hDectin-1 Is Involved in the Uptake of Apoptotic and Infected Cells and Mediates Cross-Presentation of Engulfed Antigens.

Dectin-1 is a member of the c-type-lectin-like receptor family that was shown to be the major receptor for fungal beta-glucans and to play an important role in the cellular responses mediated by these carbohydrates. In the present study we identified the hDectin-1b splice variant as the major isoform expressed in in vitro generated mDC using a quantitative RT-PCR and western blot analysis. Interestingly, stimulation of immature DC with the toll like receptor ligands Poly I:C (TLR3) or LPS (TLR4) but not with TLR ligands 2 and 7 or TNF-a led to a dramatic down regulation of hDectin-1 protein expression. To further analyze the possible involvement of Dectin-1 in the phagocytosis of cellular material we recombinantly expressed the extracellular domain (ECD) of hDectin-1b and used it to stain tumor cells. The recombinant ECD showed a specific binding to several human tumor cell lines that could be increased by induction of apoptosis in malignant cells and inhibited by incubation of the extracellular domain with zymosan, a crude cell wall extract of saccharomyces cerevisiae. Furthermore, uptake of tumor cells by immature mDC was reduced by zymosan or the presence of the recombinant Dectin-1 in phagocytosis assays suggesting that Dectin-1 is involved in the engulfement of tumor cells by DC. In line with the expression analysis of hDectin-1, we found that phagocytosis of apoptotic cells was dramatically reduced upon stimulation of DCs with Poly I:C or LPS as compared to immature DC or DC activated with TLR2 or 7 ligands. We next analysed the role of Dectin-1 in the cross-presentation of cell derived antigens and used DC that were incubated with CMV infected fibroblasts and an autologous CTL line specific for the HLA-A2 binding pp65 peptide. Stimulation of CMV peptide specific CTL in ELIspot assays by DC that were incubated with CMV infected fibroblasts efficiently stimulated IFN-gamma secretion that could be inhibited by zymosan and the extracellular domain of Dectin-1. In line with the results from phagocytosis assays stimulation of CMV specific CTL was reduced by stimulation of DC with TLR2 and 4 ligands.

Our results identify hDectin-1 as a new receptor for endogenous ligands on mammalian cells and indicate an important role of this molecule in the clearing of apoptotic and infected cells and cross-presentation of cell derived antigens..