Cellular distribution of a mixed MHC class II heterodimer between DRα and a chimeric DOβ chain

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Abstract
Human MHC class II antigens include HLA-DR, -DQ, and -DP molecules that present antigens to CD4+ T cells, as well as the non-classical molecules HLA-DM and -DO. HLA-DM promotes peptide binding to class II molecules in endocytic compartments and HLA-DO, which is physically associated with HLA-DM in B lymphocytes, regulates HLA-DM function. Antibodies specific for the DOβ chain were obtained by immunization of mice with a heterodimer consisting of a chimeric DOβ chain (DR/DOβ), containing 18 N-terminal residues of DRβ, paired with the DRα chain and isolated from transfected murine fibroblasts. The specificity of this serum for the DOβ chain and the lysosomal expression of the HLA-DO protein was confirmed using mutant human B cell lines lacking DR or DO molecules. The lysosomal localization of HLA-DO in human B cells contrasts with the cell surface expression of the mixed pair in transfected murine fibroblasts and raises questions concerning the role of the putative targeting motifs in HLA-DO. Transfection of the chimeric DR/DOβ chain along with DRα into human epithelial HeLa cells resulted in high levels of expression of the mixed isotypic pair at the surface of transfectants as well as in lysosomes. The same pattern was observed in HeLa cells transfected with the DOβ chimera and a DRα chain lacking the cytoplasmic tail. Taken together, these results suggest that functional sorting motifs exist in the DOβ chain but that the tight compartmentalization of HLA-DO observed inside B lymphocytes is controlled by the HLA-DOα chain and HLA-DM.