PpL is established as a key virulence factor that triggers AP-1-mediated inflammatory signaling, structural and functional insights into PrpL inhibition are provided, and AprA is identified as a novel upstream PrpL activator, highlighting PrpL as a promising anti-virulence therapeutic target.
Abstract
The opportunistic pathogen Pseudomonas aeruginosa secretes numerous proteases that disrupt host defenses. Among them, the lysyl endopeptidase PrpL has been implicated in virulence, but its role in inflammatory responses has remained unclear. This study shows that purified PrpL activates the AP-1 transcription factors following instillation into mouse lungs, and drives robust production of IL-1β, IL-6, and TNF, as well as clinical symptoms. A proteolytically inactive mutant of PrpL fails to elicit these responses. Structural analysis of the complex of PrpL and its natural inhibitory propeptide (PrpLPP) using X-ray crystallography revealed the inhibitory mechanism of PrpLPP, which guided the identification of a pre-existing dipeptide inhibitor (LasBi) that blocked PrpL activity and restricted AP-1-driven cytokine induction and clinical symptoms in vivo. This study shows that the alkaline protease AprA degrades PrpLPP and functions redundantly with LasB to liberate mature, active PrpL. Our findings: 1) establish PrpL as a key virulence factor that triggers AP-1-mediated inflammatory signaling, 2) provide structural and functional insights into PrpL inhibition, and 3) identify AprA as a novel upstream PrpL activator. Together, these results highlight PrpL as a promising anti-virulence therapeutic target. Given the functional redundancy of inflammatory effectors produced by P. aeruginosa, strategies aimed at mitigating P. aeruginosa-induced lung inflammation through PrpL inhibition would likely need to be combined with approaches targeting additional pro-inflammatory bacterial factors. Author Summary We identify PrpL as a critical Pseudomonas aeruginosa protease that activates the inflammatory transcription factor AP-1 and drives potent lung inflammation. Structural analysis of a natural PrpL inhibitor guided development of an existing dipeptide inhibitor that significantly reduced the production of proinflammatory cytokines and clinical responses in vivo. Moreover, discovery of the AprA protease as an additional PrpL activator refines the activation model and suggests potential upstream therapeutic strategies. These findings highlight PrpL as both a driver of pathogenesis and a promising therapeutic target for treating P. aeruginosa infections.
Chronic infections by Pseudomonas aeruginosa in people with cystic fibrosis are characterized by persistent inflammation and oxidative stress, yet the mechanisms enabling bacterial persistence are not fully understood. Here, we identify a persistence mechanism mediated by pyruvate secretion resulting from mutations in the pyruvate dehydrogenase complex in clinical isolates of P. aeruginosa, with putative analogous mutations also identified in Staphylococcus aureus, Haemophilus influenzae and Stenotrophomonas maltophilia. These mutations lead to elevated extracellular pyruvate, which dampens host inflammatory responses and favors bacterial persistence. Pyruvate exerts multiple roles: scavenges reactive oxygen species such as H2O2, suppresses host immune activation both in airway epithelial cells and macrophages, and increases bacterial survival during phagocytosis. This metabolic crosstalk promotes bacterial persistence while reducing epithelial and macrophage inflammatory responses. Our findings reveal pyruvate as a bacterial immunometabolite that mimics host antioxidant defenses, reshaping the infection niche to favor long-term colonization. This work highlights the broader role of secreted metabolites in host-pathogen interactions and suggests new strategies targeting metabolic pathways to manage chronic infections. The study shows that metabolic mutations in Pseudomonas aeruginosa lead to pyruvate secretion, which reduces oxidative stress, dampens immune responses, promoting bacterial survival and persistence in chronic airway infection.
S. Lampadariou, P. Laborda, L. Masiero et al.· Nature Communications· 0 citations
A novel function of Ecb is revealed, the activation of basophils to produce IL-4 in an IgE-independent manner, and a dual Ecb contribution to immune evasion-by interfering with complement activation and skewing Th2 immunity-and a role in the development of allergic inflammation-by inducing IL-4 production is suggested.
Haruka Sakakibara, Shion Kamada, Rikuto Iwata et al.· Microbiology and immunology· 0 citations
ABSTRACT The serine/threonine protein kinase F (PknF) of Mycobacterium tuberculosis (Mtb) has poorly defined targets and functions but is involved in limiting NLRP3 inflammasome activation in murine macrophages and dendritic cells in vitro. The importance of PknF for the virulence of Mtb in vivo is not known. Here, we demonstrate that the Mtb CDC1551 deletion mutant of pknF (∆pknF) expresses significantly increased levels of the lipid pthiocerol dimycoserosate (PDIM), a polyketide lipid with pro-virulence properties. The ∆pknF mutant strain compared with the Mtb and complemented strains showed a 100-fold increase in growth at day 28 and about a 10-fold increase in growth at days 90–98 in the lungs of mice. The increase in pulmonary bacterial loads after infection with the ∆pknF strain was conserved even in Nlrp3-deficient mice, arguing that PknF modulates Mtb virulence independently of NLRP3-inflammasome activation in mice. Staining of lung sections revealed increased inflammation in the lungs of ∆pknF strain-infected mice when compared with Mtb and the complemented mutant strain. Highly susceptible B6.Sst1S mice displayed decreased host resistance, with significantly decreased survival when infected with the ∆pknF strain compared to the complemented strain or Mtb. In conclusion, our data suggest that expression of PknF, as a modulator of multiple downstream effector proteins, restricts the virulence of Mtb in the lungs of mice through an NLRP3 inflammasome-independent mechanism, but potentially via suppressing production of the virulence lipid, PDIM.
Flor Torres-Juárez, Shivangi Rastogi, David C. Young et al.· Infection and Immunity· 0 citations
IL-6 and MIP-2 played a role in PLY-induced PMN recruitment considering that this response was partially diminished in IL-66 gene deficient mice (P = 0.1 vs. wild type mice), and in mice treated with anti-MIP-2 antiserumm (P<0.05).
ABSTRACT Antibiotic resistance threatens the effectiveness of conventional antimicrobial agents, underscoring the need for host-directed therapies that enhance immune defense. Here, we present a cross-species discovery pipeline that couples a Caenorhabditis elegans–Staphylococcus aureus liquid-based infection screen with mammalian mechanistic validation to identify small-molecule immunomodulators. Using six C. elegans innate immunity reporter strains, we identified a known small molecule, SKI-II, as a previously unrecognized host-protective compound that activates the C. elegans SKN-1/Nrf2 oxidative-stress pathway. In mouse RAW264.7 macrophages, SKI-II binds to the ATPase pocket of VCP (Valosin-Containing Protein), activating the PERK-dependent (Protein kinase R [PKR]-like Endoplasmic Reticulum Kinase) Nrf2 signaling regulation axis that reduces the levels of pathogenic ROS (reactive oxygen species). SKI-II treatment also promotes macrophage M1 polarization and a mitochondrial metabolic shift. This work identifies VCP as a druggable node for host-directed immunomodulation and highlights SKI-II as a prototype small molecule that boosts host tolerance to infection, thereby validating our C. elegans-based screening platform for discovering immunomodulators active in mammalian systems. IMPORTANCE Enhancing host immunity is a promising strategy to combat S. aureus infection, particularly multidrug-resistant strains. In this study, we applied a C. elegans liquid-based infection screening model to identify the immunomodulatory compound SKI-II. We demonstrate that SKI-II protects against S. aureus infection in both nematodes and mouse macrophages by regulating host oxidative stress pathways rather than directly targeting the pathogen. We reveal a regulatory circuit in which VCP functions as a central node controlling cellular ROS responses and activating the downstream PERK-dependent Nrf2 antioxidant signaling pathway. These findings advance our understanding of host cellular responses to bacterial infection and highlight VCP as a druggable target for host-directed therapeutic strategies against infectious diseases. Enhancing host immunity is a promising strategy to combat S. aureus infection, particularly multidrug-resistant strains. In this study, we applied a C. elegans liquid-based infection screening model to identify the immunomodulatory compound SKI-II. We demonstrate that SKI-II protects against S. aureus infection in both nematodes and mouse macrophages by regulating host oxidative stress pathways rather than directly targeting the pathogen. We reveal a regulatory circuit in which VCP functions as a central node controlling cellular ROS responses and activating the downstream PERK-dependent Nrf2 antioxidant signaling pathway. These findings advance our understanding of host cellular responses to bacterial infection and highlight VCP as a druggable target for host-directed therapeutic strategies against infectious diseases.
Liyang Zhang, Kai Ye, Charilaos Dellis et al.· mBio· 0 citations
Tuft cells (TCs) act as crucial airway sentinels that detect bacterial metabolites and initiate immune responses, yet the underlying mechanisms remain poorly understood. Here, we identify tracheal TCs as the initial source of leukotrienes (LTs), released during bacterial infection. Tracheal TCs discriminate pathogenic from commensal bacteria by sensing extracellular ATP (eATP) released by pathogens, including Pseudomonas aeruginosa and Rodentibacter pneumotropicus, within 4 h of infection, through the transient receptor potential cation channel subfamily M member 5 (Trpm5). This induces the LT release, including LTB4, and promotes rapid recruitment of neutrophils and macrophages to the trachea and alveolar spaces. Trpm5-/- mice failed to detect bacterial eATP, exhibited neutrophil sequestration in the spleen, and became colonized following R. pneumotropicus infection, while Trpm5+/+ mice efficiently cleared the pathogen. These findings uncover a critical TC-dependent sensing mechanism in pneumonia, establishing TCs as both ATP sensors and triggers of acute innate immune responses.
M. I. Elhawy, N. A. Wadood, Maria Grammer et al.· Cell Reports· 0 citations
We use cookies to run the site and, with your consent, for analytics and to show ads.
See our Cookie Policy.
