Antigen entry into early endosomes is insufficient for MHC class II processing.
Helper T-cell recognition of Ag requires that the Ag be processed and presented by class II-expressing Ag-presenting cells. Processing involves the introduction of Ag into acidic compartments where proteolysis occurs producing peptides that bind to the class II molecules. Although Ag can enter the processing pathway through fluid phase pinocytosis, Ag processing can be made over 1000-fold more efficient by binding the Ag to a variety of Ag-presenting cell surface structures. The increased efficiency in processing is presumably the result of the ability of such structures to deliver the bound Ag to compartments involved in processing. Here we report that Ag bound to the transferrin receptor (TfR), which cycles predominantly through early endosomal compartments, does not enter the processing pathway. We found that cytochrome c(c)covalently coupled to monovalent iron-saturated transferrin (Tf), (c-Tf), is not processed or presented significantly better than unconjugated c, indicating that the majority of cycling TfR does not enter compartments where processing proceeds. The conjugation of Tf to c does not affect its binding to the TfR, as the binding is both saturable and compatible with unmodified Tf. Moreover, c-Tf and unmodified Tf cycle equivalently with a t1/2 of internalization of 3 to 5 min and are released outside the cell with little detectable degradation. Significantly, we found that c-Tf is efficiently processed and presented when the TfR is cross-linked, altering its normal cycling. Indeed, c covalently coupled to polymerized Tf is presented at 1/100th the concentration of c alone. Cross-linking of c-Tf bound to the TfR using c-specific antibodies also results in efficient processing and presentation. Thus, the endosomal compartments through which Tf normally cycles are not sites of processing, whereas compartments into which cross-linked Tf is diverted allow efficient processing and presentation of Ag.
Department of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, Evanston, IL 60208.
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Niebling
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Pierce
D000818:Animals; D000938:Antigen-Presenting Cells / Q000378:metabolism*; D002451:Cell Compartmentation / Q000276:immunology*; D010856:Columbidae; D003432:Cross-Linking Reagents; D003574:Cytochrome c Group / Q000276:immunology; D004331:Drosophila melanogaster; D005260:Female; D000949:Histocompatibility Antigens Class II / Q000378:metabolism*; D016393:Lymphoma, B-Cell; D008247:Lysosomes / Q000378:metabolism; D046911:Macromolecular Substances; D051379:Mice; D008808:Mice, Inbred CBA; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism; D011990:Receptors, Transferrin / Q000378:metabolism; D014168:Transferrin / Q000276:immunology* / Q000378:metabolism; D014407:Tumor Cells, Cultured
D003432:Cross-Linking Reagents; D003574:Cytochrome c Group; D000949:Histocompatibility Antigens Class II; D046911:Macromolecular Substances; D011948:Receptors, Antigen, T-Cell; D011990:Receptors, Transferrin; D014168:Transferrin