Cellular distribution of a mixed MHC class II heterodimer between DRalpha and a chimeric DObeta chain.
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 DObeta chain were obtained by immunization of mice with a heterodimer consisting of a chimeric DObeta chain (DR/DObeta), containing 18 N-terminal residues of DRbeta, paired with the DRalpha chain and isolated from transfected murine fibroblasts. The specificity of this serum for the DObeta 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/DObeta chain along with DRalpha 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 DObeta chimera and a DRa chain lacking the cytoplasmic tail. Taken together, these results suggest that functional sorting motifs exist in the DObeta chain but that the tight compartmentalization of HLA-DO observed inside B lymphocytes is controlled by the HLA-DOalpha chain and HLA-DM.
Laboratory of Immunogenetics, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rockville, MD 20852, USA.
A
A
Samaan
J
J
Thibodeau
W
W
Mahana
F
F
Castellino
P A
PA
Cazenave
T J
TJ
Kindt
D000818:Animals; D000918:Antibody Specificity; D001402:B-Lymphocytes / Q000276:immunology*; D002451:Cell Compartmentation; D002458:Cell Fractionation; D002460:Cell Line; D019281:Dimerization; D004705:Endocytosis; D005434:Flow Cytometry; D006681:HLA-D Antigens / Q000276:immunology / Q000302:isolation & purification*; D006684:HLA-DR Antigens / Q000302:isolation & purification*; D006367:HeLa Cells; D000949:Histocompatibility Antigens Class II; D006801:Humans; D008247:Lysosomes; D008285:Major Histocompatibility Complex; D051379:Mice; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification
C416186:H-2O antigen; D006681:HLA-D Antigens; C046148:HLA-DO antigens; D006684:HLA-DR Antigens; D000949:Histocompatibility Antigens Class II; D011993:Recombinant Fusion Proteins