ERAAP Shapes the Peptidome Associated with Classical and Nonclassical MHC Class I Molecules

Abstract

The peptide repertoire presented by classical as well as non-classical MHC I molecules is altered in the absence of the ER aminopeptidase associated with antigen processing (ERAAP). To characterize the extent of these changes, peptides from cells lacking ERAAP were eluted from the cell surface and analyzed by high-throughput mass spectrometry. We found that the majority of peptides found in WT cells were retained in the absence of ERAAP. In contrast, a subset of “ERAAP-edited” peptides was lost in WT cells, and ERAAP-deficient cells presented an unique “unedited” repertoire. A substantial fraction of MHC-associated peptides from ERAAP-deficient cells contained N-terminal extensions and had a different molecular composition than those from WT cells. We found that the number and immunogenicity of peptides associated with non-classical MHC I was increased in the absence of ERAAP. Conversely, only peptides presented by classical MHC I were immunogenic in ERAAP-sufficient cells. Finally, MHC I peptides were also derived from different intracellular sources in ERAAP-deficient cells.

The Human Leukocyte Antigen (HLA)-B27 Peptidome in Vivo, in Spondyloarthritis-susceptible HLA-B27 Transgenic Rats and the Effect of Erap1 Deletion

“…Erap1 Is Not Necessary for Spondyloarthritis in Rats-The genetic linkage between ERAP1 and AS has been firmly established in humans (19,20). Deletion of ERAP1 has been shown to affect the repertoire of peptides bound to B27 in human cell lines (92) and in transgenic mice (93) and to have large-scale effects on the MHC-I peptidome in mice (43). However, HLA-B27 transgenic mice do not develop spondyloarthritis (93,94), and hence this study is the first to examine the effect of Erap1 deletion both on the B27 peptidome in vivo and on the spondyloarthritis phenotype.…”

“…Rodents lack Erap2 altogether (34), which makes rodent systems preferred models for the analysis of the effect of ERAP1 on the MHC peptidome, without interference from ERAP2. Erap1 knock-out mice (alternatively called Eraap in the mouse) show a dramatic alteration in their MHC-bound peptide repertoires, with partial loss of MHC-I expression, altered antigen presentation, binding of longer than normal peptides, and marked reciprocal cytolytic T cell alloreactivity between Erap1 Ϫ/Ϫ and wild type mice (37)(38)(39)(40)(41)(42)(43).…”

TAPBPR mediates peptide dissociation from MHC class I using a leucine lever

“…TAPBPR Øloop , in which amino acids comprising the 22-35 loop were replaced with amino acids glycine, alanine and serine, was created from TAPBPR WT pHRSIN-C56W-UbEM using the following procedure: First, amino acids 22-28 were replaced using quick-change site-directed mutagenesis using primers M22-for and M22-rev (see Table 2 for primer sequences). Subsequently, amino acids [29][30][31][32][33][34][35] were replaced using a two-step PCR. For this, the TAPBPR insert was amplified in two separate pieces, starting from each side of the mutation site (primers TAPBPR WT -BamHI-for and M29-rev for the N terminus-containing side and primers M29-for and TAPBPR WT -NotIrev for the C terminus-containing side).…”

“…Thus, technical limitations regarding the need to assign peptides to particular MHC I (by applying thresholds or cut-offs to determine which peptide was bound to which MHC I) greatly reduce the ability to detect peptides exhibiting low affinity for a particular MHC I. Other studies[17,32,33], exploring the affinity of peptides by immunopeptidomics use cells expressing only one MHC I. Therefore, the identified peptides do not need to be assigned to MHC I, thus improving the ability to see reliable changes…”

A systematic re-examination of processing of MHCI-bound antigenic peptide precursors by endoplasmic reticulum aminopeptidase 1

“…Several studies have described the effects of altered ERAP1 activity (either due to genetic manipulation or natural polymorphic variation) on the immunopeptidome of cell lines and in vivo models. ERAP1 has been found to influence a significant component of the immunopeptidome by altering both sequence and length of presented peptides (14)(15)(16)(17). These effects are generally interpreted to be the result of its aminopeptidase activity.…”