Here we identified a population of bone marrow neutrophils that constitutively express RORγt and which can produce and respond to IL-17A (IL-17). IL-6, IL-23 and RORγt, but not T cells or NK cells, are required for IL-17 production in neutrophils. IL-6 and IL-23 induced IL-17RC and Dectin-2 expression in neutrophils, and expression of IL-17RC was augmented by Aspergillus and Dectin-2 activation. Autocrine IL-17A–IL-17 receptor activity induced production of reactive oxygen species (ROS), and increased fungal killing in vitro and in a model of Aspergillus keratitis. Human neutrophils also expressed RORγt, and induced IL-17A, IL-17RC and Dectin-2 expression following IL-6 and IL-23 stimulation. These findings identify a population of human and murine neutrophils that exhibit autocrine IL-17 activity, and which likely contribute to the etiology of microbial and inflammatory diseases.

P. aeruginosa is a leading cause of blinding corneal ulcers worldwide. To determine the role of type III secretion in the pathogenesis of P. aeruginosa keratitis, corneas of C57BL/6 mice were infected with P. aeruginosa strain PAO1 or PAK, which express ExoS, ExoT and ExoY, but not ExoU. PAO1 and PAK infected corneas developed severe disease with pronounced opacification and rapid bacterial growth. In contrast, corneas infected with ΔpscD or ΔpscJ mutants that cannot assemble a Type III secretion system, or with mutants lacking the translocator proteins, do not develop clinical disease, and are rapidly killed by infiltrating neutrophils. Further, survival of PAO1 and PAK strains in the cornea and development of corneal disease was impaired in ΔexoS, ΔexoT and ΔexoST mutants of both strains, but not in a ΔexoY mutant. ΔexoST mutants were also rapidly killed in neutrophils in vitro and were impaired in their ability to promote neutrophil apoptosis in vivo compared with PAO1. Point mutations in the ADP ribosyltransferase (ADPR) regions of ExoS or ExoT also impaired pro-apoptotic activity in infected neutrophils, and exoST(ADPR-) mutants replicated the ΔexoST phenotype in vitro and in vivo, whereas mutations in rho-GAP showed the same phenotype as PAO1. Together, these findings demonstrate that the pathogenesis of P. aeruginosa keratitis in ExoS and ExoT producing strains is almost entirely due to their ADPR activities, which subvert the host response by targeting the anti-bacterial activity of infiltrating neutrophils.

SUMMARYFungal infection stimulates the canonical C-type lectin receptors (CLRs) signaling pathway via Syk activation. Here we show that SHP-2 plays a crucial role in mediating CLRs-induced Syk activation. Genetic ablation of Shp-2 (Ptpn11) in dendritic cells (DCs) and macrophages impaired Syk-mediated signaling and abrogated pro-inflammatory gene expression following fungal stimulation. Mechanistically, SHP-2 operates as a scaffold facilitating the recruitment of Syk to dectin-1 or FcRγ, through its N-SH2 domain and a previously unrecognized C-terminal ITAM motif. We demonstrate that DC-derived SHP-2 is crucial for the induction of IL-1β, IL-6 and IL-23, and anti-fungal TH17 cell responses to control Candida albicans infection. Together, these data reveal a mechanism by which SHP-2 mediates Syk activation in response to fungal infections