Global proteogenomic analysis of human MHC class I-associated peptides derived from non-canonical reading frames

Abstract

In view of recent reports documenting pervasive translation outside of canonical protein-coding sequences, we wished to determine the proportion of major histocompatibility complex (MHC) class I-associated peptides (MAPs) derived from non-canonical reading frames. Here we perform proteogenomic analyses of MAPs eluted from human B cells using high-throughput mass spectrometry to probe the six-frame translation of the B-cell transcriptome. We report that ∼10% of MAPs originate from allegedly noncoding genomic sequences or exonic out-of-frame translation. The biogenesis and properties of these ‘cryptic MAPs' differ from those of conventional MAPs. Cryptic MAPs come from very short proteins with atypical C termini, and are coded by transcripts bearing long 3′UTRs enriched in destabilizing elements. Relative to conventional MAPs, cryptic MAPs display different MHC class I-binding preferences and harbour more genomic polymorphisms, some of which are immunogenic. Cryptic MAPs increase the complexity of the MAP repertoire and enhance the scope of CD8 T-cell immunosurveillance.

Selective graft-versus-leukemia depends on magnitude and diversity of the alloreactive T cell response

“…MiHA identification is probably more efficient when EBV-LCLs are used that have been SNP genotyped more extensively, for example, by whole-genome sequencing. Recently, it has been demonstrated that a proportion of the HLA ligandome is encoded by "noncoding" genomic regions that are not known to contain functional genes (35). Here, we confirm that these genomic regions indeed encode relevant T cell epitopes, as illustrated by the characterization of LB-C16ORF-1R as a novel MiHA that is targeted by CD8 T cells in the setting of alloSCT and DLI.…”

MHC class I–associated peptides derive from selective regions of the human genome

“…To obtain a comprehensive representation of the immunopeptidome presented by HLA-A and HLA-B molecules, we applied a well-validated high-throughput proteogenomic approach that hinges on a combination of next-generation sequencing and high-throughput MS (20,27,28). Transcriptome soned that if our MS analyses randomly missed some MAPs, the proportion of MAP source versus nonsource genes should nevertheless follow a binomial distribution where the number of source genes increases as a function of the number of detected MAPs (Figure 1A).…”

“…According to the dominant paradigm, MAPs preferentially originate from defective ribosomal products (DRiPs) which can be created by several mechanisms such as nonsense-mediated decay (NMD), mRNA destabilization, or noncanonical translation in the cytosol or the nucleus (16)(17)(18)(19)(20). Large-scale mass spectrometry (MS) offers the sole direct approach to analyzing the global molecular composition of the immunopeptidome.…”

Nowhere to hide: unconventional translation yields cryptic peptides for immune surveillance

“…These unusual translation products are referred to as cryptic peptides and form a substantial and significant part of the overall pMHC I repertoire (19). When Perreault and colleagues examined the MHC I associated peptides with mass spectrometry, they discovered ~10% of these peptides were actually encoded by transcripts normally considered to be untranslated.…”