Phagocytosis‐dependent activation of a TLR 9– BTK –calcineurin– NFAT pathway co‐ordinates innate immunity to Aspergillus fumigatus

Abstract

Transplant recipients on calcineurin inhibitors are at high risk of invasive fungal infection. Understanding how calcineurin inhibitors impair fungal immunity is a key priority for defining risk of infection. Here, we show that the calcineurin inhibitor tacrolimus impairs clearance of the major mould pathogen Aspergillus fumigatus from the airway, by inhibiting macrophage inflammatory responses. This leads to defective early neutrophil recruitment and fungal clearance. We confirm these findings in zebrafish, showing an evolutionarily conserved role for calcineurin signalling in neutrophil recruitment during inflammation. We find that calcineurin– NFAT activation is phagocytosis dependent and collaborates with NF ‐κB for TNF ‐α production. For yeast zymosan particles, activation of macrophage calcineurin– NFAT occurs via the phagocytic Dectin‐1–spleen tyrosine kinase pathway, but for A. fumigatus , activation occurs via a phagosomal TLR 9‐dependent and Bruton's tyrosine kinase‐dependent signalling pathway that is independent of MyD88. We confirm the collaboration between NFAT and NF ‐κB for TNF ‐α production in primary alveolar macrophages. These observations identify inhibition of a newly discovered macrophage TLR 9– BTK –calcineurin– NFAT signalling pathway as a key immune defect that leads to organ transplant‐related invasive aspergillosis. image Whilst calcineurin inhibition is common during organ transplantation, it also increases susceptibility to life‐threatening invasive pulmonary aspergillosis. Calcineurin is found here to mediate phagocytosis‐dependent innate immune responses to Aspergillus fumigatus in macrophages. Phagocytosis‐dependent activation of TLR 9 by the human mould pathogen A. fumigatus drives calcineurin– NFAT transcriptional responses in macrophages. Activation of calcineurin– NFAT by TLR 9 is independent of MyD88 and Dectin‐1–spleen tyrosine kinase, but dependent on Bruton's tyrosine kinase. This pathway is critical for macrophage TNF ‐α production and subsequent chemotaxis of neutrophil to the airway during pulmonary aspergillosis. Direct inhibition of the TLR 9– BTK –calcineurin– NFAT pathway by calcineurin inhibitors is a key mechanism that increases clinical susceptibility to pulmonary aspergillosis.