Abstract
The class I major histocompatibility complex (MHC) presents self-developed peptides to specific T cells to induce cytotoxity against infection. The MHC proteins are encoded by multiple loci that express numerous alleles to preserve the variability of the antigen-presenting ability in each species. The mechanism regulating MHC mRNA and protein expression at each locus is difficult to analyze because of the structural and sequence similarities between alleles. In this study, we examined the correlation between the mRNA and surface protein expression of swine leukocyte antigen (SLA)-1*0401 after the stimulation of peripheral blood mononuclear cells (PBMCs) by Staphylococcus aureus superantigen toxic shock syndrome toxin-1 (TSST-1). We prepared a monoclonal antibody (mAb) against a domain composed of Y102, L103 and L109 in the α2 domain. The Hp-16.0 haplotype swine possess only SLA-1*0401, which has the mAb epitope, while other haplotypes possess 0 to 3 SLA classical class I loci with the mAb epitopes. When PBMCs from SLA-1*0401 homozygous pigs were stimulated, the SLA-1*0401 mRNA expression level increased until 24 hrs and decreased at 48 hrs. The kinetics of the interferon regulatory transcription factor-1 (IRF-1) mRNA level were similar to those of the SLA-1*0401 mRNA. However, the surface protein expression level continued to increase until 72 hrs. Similar results were observed in the Hp-10.0 pigs with three mAb epitopes. These results suggest that TSST-1 stimulation induced both mRNA and surface protein expression of class I SLA in the swine PBMCs differentially and that the surface protein level was sustained independently of mRNA regulation.
Highlights
The class I major histocompatibility complex (MHC) antigens are constitutively expressed cellular membrane-bound glycoproteins that associate non-covalently with β-hamicroglobulin (β2M) to present intracellularly processed peptide antigens to T-cell receptors of specific CD8+ T cells [1–3]
Swine swine leukocyte antigen (SLA)-1Ã0401 and β2 microglobulin cDNAs were inserted into N576, an expression vector containing a monomer Venus yellow—green fluorescent protein-reporter gene, and the modified vector was transfected into A20 cells
We selected a specific X2F6 subclone (Fig 1 and S1 Fig) for further use because this clone secreted a monoclonal antibody that reacted with the HEK293 transfectants expressing SLA-1Ã0401 and the monomer Venus yellow—green fluorescent protein (mVenus) fluorescent protein with similar or identical staining patterns
Summary
The class I major histocompatibility complex (MHC) antigens are constitutively expressed cellular membrane-bound glycoproteins that associate non-covalently with β-hamicroglobulin (β2M) to present intracellularly processed peptide antigens to T-cell receptors of specific CD8+ T cells [1–3]. MHC class I proteins are encoded by polymorphic genes at multiple loci, and they act as ligands for killer-cell immunoglobulin-like receptors (KIRs) [4–6]. This polymorphism results in numerous alleles in a population, presumably to preserve the variability of the antigen presenting ability and help the species to defend against various infectious agents, MHC variability may cause autoimmune responses [7–9]. It is important to distinguish between the classical and nonclassical HLA alleles and their regulation at the level of expressed mRNAs and allele-specific surface proteins, as these different classes of MHC molecules have contrary functions. There are relatively few studies on the surface expression of MHC alleles, probably because of the lack of allele-specific monoclonal antibodies due to the similarity of the alleles among the MHC sequences
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