AKAPs in lipid rafts are required for optimal antigen presentation by dendritic cells

Dendritic cell (DC) maturation and antigen presentation are regulated by activation of protein kinase A (PKA) signaling pathways, through unknown mechanisms. We have recently shown that interfering with PKA signaling through the use of anchoring inhibitor peptides hinders antigen presentation and DC maturation. These experiments provide evidence that DC maturation and antigen presentation are regulated by A-kinase anchoring proteins (AKAPs). Herein, we determine that the presence of AKAPs and PKA in lipid rafts regulates antigen presentation. Using a combination of western blotting and immuno-cytochemistry, we illustrate the presence of AKAP149, AKAP79, Ezrin and the regulatory subunits of PKA in DC lipid rafts. Incubation of DCs with the type II anchoring inhibitor, AKAP-in silico (AKAP-IS), removes Ezrin and RII from the lipid raft without disrupting raft formation. Addition of a lipid raft disruptor, methyl-β-cyclodextrin, blocks the efficacy of AKAP-IS, suggesting that the lipid raft must be intact for AKAP-IS to inhibit antigen presentation. Ezrin and AKAP79 are present in the lipid raft of stimulated KG1 cells, but Ezrin is not present in the lipid raft of unstimulated KG1 cells and AKAP79 levels are greatly diminished, suggesting that Ezrin and AKAP79 may be the key AKAPs responsible for regulating antigen presentation.