Molecular mechanisms of LC3-associated phagocytosis in the macrophage response toParacoccidioidesspp

Abstract

Paracoccidiomycosis is a systemic fungal infection that is endemic in Latin America. The etiologic agents are thermodimorphic fungi from the Paracoccidiodes genus, which are facultative intracellular parasites of macrophages. LC3-associated phagocytosis (LAP), a noncanonical form of autophagy, is important in the immune response to similar pathogens, so we sought to determine the role LAP plays in the macrophage response to Paracoccidioides spp. By immunofluorescence, we found that LC3 was recruited to phagosomes containing Paracoccidioides spp. in both RAW264.7 and J774.16 cell lines and in bone marrow-derived macrophages. Interference with autophagy using RNAi against ATG5 reduced the antifungal activity of J774.16 cells, showing that LC3 recruitment is important for proper control of the fungus by macrophages. Finally, we used pharmacological Syk kinase and NAPH oxidase inhibitors, which inhibit signaling pathways necessary for macrophage LAP against Aspergillus fumigatus and Candida albicans, to dissect part of the signaling pathways that trigger LAP against Paracoccidioides spp. Interestingly, these inhibitors did not decrease LAP against P. brasiliensis, possibly due to differences in the fungal cell surface compositions. These observations suggest a potential role for autophagy as target for host-directed paracoccidioidomycosis therapies.