Importance of thioredoxin in the proteolysis of an immunoglobulin G as antigen by lysosomal Cys-proteases.
For disulphide-bonded antigens, reduction has been postulated to be a prerequisite for proteolytic antigen processing, with subsequent production of major histocompatibility complex (MHC) class II binding fragments. The murine monoclonal immunoglobulin G (IgG) CE25/B7 was used as a multimeric antigen in a human model. Native IgG is highly resistant to proteolysis and has been previously found to be partially reduced at early steps of cell processing to become a suitable substrate for endopeptidases. The role of the oxidoreductase thioredoxin (Trx) was assessed in the reduction of the IgG by cleavage of H-L and H-H disulphide bonds. Recombinant human Trx (rTrx) has been assayed in a proteolytic in vitro system on IgG using endosomal and lysosomal subcellular fractions from B lymphoblastoid cells. rTrx is required in a dose-dependent manner for development of efficient proteolysis, catalysed by thiol-dependent Cys-proteases, such as cathepsin B. We demonstrated that cathepsin B activity was stimulated by the addition of rTrx. Thus, we propose that Trx-dependent IgG proteolysis occurred, on the one hand by means of the unfolding of the IgG after disulphide reduction, becoming a substrate of lysosomal proteases, and on the other hand by Cys-proteases such as cathepsin B that are fully active upon the regeneration of their activity by hydrogen donors.
Kerblat
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CEA Laboratoire d'Immunochimie, INSERM U238, Grenoble, France.
Drouet
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C
Chesne
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Marche
P N
PN
D002401:Cathepsin B / Q000378:metabolism; D018929:Cell Culture Techniques; D002458:Cell Fractionation; D003546:Cysteine Endopeptidases / Q000276:immunology*; D004306:Dose-Response Relationship, Immunologic; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology*; D008247:Lysosomes / Q000276:immunology; D010084:Oxidation-Reduction; D013879:Thioredoxins / Q000276:immunology*; D014407:Tumor Cells, Cultured
D007074:Immunoglobulin G; D013879:Thioredoxins; D003546:Cysteine Endopeptidases; D002401:Cathepsin B