Activated dectin-1 localizes to lipid raft microdomains for signaling and activation of phagocytosis and cytokine production in dendritic cells.

ABSTRACT

Lipid rafts are plasma membrane microdomains that are enriched in cholesterol, glycosphingolipids, and glycosylphosphatidylinositol-anchored proteins and play an important role in the signaling of ITAM-bearing lymphocyte antigen receptors. Dectin-1 is a C-type lectin receptor (CLR) that recognizes beta-glucan in the cell walls of fungi and triggers signal transduction via its cytoplasmic hemi-ITAM. However, it is not known if similar to antigen receptors, Dectin-1 would also signal via lipid rafts and if the integrity of lipid raft microdomains is important for the physiological functions mediated by Dectin-1. We demonstrate here using sucrose gradient ultracentrifugation and confocal microscopy that Dectin-1 translocates to lipid rafts upon stimulation of dendritic cells (DCs) with the yeast derivative zymosan or beta-glucan. In addition, two key signaling molecules, Syk and PLCgamma2 are also recruited to lipid rafts upon the activation of Dectin-1, suggesting that lipid raft microdomains facilitate Dectin-1 signaling. Disruption of lipid raft integrity with the synthetic drug, methyl-beta-cyclodextrin (betamD) leads to reduced intracellular Ca2+ flux and defective Syk and ERK phosphorylation in Dectin-1-activated DCs. Furthermore, betamD-treated DCs have significantly attenuated production of IL-2, IL-10, and TNFalpha upon Dectin-1 engagement, and they also exhibit impaired phagocytosis of zymosan particles. Taken together, the data indicate that Dectin-1 and perhaps also other CLRs are recruited to lipid rafts upon activation and that the integrity of lipid rafts is important for the signaling and cellular functions initiated by this class of innate receptors.