nrf2 in the cytosol thereby repressing the transcription of Nrf2-dependent anti-oxidative genes. The impaired ability to respond to S. Typhimurium-induced oxidative stress results in ROS-mediated mitochondrial damage, ATP depletion, transient induction of autophagy, and autophagy-mediated degradation of p62, which impairs p62-Keap1 interaction. Consequently, Keap1 interacts more with Nrf2, which further represses Nrf2 function and additionally impairs anti-oxidative stress responses to S. Typhimurium infection thereby sensitizing macrophages to cell death. Taken together, we identify impaired Nrf2-dependent redox homeostasis of S. Typhimurium-infected macrophages as an important mechanism that drives cell death downstream of IFN-I/RIP3.

International audience; We performed a multicentre retrospective cohort study including 606,649 acute inpatient episodes at 10 European hospitals in 2010 and 2011 to estimate the impact of antimicrobial resistance on hospital mortality, excess length of stay (LOS) and cost. Bloodstream infections (BSI) caused by third-generation cephalosporin-resistant Enterobacteriaceae (3GCRE), meticillin-susceptible (MSSA) and -resistant Staphylococcus aureus (MRSA) increased the daily risk of hospital death (adjusted hazard ratio (HR) = 1.80; 95% confidence interval (CI): 1.34-2.42, HR = 1.81; 95% CI: 1.49-2.20 and HR = 2.42; 95% CI: 1.66-3.51, respectively) and prolonged LOS (9.3 days; 95% CI: 9.2-9.4, 11.5 days; 95% CI: 11.5-11.6 and 13.3 days; 95% CI: 13.2-13.4, respectively). BSI with third-generation cephalosporin-susceptible Enterobacteriaceae (3GCSE) significantly increased LOS (5.9 days; 95% CI: 5.8-5.9) but not hazard of death (1.16; 95% CI: 0.98-1.36). 3GCRE significantly increased the hazard of death (1.63; 95% CI: 1.13-2.35), excess LOS (4.9 days; 95% CI: 1.1-8.7) and cost compared with susceptible strains, whereas meticillin resistance did not. The annual cost of 3GCRE BSI was higher than of MRSA BSI. While BSI with S. aureus had greater impact on mortality, excess LOS and cost than Enterobacteriaceae per infection, the impact of antimicrobial resistance was greater for Enterobacteriaceae.

Salmonella enterica serovar Typhimurium (S. Typhimurium) is a Gram-negative bacterium that induces cell death of macrophages as a key virulence strategy. We have previously demonstrated that the induction of macrophage death is dependent on the host’s type I IFN (IFN-I) response. IFN-I signaling has been shown to induce tripartite motif (TRIM) 21, an E3 ubiquitin ligase with critical functions in autoimmune disease and antiviral immunity. However, the importance and regulation of TRIM21 during bacterial infection remains poorly understood. In this study, we investigated the role of TRIM21 upon S. Typhimurium infection of murine bone marrow–derived macrophages. Although Trim21 expression was induced in an IFN-I–dependent manner, we found that TRIM21 levels were mainly regulated posttranscriptionally. Following TLR4 activation, TRIM21 was transiently degraded via the lysosomal pathway by chaperone-mediated autophagy (CMA). However, S. Typhimurium–induced mTORC2 signaling led to phosphorylation of Akt at S473, which subsequently impaired TRIM21 degradation by attenuating CMA. Elevated TRIM21 levels promoted macrophage death associated with reduced transcription of NF erythroid 2–related factor 2 (NRF2)–dependent antioxidative genes. Collectively, our results identify IFN-I–inducible TRIM21 as a negative regulator of innate immune responses to S. Typhimurium and a previously unrecognized substrate of CMA. To our knowledge, this is the first study reporting that a member of the TRIM family is degraded by the lysosomal pathway.

Significance In the present study, we identify and describe an important cross-talk between leptin signaling and macrophage functions in the context of Salmonella Typhimurium infection. Genetic ablation of leptin receptor or pharmacological antagonization of leptin augmented lysosomal functions in macrophages, reduced S. Typhimurium burden, and diminished inflammation both in vitro and in vivo. Leptin signaling activates mTORC2/Akt pathway through the down-regulation of Phlpp1 phosphatase, thus impairs lysosome-mediated pathogen clearance., The dynamic interplay between metabolism and immune responses in health and disease, by which different immune cells impact on metabolic processes, are being increasingly appreciated. However, the potential of master regulators of metabolism to control innate immunity are less understood. Here, we studied the cross-talk between leptin signaling and macrophage function in the context of bacterial infections. We found that upon infection with Gram-negative pathogens, such as Salmonella Typhimurium, leptin receptor (Lepr) expression increased in both mouse and human macrophages. Unexpectedly, both genetic Lepr ablation in macrophages and global pharmacologic leptin antagonization augmented lysosomal functions, reduced S. Typhimurium burden, and diminished inflammation in vitro and in vivo. Mechanistically, we show that leptin induction activates the mTORC2/Akt pathway and subsequently down-regulates Phlpp1 phosphatase, allowing for phosphorylated Akt to impair lysosomal-mediated pathogen clearance. These data highlight a link between leptin signaling, the mTORC2/Phlpp1/Akt axis, and lysosomal activity in macrophages and have important therapeutic implications for modulating innate immunity to combat Gram-negative bacterial infections.

During intracellular infections, autophagy significantly contributes to the elimination of pathogens, regulation of pro-inflammatory signaling, secretion of immune mediators and in coordinating the adaptive immune system. Intracellular pathogens such as S. Typhimurium have evolved mechanisms to circumvent autophagy. However, the regulatory mechanisms targeted by S. Typhimurium to modulate autophagy have not been fully resolved. Here we report that cytosolic energy loss during S. Typhimurium infection triggers transient activation of AMPK, an important checkpoint of mTOR activity and autophagy. The activation of AMPK is regulated by LKB1 in a cytosolic complex containing Sirt1 and LKB1, where Sirt1 is required for deacetylation and subsequent activation of LKB1. S. Typhimurium infection targets Sirt1, LKB1 and AMPK to lysosomes for rapid degradation resulting in the disruption of the AMPK-mediated regulation of mTOR and autophagy. The degradation of cytosolic Sirt1/LKB1/AMPK complex was not observed with two mutant strains of S. Typhimurium, ΔssrB and ΔssaV, both compromising the pathogenicity island 2 (SPI2). The results highlight virulence factor-dependent degradation of host cell proteins as a previously unrecognized strategy of S. Typhimurium to evade autophagy., Author summary S. Typhimurium is a facultative intracellular pathogen which uses its type III secretion system to avoid cell-autonomous defense mechanisms such as autophagy. Here we show that S. Typhimurium induces energy depletion resulting in an early but transient activation of AMPK and autophagy. Salmonella virulence factors target Sirt1/LKB1/AMPK for lysosomal degradation, which enables sustained mTOR-activation and inhibition of autophagy. Activation of mTOR establishes a molecular feedback loop that enhances lysosomal degradation of Sirt1/LKB1/AMPK.

Type I interferon (IFN-I) triggers necroptosis in macrophages infected with S. Typhimurium by an unclear mechanism. Hos et al. now demonstrate that RIP3 enhances the interaction of Nrf2 with Pgam5 in response to IFN-I signaling in S. Typhimurium–infected macrophages, which abates Nrf2-dependent cytoprotective pathways and increases cell death., Salmonella enterica serovar Typhimurium exploits the host’s type I interferon (IFN-I) response to induce receptor-interacting protein (RIP) kinase–mediated necroptosis in macrophages. However, the events that drive necroptosis execution downstream of IFN-I and RIP signaling remain elusive. In this study, we demonstrate that S. Typhimurium infection causes IFN-I–mediated up-regulation of the mitochondrial phosphatase Pgam5 through RIP3. Pgam5 subsequently interacts with Nrf2, which sequesters Nrf2 in the cytosol, thereby repressing the transcription of Nrf2-dependent antioxidative genes. The impaired ability to respond to S. Typhimurium–induced oxidative stress results in reactive oxygen species–mediated mitochondrial damage, energy depletion, transient induction of autophagy, and autophagic degradation of p62. Reduced p62 levels impair interaction of p62 with Keap1, which further decreases Nrf2 function and antioxidative responses to S. Typhimurium infection, eventually leading to cell death. Collectively, we identify impaired Nrf2-dependent redox homeostasis as an important mechanism that promotes cell death downstream of IFN-I and RIP3 signaling in S. Typhimurium–infected macrophages.