MHC I–associated peptides preferentially derive from transcripts bearing miRNA response elements

Abstract

MHC I-associated peptides (MIPs) play an essential role in normal homeostasis and diverse pathologic conditions. MIPs derive mainly from defective ribosomal products (DRiPs), a subset of nascent proteins that fail to achieve a proper conformation and the physical nature of which remains elusive. In the present study, we used high-throughput proteomic and transcriptomic methods to unravel the structure and biogenesis of MIPs presented by HLA-A and HLA-B molecules on human EBV-infected B lymphocytes from 4 patients. We found that although HLA-different subjects present distinctive MIPs derived from different proteins, these MIPs originate from proteins that are functionally interconnected and implicated in similar biologic pathways. Secondly, the MIP repertoire of human B cells showed no bias toward conserved versus polymorphic genomic sequences, were derived preferentially from abundant transcripts, and conveyed to the cell surface a cell-type-specific signature. Finally, we discovered that MIPs derive preferentially from transcripts bearing miRNA response elements. Furthermore, whereas MIPs of HLA-disparate subjects are coded by different sets of transcripts, these transcripts are regulated by mostly similar miRNAs. Our data support an emerging model in which the generation of MIPs by a transcript depends on its abundance and DRiP rate, which is regulated to a large extent by miRNAs. (Blood. 2012;119(26):e181-e191)

Expanding the detectable HLA peptide repertoire using electron-transfer/higher-energy collision dissociation (EThcD)

“…Comparison of the complete dataset with recent large-scale studies revealed that 81% of the peptides have not been reported before (SI Appendix, Table S2) (16)(17)(18). Remarkably, the unique peptides identified in this study have distinct sequence characteristics compared with literature data (SI Appendix, Fig S4).…”

“…In contrast, biases in allele-specific peptide identification results could be directly translated to the known limitations of conventional peptide fragmentation techniques. This suggests that the outcome of HLA class I peptide identification studies that are based on CID (HCD) (16,18) or ETD (29) alone may only partially reflect the actual landscape of peptides presented at the cell surface. EThcD as the universal fragmentation method is therefore an attractive alternative to current MS sequencing technology.…”

“…However, although the major part of the proteome seemed suited for HLA sampling, the actual small number of peptides identified per source protein confirms that considerable selection of peptides takes place, related to proteolytic cleavage, efficiency of TAP and the binding affinity of the available HLA molecule (16,20). Earlier studies reporting fewer numbers of peptides (hundreds) demonstrated that most proteins were represented by only a single peptide (18). Hence, fragmentation techniques that can capture the full landscape of HLA class I peptides in cells of interest, such as EThcD, can make an important step toward the next stage of discovery.…”

The Effect of Proteasome Inhibition on the Generation of the Human Leukocyte Antigen (HLA) Peptidome

“…Furthermore, although the proteasome was proposed as the main protease responsible for degradation of newly synthesized proteins, including DRiPs and SLiPs, autophagy and lysosomal proteolysis were mostly associated with degradation of long-lived, house-keeping proteins and whole organelles at the end of their functional lifecycles (64). Of interest is a recent study that suggests that a significant fraction of the MHC peptidome is derived from proteins whose transcripts have micro-RNA elements, which may affect the DRiPs production rates of the encoded proteins (69). The quantitative data provided in this study, points to the possible large involvement of such nonproteasomal pathways with the production of the HLA class I peptidome, including contributions from newly synthesized proteins.…”

The nature and extent of contributions by defective ribosome products to the HLA peptidome

“…In cultured cells, upward to a third of the MHC peptidome was suggested to be derived from the latter. These rapidly degraded defective ribosome products (DRiPs) (4,9) were proposed to be produced by exotic mechanisms, such as downstream initiation (10), transcripts associated with specific microRNAs (11), premature termination of translation (12)(13)(14), short protein segments, produced by the pioneer round of translation (15) during the nonsense mediated decay (NMD) quality control of mRNA (16,17), and even out-of-frame translated segments of proteins (18). MHC peptides derived from newly synthesized proteins were also suggested to be produced by degradation of normal, fully functional polypeptides, which are the excess subunits of large protein complexes (19,20).…”