Linda D. Barber

Overlap in the repertoires of peptides bound in vivo by a group of related class I HLA-B allotypes

BACKGROUND Polymorphism among class I molecules of the major histocompatibility complex (MHC) confers allotypic specificity on the peptides that these molecules bind and present to cytotoxic T lymphocytes. Evolution of new human HLA class I alleles usually involves gene recombination events that replace a segment of one allele with the homologous region of another. In this study, the impact of these evolutionary changes has been assessed by comparison of the peptide-binding specificities of six related HLA-B allotypes. RESULTS Endogenous peptides bound by HLA-B*5401, HLA-B*5501, HLA-B*5502, HLA-B*5601, HLA-B*6701 and HLA-B*0702 were characterized. Despite differing by 1-9 of the amino-acid residues comprising their peptide-binding sites, all these allotypes share a dominant preference for peptides that have proline at position 2. Polymorphism results in differing selection of carboxy-terminal and secondary anchor residues, but the peptide-binding specificities are sufficiently similar that there is overlap in the repertoires of peptides bound by these allotypes. Complete sequence determination of individual peptides revealed four that could be isolated from two or more allotypes. Members of the closely related HLA-B22 family--HLA-B*5401, HLA-B*5501, HLA-B*5502 and HLA-B*5601--show only minor differences in their peptide-binding specificities. This marked similarity is reflected at the functional level, as alloreactive cytotoxic T lymphocytes generated against HLA-B*5401 and HLA-B*5501 exhibited cross-reactive recognition. CONCLUSION The isolation of identical endogenously bound peptides from six HLA-B allotypes demonstrates overlap in the repertoires of peptides bound in vivo by different allotypes. We speculate that the shared preference for binding peptides with proline at position 2 reflects a selective pressure to retain this specificity, which may be based upon peptide availability in vivo. Characterization of the overlap between the repertoires of peptides bound by HLA-B allotypes could simplify the development of peptide-based vaccines that are targeted to cytotoxic T cells, as single peptides would be effective for humans of different HLA types.

The Bw4/Bw6 difference between HLA-B*0802 and HLA-B*0801 changes the peptides endogenously bound and the stimulation of alloreactive T cells

Abstract HLA-B*0801 is unique among HLA-B allotypes in having dominant amino acid anchors at positions 3 and 5 of the peptide-binding motif. HLA-B*0802 is a variant of HLA-B*0801 in which the Bw6 sequence motif is replaced by a Bw4 sequence motif. This change, involving substitutions at positions 77, 80, 81, 82, and 83 of the B*08 heavy chain, is probably the result of a single evolutionary event of interallelic conversion. Moreover, the difference between B*0802 and B*0801 is sufficient to stimulate a cytotoxic T-cell response. To assess further the functional impact of the Bw4 motif on a B8 background, we compared the peptide-binding specificity of the B*0801 and B*0802 allotypes by sequencing the mixture of peptides endogenously bound to B*0802 and 12 individual peptides purified from that mixture. The HLA-B*0802 allotype, while able to bind some peptides bound by B*0801, has a broader repertoire of endogenously bound peptides than B*0801: the peptides bound by B*0802 are more variable in length and exhibit greater diversity in the carboxyl-terminal amino acid which interacts with the F pocket.

Localization of class i histocompatibility molecule assembly by subfractionation of the early secretory pathway

Class I molecules of the major histocompatibility complex bind peptides derived from cytosolic proteins and display them on the cell surface. This function alerts cytotoxic T cells to the presence of intracellular pathogens. Class I molecule assembly requires the association of the heavy chain with beta 2-microglobulin, accompanied by peptide loading via specific transporters. This study localizes where these assembly steps take place, using monoclonal antibodies recognizing class I molecules in different assembly states to analyze subcellular fractions of the early secretory pathway. The distribution of peptide-loaded class I molecules was more localized than the distribution of the total pool of class I molecules in the early secretory pathway. Loaded molecules colocalized with the peptide transporter, free heavy chains, and the chaperone calnexin in high density rough endoplasmic reticulum (RER) membranes. These data suggest that subunit assembly and peptide acquisition occur at the same intracellular site. Class I molecules also localized to less dense subfractions of the early secretory pathway, which contained comparatively less peptide-loaded molecules than the high density RER fractions, at steady state. Following a 15 degrees C temperature block, class I molecules accumulated in these less dense membrane fractions, indicating that these fractions represent the intermediate compartment where empty class I molecules are trapped in mutant cells. In the presence of cycloheximide, a pool of class I molecules recycling to the RER was detected suggesting empty molecules recycle to acquire peptide.

HLA and Disease

Publisher Summary This chapter studies correlation of HLA polymorphisms with susceptibility and resistance to various diseases. In 1973, the HLA-B27 antigen was at very high frequency (∼95%) in patients suffering from ankylosing spondylitis. By contrast, the frequency of HLA-B27 in the general population today is 2%. This striking observation prompted investigation of other diseases and numerous associations were found. Among these, the significance of the associations was highly variable and none were as strong as that first seen for HLA-B27 and ankylosing spondylitis. The chapter gives a current view of HLA-associated autoimmune disease. The development of disease involves a genetic predisposition resulting from a combination of factors at HLA and other genes. Even amongst the individuals who have the genetic predisposition, only a small minority of them become diseased. In those who do, the disease is triggered by the immune response to infection, during which one or a few T-cell clones escape from tolerance and become reactive.

HLA POlymorphism, Peptide-binding Motifs and T-Cell Epitopes

Comparing the sequences of alleles of the polymorphic class I and class II loci shows that nucleotide substitutions are concentrated in the exons that encode the peptide-binding groove and the site of interaction with the T-cell receptor. Differences in amino acid sequence between allotypes and isotypes do not change the basic structure of HLA class I or class II molecules. Conserved amino acids in the peptide-binding site allow a network of hydrogen bonds to be formed with the peptide main chain, which provides most of the binding affinity. This enables a broad spectrum of peptides to be displayed by HLA molecules for effective surveillance of pathogen-derived peptides. However, the diversity is not infinite. To overcome these problems, the more sensitive technique of mass spectrometry has been applied to analysis of HLA-bound peptides. For the pool of peptides bound by an HLA class I or II allotype, a description of the anchor residues and their positions within the peptide sequence is called the peptide-binding motif of the allotype. The ambiguous nature of class II peptide-binding motifs means that their application to the identification of CD4 T-cell epitopes is often not informative.

The Organization of HLA Genes Within the HLA Complex

The genes that encode the HLA class I and II alloantigen are closely linked to each other on the short arm of human chromosome 6. This part of the genome constitutes the human major histocompatibility complex (MHC) and is called the HLA complex 1,2 . The HLA complex encompasses some four million base pairs of DNA and is of a size comparable to the genome of the common intestinal bacterium Escherichia coli. This chapter presents the HLA class I region, it shows the class I genes and related genes. HLA-H, -J, -K and -L are complete class 1 genes that are not expressed, that is, they are pseudo genes. The MIC gene family consists of five genes, of which MICA and MICE are expressed, MICC, MICD and MICE are pseudo genes. The unlabelled genes (short vertical bars) are fragments of class I genes. HFE is a functional class I-like gene found ∼4Mb telomeric of HLA-F. In addition to the genes, some additional genes, which are interspersed amongst the class I genes and are structurally unrelated to them are also reviewed.

12 – Evolution and Anthropology of HLA

Comparison of the major histocompatibility complex in different vertebrate species reveals a diversity in the class I and class II genes, which is imposed upon a common background of other genes. In the laboratory, mice that express neither class I genes nor class II genes develop, survive and reproduce. Such experiments demonstrate that polymorphic genes like HLA-A, -B, -C, -DP, -DQ and -DR are not involved in development or housekeeping functions of the mammalian body, but are dedicated to its protection once it is up, running and exposed to the microbiological environment. An important effect of the evolution of multiple polymorphic class I and II HLA genes is that it provides individual human beings with a diversity of class I and II antigen-presenting functions. Thus, this chapter discusses the evolution and anthropology of HLA. The number of alleles that can be maintained in a population under a given level of selection is correlated with the size of the population. In small populations alleles are more likely to be lost by chance, a consequence of what is called genetic drift.

HLA Class I and II Molecules Present Peptide Antigens to Different Types of T Cell

This chapter examines how CDS T cells recognize peptide antigens presented by HLA class I molecules, and illustrates the pathway by which extracellular antigens are processed and presented by HLA class II molecules. It explains extracellular proteins are taken into the cell by endocytosis or phagocytosis, and are then degraded to peptides within endosomes and lysosomes. The peptides are then sorted into MIIC vesicles where they can meet HLA class II molecules. HLA class II α and β chains and the invariant chain (li) are synthesized on ribosomes and translocated into the lumen of the ER, where they assemble into heterotrimers that cannot bind peptides because the invariant chain occupies the peptide-binding site. The class II heterotrimers leave the ER and pass through the Golgi apparatus to enter MIIC vesicles. There the invariant chain is degraded and with the help of HLADM and HLA-DO (not shown) a peptide can be bound. Complexes of HLA class II and peptide are then taken to the plasma membrane where they can be recognized by CD4 T cells. This pathway is used to respond to infection by the species of bacteria that live and replicate in the connective tissues. Macrophages phagocytose bacteria and present bacterial peptides to CD4 T cells. Some CD4 T cells are activated to secrete cytokines that directly act on macrophages to improve the rate at which they kill bacteria; other CD4 T cells stimulate B cells to produce bacteria-specific antibodies, which by coating bacteria make them more susceptible to phagocytosis by macrophages

Guide to FactsBook Tables

This chapter provides a guide to facts, such as peptide-binding specificity, alleles, serological specificity, and cells sequenced. The peptide-binding motif of an HLA allotype describes the anchoring residues preferentially bound and their position within the peptide sequence. Dominant anchors are highlighted in boldface and are those positions where one or a few closely related amino acids exclusively occupy the position. Amino acids that are enriched at a position are shown in plain text. Residues are listed in descending order of preference. Analysis of peptide binding by HLA-DR is complicated when the individual contributions of the two functional DRB genes expressed by most haplotypes are not distinguished.

Alloreactions in Transplantation

This chapter discusses that bone marrow transplantation (BMT) is more sensitive to HLA difference than solid-organ transplantation. The success of transplantation depends upon the HLA match between donor and recipient. Various other minor histocompatibility antigens have been defined by studying alloreactive CDS T-cell responses in patients who have received HLA-identical bone marrow transplants. For those patients who need a bone marrow transplant and do not have an HLA compatible donor within the family, a search can be made for an unrelated donor, who is HLA matched. Graft-versus-host disease (GVHD) is caused by mature T cells in the transplanted bone marrow, which may even derive from the peripheral blood that contaminates all bone marrow aspirates. In bone marrow transplantation the immunological situation is different from that in solid organ transplantation. Here the recipients immune system is deliberately destroyed by treatment with radiation and cytotoxic drugs, before transplantation with a source of pluripotent haematopoietic stem cells that will in time reconstitute the immune system. The dominant alloreactions that arise after bone marrow transplantation are caused by mature T cells in the transplanted bone marrow, which respond to the major and minor histocompatibility antigens expressed by the cells and tissues of the recipient.