Biochemical characterization of activation‐associated bovine class I major histocompatibility complex antigens

Abstract

Utilizing a 'sandwich' ELISA assay we have been able to demonstrate that mAb W6/32, B1G6 and IL-A19 are reactive with three different monomorphic determinants on bovine class I major histocompatibility complex (MHC) molecules.Sequential immunoprecipitations performed with the mAb revealed that class I molecules on PBM comprise a single population with respect to reactivity with the mAb in that the P2m-associated proteins bear all three epitopes. By contrast, TCGF-driven lymphoblasts and cells transformed by Theileria parva (Tp) additionally express molecules of Mr 45000 bound to P2m which are recognized by mAb B1G6 and IL-A19 but not by W6/32. These two subclasses of molecules were further distinguished on the basis that, when tunicamycin was added to cultures in the preparation of cells for analysis, mAb W6/32 precipitated class I heavy chains of Mr 39000 while the extra molecules detected only by mAb B1G6 and IL-A19 were of Mr 37000 and 39000. On thymocytes, the mAb W6/32-non-reactive class I molecules are present in low amounts and are expressed by cells in the medulla area, unlike BoTl (analogous to human CD1) molecules which are expressed by the cortical cells. Our studies also revealed that the supposed P2m-specific mAb B1G6 does not recognize the P2m-associated molecules (BoT1) precipitated by mAb TH97A and thus the specificity of mAb B1G6 in cattle is for an epitope on bovine p2m which is strongly influenced by the nature of the heavy chain with which the P2m is associated. 241 242 A. Bensaid et al. heterozygous animal (Bensaid et al. 1988). Here we describe studies utilizing mAb IL-A19, detecting a monomorphic epitope of BOLA class I proteins, and mAb TH97A, defining BoTl molecules analogous to human CD1 (MacHugh et al. 1988).We have compared, with histo-and immunochemical techniques, the properties of the molecules carrying the determinants recognized by these mAb with those of the molecules carrying determinants for the cross-reacting mAb W6/32 and B1G6 in order to gain further insights into the diversity, structure and expression of class I MHC molecules in cattle. Materials and methods AnimalsA 3-month-old Boran (Bos indicus) calf was used as a donor of thymus cells. Peripheral blood mononuclear cells were collected from a 4-year-old Boran steer. Monoclonal antibodiesMAb W6/32 (IgG2a) recognizes an epitope on all HLA-A, B and C heavy chains associated with p2m (Barnstable et al. Parham et al. 1979). The mAb B1G6 (IgG2b) is directed against human P2m and recognizes this molecule both free and complexed with HLA heavy chains (Liabeuf etal. 1981). B1G6 has also been found to be reactive with bovine P2m (Chardon et al. 1983).The mAb IL-A19 (Ig2a) was derived in our laboratory from a hybridoma produced by fusion of the spleen cells of a Balb/c mouse with myeloma X63 .Ag. Prior to fusion, the mouse had received multiple inoculations with PBM from B. indicus and B. tuurus animals. Hybridoma selection and cloning was by standard methods. This mAb reacts with an epitope present on the surface of PBM of all ...

Bovine Neonatal Pancytopenia is a heritable trait of the dam rather than the calf and correlates with the magnitude of vaccine induced maternal alloantibodies not the MHC haplotype

“…The binding of certain monoclonal antibodies to the B2M-MHC class I heavy chain heterodimer can depend on the associated B2M allele [ 23 ] or MHC class I allele [ 24 ]. Although polymorphisms within the bovine B2M gene are known, none lead to changes in the amino acid sequence [ 25 ].…”

Living with the enemy or uninvited guests: Functional genomics approaches to investigating host resistance or tolerance traits to a protozoan parasite, Theileria annulata, in cattle

“…An additional complexity of cattle DQ, is that DQA and DQB can be duplicated. Thus, it could be that the Sahiwals have duplicated a Specificity of antibody: Anti-DR antibody = J11, anti-DQ␣ antibody = VPM36 and anti-DQ␤ = VPM44 (confirmed as specifically recognising the BoLA equivalents by Russell et al, 2000); anti-class I antibody = IL-A19 (Bensaid et al, 1989). b Mean %.…”

Biochemical evidence of the expression of two major histocompatibility complex class I genes on bovine peripheral blood mononuclear cells

“…The finding that bovine class I products from particular haplotypes are not precipitated by mAb raised against monomorphic determinants on human MHC class I molecules has been demonstrated before (Joosten et al 1988;Oliver et al 1989). The fact that Bensaid et al (1987Bensaid et al ( , 1989 claim that B1.1G6 and W6/32 recognize the same set of p2-microglobulin associated molecules expressed on bovine PBMC must therefore be ascribed to their use of a single heterozygous animal. One explanation for the non-reactivity or low affinity of W6/32 for some of the class I products (variants) could be that substitutions in the a1 or a 2 domain induce a conformational change which could influence the binding of P2-microglobulin to the heavy chain in sych a way that the conformational determinant recognized by W6/32 is masked or even lost.…”