A mutant human histocompatibility leukocyte antigen DR molecule associated with invariant chain peptides.
From a human histocompatibility leukocyte antigen (HLA)-DR/DQ hemizygous, B lymphoblastoid progenitor, we isolated a cell line, 10.24.6, with a DR alpha missense mutation (96P-->96S), which results in an N-linked carbohydrate addition at position 94 in the DR alpha 2 domain. Several features of 10.24.6 cells suggest that the mutation disrupts normal intracellular formation of peptide/DR complexes. The mutant HLA-DR dimers, though expressed at the cell surface, lack the conformation of the mature, peptide-loaded class II molecules of the progenitor cell, as assessed by their loss of binding of certain antibodies and by the lack of stability in detergent (sodium dodecyl sulfate) solution. In addition, presentation of endocytosed antigen to HLA-DR-restricted T cells is defective in the mutant, but can be restored by transfection of a wild type DRA gene. Assays with synthetic peptides indicate that the 10.24.6 phenotype is not due to an intrinsic inability of the mutant DR molecules to bind peptides. Therefore, to directly evaluate peptide occupancy of the mutant molecules, we analyzed acid-eluted, HLA-DR-associated peptides. The predominant species from the 10.24.6 mutant is a nested set of invariant chain (Ii)-derived peptides that are undetectable in the DR eluate from progenitor cells. The region of DR alpha altered in the mutant molecules is thus implicated in normal formation of peptide/DR complexes. Further, the same set of Ii peptides associated with the DR molecules is present in the eluate from an antigen presentation mutant with a defect in an major histocompatibility complex (MHC)-linked gene. These results suggest that DR molecules in 10.24.6 and in certain presentation mutants are affected at the same or related steps in class II molecule biosynthesis, raising the possibility that class II molecules interact with an MHC-encoded accessory molecule during antigen presentation.
Mellins
E
E
Department of Pediatrics, University of Pennsylvania, Philadelphia 19104.
Cameron
P
P
Amaya
M
M
Goodman
S
S
Pious
D
D
Smith
L
L
Arp
B
B
D000595:Amino Acid Sequence; D000938:Antigen-Presenting Cells / Q000276:immunology; D000944:Antigens, Differentiation, B-Lymphocyte; D002241:Carbohydrates / Q000737:chemistry; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D017124:Conserved Sequence; D003902:Detergents; D006684:HLA-DR Antigens / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D000949:Histocompatibility Antigens Class II / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000737:chemistry / Q000276:immunology*; D010641:Phenotype; D011487:Protein Conformation; D014162:Transfection
D000944:Antigens, Differentiation, B-Lymphocyte; D002241:Carbohydrates; D003902:Detergents; D006684:HLA-DR Antigens; D000949:Histocompatibility Antigens Class II; D010455:Peptides; C045084:invariant chain