Anita J. Zaitouna

Major histocompatibility class I (MHC-I) proteins mediate immunosurveillance against pathogens and cancers by presenting antigenic or mutated peptides to antigen receptors of CD8+ T cells and by engaging receptors of natural killer (NK) cells. In humans, MHC-I molecules are highly polymorphic. MHC-I variations permit the display of thousands of distinct peptides at the cell surface. Recent mass spectrometric studies have revealed unique and shared characteristics of the peptidomes of individual MHC-I variants. The cell surface expression of MHC-I–peptide complexes requires the functions of many intracellular assembly factors, including the transporter associated with antigen presentation (TAP), tapasin, calreticulin, ERp57, TAP-binding protein related (TAPBPR), endoplasmic reticulum aminopeptidases (ERAPs), and the proteasomes. Recent studies provide important insights into the structural features of these factors that govern MHC-I assembly as well as the mechanisms underlying peptide exchange. Conformational sensing of MHC-I molecules mediates the quality control of intracellular MHC-I assembly and contributes to immune recognition by CD8 at the cell surface. Recent studies also show that several MHC-I variants can follow unconventional assembly routes to the cell surface, conferring selective immune advantages that can be exploited for immunotherapy.

The diversity of peptides displayed by class I human leukocyte antigen (HLA) plays an essential role in T cell immunity. The peptide repertoire is extended by various posttranslational modifications, including proteasomal splicing of peptide fragments from distinct regions of an antigen to form nongenomically templated cis-spliced sequences. Previously, it has been suggested that a fraction of the immunopeptidome constitutes such cis-spliced peptides; however, because of computational limitations, it has not been possible to assess whether trans-spliced peptides (i.e., the fusion of peptide segments from distinct antigens) are also bound and presented by HLA molecules, and if so, in what proportion. Here, we have developed and applied a bioinformatic workflow and demonstrated that trans-spliced peptides are presented by HLA-I, and their abundance challenges current models of proteasomal splicing that predict cis-splicing as the most probable outcome. These trans-spliced peptides display canonical HLA-binding sequence features and are as frequently identified as cis-spliced peptides found bound to a number of different HLA-A and HLA-B allotypes. Structural analysis reveals that the junction between spliced peptides is highly solvent exposed and likely to participate in T cell receptor interactions. These results highlight the unanticipated diversity of the immunopeptidome and have important implications for autoimmunity, vaccine design, and immunotherapy. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Human leukocyte antigen class I (HLA-I) variants can be linked to many thousands of different peptides, termed the ‘peptidome’. In this paper, Sarkizova et al. describe the development of a tool to provide prediction models for endogenous peptide presentation with the goal of improving epitope predictions for cancer vaccine design and immunotherapies. They were thorough in characterizing a total of 96 HLA-A, HLA-B, HLA-C, and HLA-G peptidomes, using this and other information to develop a prediction tool, which was validated using cancer cell lines. This Recommendation is of an article referenced in a Faculty Review also written by Anita J. Zaitoua, Amanpreet Kaur and Malini Raghavan.

Immunoaffinity purification is widely used in the pursuit of the ‘peptidomes’ of major histocompatibility complex class I (MHC-I) molecules within primary tissues and cells, via the use of antibodies specific to MHC-I. Partridge et al. are able to identify peptides that co-isolate with and contaminate peptidome preparations, which suggests that peptidome data must be interpreted more carefully to reduce incorrect identifications of T cell epitope candidates. This Recommendation is of an article referenced in a Faculty Review also written by Anita J. Zaitoua, Amanpreet Kaur and Malini Raghavan.

A number of recent studies {1,2} have attempted to quantify the fraction of human leukocyte antigen class I (HLA-I) ligandomes that correspond to spliced epitopes, both cis-spliced (spliced peptides originating from the same precursor) and trans-spliced (spliced peptides originating from different precursors). This study by Mylonas et al. highlights that the validity of some of the approaches used for quantifying the spliced peptide fraction remains somewhat controversial. To date, only a small number of the possible spliced peptides have been confirmed as products of proteasomal splicing or directly validated as spliced epitopes based on other approaches. This Recommendation is of an article referenced in a Faculty Review also written by Anita J. Zaitoua, Amanpreet Kaur and Malini Raghavan.

Major histocompatibility complex-I-β2m dimers (MHC-I) bind peptides derived from intracellular proteins, enabling the immune system to distinguish between normal cells and those expressing pathogen-derived or mutant proteins. The peptides bind to MHC-I in the endoplasmic reticulum (ER), and this binding is facilitated by the peptide loading complex (PLC), which contains calreticulin (CRT). CRT associates with MHC-I via a conserved glycan present on MHC-I and recruits it to the PLC for peptide binding. Somatic frameshift mutations in CRT (CRT-FS) drive the proliferation of a subset of myeloproliferative neoplasms, which are chronic blood tumors. All CRT-FS proteins have a C-terminal sequence lacking the normal ER-retention signal and possessing a net negative charge rather than the normal positive charge. We characterized the effect of CRT-FS on antigen presentation by MHC-I in human cells. Our results indicate that CRT-FS cannot mediate CRT's peptide loading function in the PLC. Cells lacking CRT exhibited reduced surface MHC-I levels, consistent with reduced binding of high-affinity peptides, and this was not reversed by CRT-FS expression. CRT-FS was secreted and not detectably associated with the PLC, leading to poor MHC-I recruitment, although CRT-FS could still associate with MHC-I in a glycan-dependent manner. The addition of an ER-retention sequence to CRT-FS restored its association with the PLC but did not rescue MHC-I recruitment or its surface expression, indicating that the CRT-FS mutants functionally compromise the PLC. MHC-I down-regulation permits tumor cells to evade immune surveillance, and these findings may therefore be relevant for designing effective immunotherapies for managing myeloproliferative neoplasms. © 2018 Arshad and Cresswell.

Herpesviruses are able to evade immune recognition by many mechanisms, including disrupting the expression or function of the transporter associated with antigen processing (TAP), which transports peptides into the endoplasmic reticulum (ER) for assembly with major histocompatibility complex class I (MHC-I) molecules. This recent study shows that tapasin, a key component of the MHC-I pathway, becomes a degradation target in molluscum contagiosum virus (MCV)-infected cells, thereby negatively affecting MHC-I assembly. Harvey et al. find that MC80, a MCV protein, binds to the tapasin lumenal domain, targeting it for degradation. TAP, which binds to tapasin, is also degraded, leading to a reduction both in the ER peptide pool and in tapasin function in MCV-infected cells. This Recommendation is of an article referenced in a Faculty Review also written by Anita J. Zaitoua, Amanpreet Kaur and Malini Raghavan.

Calreticulin, an endoplasmic reticulum (ER) resident chaperone, is translocated from ER to plasma membrane under certain conditions of cell stress and cell death. The cell surface calreticulin acts as an ‘eat me signal’ and facilitates phagocytosis of stressed cells by macrophages. Mutant forms of calreticulin have been shown to drive cellular transformation and proliferation in myeloproliferative neoplasms (MPNs) (see {1,2}). This study by Liu et al. demonstrates an immunosuppressive role for MPN mutant calreticulin proteins. The authors show that MPN calreticulin mutants are secreted and that the secreted forms of MPN calreticulin mutants inhibit phagocytosis of dying cancer cells by dendritic cells. Such immune evasion imparted by extracellular forms of mutant calreticulin proteins abrogates the protective effects of anti-tumor drugs that activate immunological cell death via cell surface calreticulin expression. This study suggests that secretion of MPN calreticulin mutants contributes to immunosuppression and tumor progression. This Recommendation is of an article referenced in a Faculty Review also written by Anita J. Zaitoua, Amanpreet Kaur and Malini Raghavan.

This study by Lorente et al., involving the analysis of major histocompatibility complex class I (MHC-I) peptidomes from spleen cells of mice deficient in transporter associated with antigen processing (TAP), reports that the peptidome is enriched in signal sequence-derived peptides, as well as peptides derived from precursors in the extracellular space. Notably, endoglycosidase H (Endo-H) resistant forms of HLA-B are detectable on the cell-surface under TAP-deficiency conditions, indicating a role for unconventional secretory pathways in trafficking of human leukocyte antigen (HLA)-B from the ER to the cell surface in TAP-deficient cells (see {1}). While the nature of these pathways remains to be elucidated, it is possible that this mode of trafficking allows for peptide loading of endocytosed extracellular antigens within endolysosomal compartments. This Recommendation is of an article referenced in a Faculty Review also written by Anita J. Zaitoua, Amanpreet Kaur and Malini Raghavan.

Deficiencies of components of antigen presentation machinery generally allow cancer cells to evade immune surveillance by CD8⁺ T cells. However, defective antigen presentation via conventional routes in cancers increase the likelihood of presentation of T-cell epitopes associated with impaired peptide processing (TEIPP), that can be exploited for cancer immunotherapies. This study by Marijt et al. reports the identification of 16 human leukocyte antigen (HLA)-A*02:01-restricted TEIPP neoantigens that are potential candidates for cancer immunotherapy. CD8⁺ T cells specific to these epitopes are present in healthy donors and are not affected by tolerance mechanisms. T cells against an epitope derived from the leader sequence of LDL receptor-associated protein 1 (LRPAP1) are shown to recognize transporter associated with antigen processing (TAP)-deficient tumors cells of different histological origins, but not healthy cells. This Recommendation is of an article referenced in a Faculty Review also written by Anita J. Zaitoua, Amanpreet Kaur and Malini Raghavan.