LC3-Associated Phagocytosis Is Required for Dendritic Cell Inflammatory Cytokine Response to Gut Commensal Yeast Saccharomyces cerevisiae

Abstract

The human fungal microbiota known as mycobiota is increasingly recognized as a critical factor in human gut health and disease. Non-pathogenic commensal yeasts such as Saccharomyces cerevisiae promote homeostasis in the gut, whereas dysbiosis of the gut mycobiota is associated with inflammation. Glycan-binding receptors (lectins) are key host factors in host–mycobiota interaction in the gut. They are expressed on immune cells such as dendritic cells (DCs) and recognize fungal polysaccharides. This interaction is imperative to mount appropriate immune responses for immune homeostasis in the gut as well as clearance of fungal pathogens. Recent studies demonstrate that microtubule-associated protein light-chain 3 (LC3)-associated phagocytosis (LAP) is involved in lectin–fungi interactions. Yet, the biological impact of LAP on the lectin function remains largely elusive. In this report, we demonstrate that in mouse LAP is linked to dendritic cell-associated lectin 2 (Dectin-2), a C-type lectin specific to fungal α-mannan polysaccharide. We found that mouse Dectin-2 recognizes commensal yeast S. cerevisiae and Kazachstania unispora. Mouse bone marrow-derived DCs (BMDCs) produced inflammatory cytokines TNFα and IL-1β in response to the yeasts in a Dectin-2 and spleen tyrosine kinase (Syk)-dependent manner. We found that S. cerevisiae and K. unispora induced LAP in mouse BMDCs upon internalization. Furthermore, LC3 was activated by stimulation of BMDCs with the yeasts in a Dectin-2 and Syk-dependent manner. To address the biological impact of LAP on Dectin-2 yeast interaction, we established a knock-in mouse strain (Atg16L1E230, thereafter called E230), which BMDCs exhibit autophagy-active and LAP-negative phenotypes. When stimulated with yeasts, E230 BMDCs produced significantly less amounts of TNFα and IL-1β. Taken together, we revealed a novel link between Dectin-2 and LAP that enables host immune cells to respond to mycobiota.

The ATG5-binding and coiled coil domains of ATG16L1 maintain autophagy and tissue homeostasis in mice independently of the WD domain required for LC3-associated phagocytosis

“…reported previously [22,23] these observations indicate that the WD domain is required for LAP in myeloid cells, and confirmed that the atg16l1 E230 mouse would provide a LAPdeficient mouse model to study the role played by LAP in maintaining tissue homeostasis in vivo.…”

“…These include newly formed macropinosomes, phagosomes containing latex beads or apoptotic corpses, endosomes swollen by monensin or a combination of ammonium chloride and the vacuolating toxin A from Helicobacter pylori, and endosomes targeted by the M2 proton channel encoded by influenza virus. Furthermore, loss of the WD domain from ATG16L1 in dendritic cells results in reduced secretion of TNF and IL1B in response to CLEC4N/Dectin2 signalling [23] and reduced antigen presentation by dendritic cells [22]. The requirement of the WD domain for LAP suggests that the scaffold provided by the WD domains of ATG16L1 has evolved a specialized role, independent of autophagy, to ensure the quality control of endocytic pathways by conjugating LC3 to phagosomes containing pathogens or apoptotic cells, or endocytic compartments showing signs of damage.…”

The WD and linker domains of ATG16L1 required for non-canonical autophagy limit lethal respiratory infection by influenza A virus at epithelial surfaces

“…Recent work shows that autophagy and phagocytosis can be linked by non-canonical autophagy. This is best characterized in 10 phagocytic cells where LC3 associated phagocytosis (LAP) is activated by Toll-like receptor signaling resulting in recruitment of autophagy marker protein LC3 to the phagosome membrane to enhance phagosome maturation [1][2][3][4][5] . In non-phagocytes a similar non-canonical pathway recruits LC3 to endo-lysosome compartments during the uptake of particulate material such as apoptotic cells and aggregated β-amyloid and 15 following membrane damage during pathogen entry or osmotic imbalance induced by lysosomotropic drugs 6,7 8-10 .…”

The Role of ATG16 in Autophagy and The Ubiquitin Proteasome System

“…The WD40 repeat domain is dispensable for canonical autophagy but is required for other processes [93]. Its deletion inhibited MHC class II antigen presentation in mouse dendritic cells during influenza A virus infection [79].…”