TLR9 Activation Increases TAP-Independent Vesicular MHC Class I Processing In vivo

Cross-presentation of soluble protein antigens on major histocompatibility complex (MHC) class I by dendritic cells (DC) can occur in vesicular, endolysosomal compartments and be either dependent or independent of TAP peptide transporters. Here we investigate if an immunostimulatory CpG oligodeoxynucleotide can increase the activity in a TAP-independent endolysosomal vesicular pathway (el-VP) in vivo as we have earlier found in in vitro cultured DC. We use the in vivo response of CFSE labelled OT-1 T cells, transgenic for a T-cell receptor (TCR) that recognizes an ovalbumin (OVA)-derived peptide (SIINFEKL) presented by H-2Kb, transferred into TAP1-/- mice, as a functional read-out for activity in the el-VP. We have found a poor OT-1 T-cell response to soluble OVA which, however, could be strongly enhanced by the simultaneous administration of CpG. This increased responsiveness required both the endolysosomal cathepsin S (CatS) and Toll like receptor (TLR)9, the CpG receptor, both of which are present in the el-VP. Confocal microscopy demonstrated a co-localization of H-2Kb/SIINFEKL and the endolysosomal marker LAMP1 in CD11c positive DC which was markedly increased by CpG administration. No complexes were found in the ER and cis-Golgi compartments in TAP1-/- mice, indicating the lack of classical MHC-I processing. In DC isolated from CatS-/- mice the opposite was found, complexes were present in the ER but not in the el-VP. We conclude that in vivo activation of TLR9 by CpG increases the efficiency of TAP independent el-VP and that this might contribute to the potent adjuvant activity of this type of compound. The cellular mechanisms remain to be established.