Harry T. Orr

Major histocompatibility antigens: The human (HLA-A,-B,-C) and murine (H-2K, H-2D) class I molecules

Hidde L. Ploegh,* Harry T. Orrt and Jack L. Strominger Harvard University The Biological Laboratories Cambridge, Massachusetts 02138 Introduction More than 40 years ago skin grafting experiments carried out in mice showed that the rejection of grafts is under genetic control. Differences between individ- ual mice at any of a number of loci could lead to graft rejection. However, one locus stood out in its ability to provoke graft rejection; it was subsequently named the H-2 locus, or major histocompatibility complex (MHC) (Klein, 1975). Not until some 25 years later, by a rather different route, could a similar region be identified in humans, which has been named the HLA region (Bach and van Rood, 1976). Although originally discovered through grafting ex- periments, the MHC has been analyzed most exten- sively with serological reagents. In mice, individual animals will produce antibodies after deliberate im- munization with lymphoid cells or after skin grafting when differences exist between the immunizing and responding animal with respect to histocompatibility loci. Sera of exquisite specificity can be obtained through the use of congenic mouse strains (strains of mice that have identical genetic backgrounds, but differ only in a well defined segment of the genome- for example, in the MHC or a portion of it [Snell, 1980]). With the advent of monoclonal antibodies (Moller, 1979) the specificity of serologic reagents can be even more precise. In humans, most antibodies for the analysis of the HLA system are derived from women who have been pregnant multiple times and are immunized by pater- nal antigens present on fetal cells in the maternal circulation. Depending on the differences in HLA type and the degree of antigenic stimulation, these re- agents can be of sufficiently high titer and specificity to be used as tissue-typing reagents. Again, mono- clonal antibodies may improve the serological defini- tion of HLA antigens (Parham and Bodmer, 1978; Brodsky et al., 1979). One of the features of the MHC that was quickly uncovered is its extensive polymorphism (Klein, 1979). It is now thought that at each of the recognized loci in the H-2 complex up to a hundred alleles may exist in the population. For the HLA loci, a high degree of polymorphism has also been demonstrated (Figure 1). By applying serological techniques to the study of inbred strains of mice and human families carrying

Assembly and maturation of HLA-A and HLA-B antigens in vivo.

HLA-A and HLA-B antigens are integral membrane glycoproteins which consist of a glycosylated heavy chain embedded in the membrane in noncovalent association with beta 2-microglobulin, a water-soluble polypeptide. The assembly and maturation of these antigens has been studied in vivo in the human B lymphoblastoid cell line T5-1 (HLA-A1, -A2, -B8, -B27). Two antigenically distinct populations of HLA-A and -B heavy chains can be detected by antisera which recognize determinants sensitive to the conformation of the heavy chain. One heavy chain population is associated with beta 2-microglobulin, whereas the other population is not. These populations can be further distinguished by their oligosaccharide structure and their localization within the cell. Pulse-chase experiments demonstrate a precursor-product relationship between these heavy chain populations and suggest the following pathway for the assembly and maturation of HLA-A and -B antigens. The completed heavy chains initially carry high mannose oligosaccharides and are largely or wholly associated with beta 2-microglobulin. During the next 10-15 min, association with beta 2-microglobulin occurs and the heavy chain conformation is altered. Beginning at about 30 min after synthesis, the oligosaccharides are converted from the high mannose form to the complex form, and between 60 and 80 min after synthesis, the mature antigens appear at the cell surface. These observations are discussed in relation to in vivo and in vitro studies on the biosynthesis of a variety of secreted proteins and membrane proteins.

An Evaluation of the Significance of Amino Acid Sequence Homologies in Human Histocompatibility Antigens (HLA‐A and HLA‐B) with Immunoglobulins and Other Proteins, Using Relatively Short Sequences

A computer search was carried out for homologies between HLA‐A and HLA‐B antigen sequences and the sequences of constant and variable regions of immunoglobulins and of all other sequenced proteins. Searches were made both with relatively short peptide sequences from the HLA antigens and with those longer peptide sequences which were available in 1978. Significant homology of HLA antigen sequences to immunoglobulin constant region sequences was found in two cases: (1) a short decapeptide sequence which includes the fourth cysteine residue of HLA‐B7 and (2) an 89‐amino‐acid residue (Ac‐2) C‐terminal fragment of the papain‐solubilized HLA‐B7 molecule. The difficulty of establishing statistically significant sequence homology with relatively short peptide sequences is emphasized by computer‐based comparisons of the decapeptide sequence with randomly generated peptide sequences. It is concluded that statistically significant homology with short sequences can be assured only when extraordinarily high degrees of homology are present and additional constraints are included in the matches, for example, matches at relatively rare amino acid residues such as Cys, His and Trp. The homology of the 89‐amino‐acid residue sequence to constant region sequences of immunoglobulins is as great as or greater than that of β2‐microglobulin. These findings and the unique domain structure involving a disulphide loop of comparable size strongly favour a common evolutionary origin for this region of HLA‐A and ‐B, β2‐microglobulin and immunoglobulin constant regions.

Structure, Function, and Biosynthesis of the Major Human Histocompatibility Antigens (HLA-A and HLA-B)

Summary

Monoclonal Antibodies for Analysis of HLA Antigens: Further Studies with the W6/32 Antibody

Many mouse monoclonal antibodies against HLA antigens have been described and their applications to tissue-typing, biological and biochemical research are being explored. A number of our studies revealed certain problems associated with an ignorance of the basic immunochemical properties of the monoclonal reagents we were using.

Complete Primary Structure of Human Histocompatibility Antigen HLA-B7

Tissue grafts cannot be exchanged successfully in the absence of immuno-suppression, except between syngeneic strains or identical twins. From a biochemical standpoint, each individual of a species carries on the surface of its cells substances now known to be glycoproteins that are recognized by every other individual of that species as foreign. These glycoproteins are the products of a highly polymorphic genetic system, first discovered through mouse-breeding studies. Largely through the efforts of Peter Gorer in England and George Snell in Bar Harbor, a region was identified, called the H-2 region, differences at which are the two major histocompatibility antigens, called H-2K and H-2D, were products of two genetic loci in the H-2 region on the 17th mouse chromosome. Subsequently, a variety of othr loci have been identified that are in some manner involved in graf rejection or acceptance. This region is organized similarly in all species that have been examined and is now called the major histocompatibility complex (MHC).

Chapter 4 Complete Primary Structure of Human Histocompatibility Antigen Hla-B7: Evolutionary and Functional Implications

Publisher Summary This chapter focuses on the complete primary structure of human histocompatibility antigen— HLA-B7. The histocompatibility antigens and immunoglobulins are both probably the descendants of some common ancestral gene, which may have originally coded for a cell-bound defense molecule. The development of animals with circulatory systems and duplication of that gene resulted later in an evolution in the secretion of the product of some of these genes into the circulation—i.e., the immunoglobulins. The product of the other genes remained membrane bound and probably went on to evolve to become the histocompatibility antigens. The histocompatibility antigens also serve a defense function for the organism. In this case, the cell-bound defense mechanism may serve to protect the organism from invasion by closely related cells, perhaps serving in primitive organisms to maintain colonial identity on the sea floor. At a different level, the organization of information within a genome can be approached via the cloning of major histocompatibility complex genes by recombinant DNA techniques.