The Dendritic Cell MHC II Peptidome Derives from a Variety of Processing Pathways and Includes Peptides with a Broad Spectrum of HLA-DM Sensitivity

Abstract

Abstract The repertoire of peptides displayed in vivo by MHC II molecules derives from a wide spectrum of proteins produced by different cell types. While intracellular endosomal processing in dendritic cells and B cells has been characterized for few antigens the overall range of processing pathways responsible for generating the MHCII peptidome are currently unclear. To determine the contribution of non-endosomal processing pathways we eluted and sequenced over 3000 HLA-DR1-bound peptides presented in vivo by dendritic cells. Processing enzymes were identified by reference to a database of experimentally determined cleavage sites and experimentally validated for four epitopes derived from complement 3, collagen II thymosin b4 and gelsolin. We determined that self-antigens processed by tissue-specific proteases, including complement, matrix metalloproteases, caspases, and granzymes, and carried by lymph, contribute significantly to the MHC II self-peptidome presented by conventional dendritic cells in vivo. Additionally, presented peptides exhibited a wide spectrum of binding affinity and HLA-DM susceptibility. The results indicate that the HLA-DR1-restricted self-peptidome presented under physiological conditions derives from a variety of processing pathways. Non-endosomal processing enzymes add to the number of epitopes cleaved by cathepsins, altogether generating a wider peptide repertoire. Together with HLA-DM-dependent and HLA-DM-independent loading pathways, this ensures that a broad self-peptidome is presented by dendritic cells.