Dataset Information

Mass spectrometry of HLA-I peptidomes reveals strong effects of protein abundance and turnover on antigen presentation

ABSTRACT: HLA class I molecules reflect the health state of cells to cytotoxic T-cells by presenting a repertoire of endogenously derived peptides. However, the extent to which the proteome shapes the peptidome is still largely unknown. Here we present a high-throughput mass-spectrometry-based workflow that allows stringent and accurate identification of thousands of such peptides and direct determination of binding motifs. Applying the workflow to seven cancer cell lines and primary cells, yielded more than 22,000 unique HLA peptides across different allelic binding specificities. By computing a score representing the HLA-I sampling density, we show a strong link between protein abundance and HLA-presentation (P<0.0001). When analyzing over-presented proteins - those with at least five-fold higher density score than expected for their abundance – we noticed that they are degraded almost 3 hours faster than similar but non-presented proteins (top 20% abundance class; median half-life 20.8h vs. 23.6h, p<0.0001). This validates protein degradation as an important factor for HLA presentation. Ribosomal, mitochondrial respiratory chain and nucleosomal proteins as particularly well presented. Taking a set of proteins associated with cancer, we compared the predicted immunogenicity of previously validated T-cell epitopes with other peptides from these proteins in our dataset. The validated epitopes indeed tend to have higher immunogenic scores than the other detected HLA peptides, suggesting the usefulness of combining MS-analysis with immunogenesis prediction for ranking and selection of epitopes for therapeutic use.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): B Cell, Fibroblast

SUBMITTER: Mario Oroshi

LAB HEAD: Matthias Mann

PROVIDER: PXD000394 | Pride | 2015-01-12

REPOSITORIES: Pride

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Publications

Mass spectrometry of human leukocyte antigen class I peptidomes reveals strong effects of protein abundance and turnover on antigen presentation.

Bassani-Sternberg Michal M Pletscher-Frankild Sune S Jensen Lars Juhl LJ Mann Matthias M

Molecular & cellular proteomics : MCP 20150109 3

HLA class I molecules reflect the health state of cells to cytotoxic T cells by presenting a repertoire of endogenously derived peptides. However, the extent to which the proteome shapes the peptidome is still largely unknown. Here we present a high-throughput mass-spectrometry-based workflow that allows stringent and accurate identification of thousands of such peptides and direct determination of binding motifs. Applying the workflow to seven cancer cell lines and primary cells, yielded more t ...[more]

PMID: 25576301

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Project description:HLA-DR molecules are highly expressed in synovial tissue (ST), the target of the immune response in chronic inflammatory arthritides, including in rheumatoid arthritis (RA) and Lyme arthritis (LA). In the current study, we identified HLA-DR-presented self-peptides (T cell epitopes) in ST, synovial fluid mononuclear cells (SF), and peripheral blood mononuclear cells (PB) from five patients with RA and eight with LA. Altogether, from 22 samples, 1,532 non-redundant HLA-DR-presented peptides were identified that were derived from 778 source proteins. Moreover, as the study progressed, application of newer, high sensitivity LC-MS instruments resulted in the identification of larger numbers of HLA-DR-presented peptides. Surprisingly, the source proteins for HLA-DR-presented peptides were as likely to have intracellular as extracellular locations. Although the number of peptides identified was greater in ST than in SF or PB, 68% of the peptides identified in SF were found in ST, and 55% of the peptides in PB were found in ST. Two RA patients had the same HLA-DR genotype (DRB1*0401 and 0101), and 44% of the peptides identified in these two patients were the same. In contrast, among the patients with RA or LA who had no shared HLA-DR alleles, only 6% of the peptides were the same. In the RA patients, HLA-DR-presented peptides were identified from source proteins that are thought to serve as potential autoantigens: collagen, enolase, fibrinogen, fibronectin, immunoglobulin and vimentin. Interestingly, peptides from these same source proteins were also commonly found in LA patients. However, citrullinated T cell epitopes were not identified in any patient. Thus, LC-MS/MS techniques are now sensitive enough to identify large numbers of T cell epitopes from tissue or fluids in individual patients. Importantly, analysis of SF or PB allowed us to identify a broader range of HLA-DR-presented self-peptides from individual patients and from patients seen earlier in the disease.

Dataset Information

Mass spectrometry of HLA-I peptidomes reveals strong effects of protein abundance and turnover on antigen presentation

Publications

Mass spectrometry of human leukocyte antigen class I peptidomes reveals strong effects of protein abundance and turnover on antigen presentation.

Similar Datasets

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