Cathepsin B cleavage and release of invariant chain from MHC class II molecules follow a staged pattern.
A staged pattern of cathepsin B cleavage of MHC class II alpha, beta-bound invariant (Ii) chain and release of fragments was defined. Charge-loss mutations in the Ii chain were created in three clusters of cathepsin B putative cleavage sites R78K80K83K86, K137K143, and R151K154. Products of HLA-DR1 alpha, beta and wild type (WT) or mutant Ii genes, co-transfected into COS1 cells, were cleaved by cathepsin B and immunoprecipitated by antibodies either to MHC class II chains or to different Ii epitopes. In WT Ii, cathepsin B digestion generated two forms of p21 Ii fragments: a p21 recognized by anti-C-terminus antibodies and a p21 recognized by an antibody to a determinant near the N-terminus. C-terminal p21 was released from MHC class II alpha, beta chains upon its formation while N-terminal p21 remained associated with MHC class II alpha, beta chains. Mutations at K137K143 inhibited the generation of N-terminal p21 by cathepsin B. Mutation at R78K80K83K86 led to an accumulation of MHC class II-bound N-terminal p21 without the appearance of MHC class II-bound p14, p10, and p6 fragments after cathepsin B digestion. These results indicate that cathepsin B cleaves wild type Ii first about K137K143 to produce a MHC class II-associated N-terminal p21, which is then cleaved about R78K80K83K86 to generate p14, p10 and finally p6 which still associates with MHC class II alpha, beta chains. This pattern of staged cleavage and release of Ii might be related to a concerted mechanism regulating the binding of antigenic peptides to MHC class II molecules.
Department of Pharmacology, University of Massachusetts Medical School, Worcester 01655.
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Xu
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Capraro
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Daibata
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Reyes
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Humphreys
D000595:Amino Acid Sequence; D000944:Antigens, Differentiation, B-Lymphocyte; D002401:Cathepsin B / Q000378:metabolism*; D002460:Cell Line; D006684:HLA-DR Antigens / Q000235:genetics / Q000378:metabolism*; D000949:Histocompatibility Antigens Class II / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000378:metabolism*; D011233:Precipitin Tests; D013462:Sulfur Radioisotopes; D014162:Transfection
D000944:Antigens, Differentiation, B-Lymphocyte; D006684:HLA-DR Antigens; D000949:Histocompatibility Antigens Class II; D010446:Peptide Fragments; D013462:Sulfur Radioisotopes; C045084:invariant chain; D002401:Cathepsin B