Role of thiols in the presentation of a snake toxin to murine T cells.
We isolated and characterized two T hybridomas specific for a highly stable snake toxic protein. One hybridoma, called T1C9, is I-E(d)-restricted and stimulated by both the native and reduced and carboxymethylated (RCM) toxins and by synthetic fragments containing the region 24-36. The other hybridoma, called T1B2, is I-A(d)-restricted and stimulated by the native toxin, only. Neither the RCM toxin nor any of the initial synthetic peptides used in our study could stimulate it. We show that this lack of effect is associated with the presence, in the epitope-containing fragment, of irreversible blocking groups on cysteine residues. Indeed, when the fragment 32-49 has its cysteines involved in either intra-(32-49SS) or mixed disulfides, a stimulation of T1B2 was observed. Fixed APC do not present native toxin to either hybridomas but present RCM toxin to T1C9. Strikingly, fixed APC present the peptide 32-49SS to T1B2; however, we show that this is possible only because the peptide disulfide is reduced. The thiol dependence of this epitope suggests that the native toxin can stimulate T1B2 only after disulfide reduction. This reaction may constitute a major step during the processing of the toxin and more generally of any disulfide-containing Ag.
Maillère
B
B
Département d'Ingénierie et d'Etudes des Protéines, CEA-C.E. Saclay, Gif-sur-Yvette, France.
Cotton
J
J
Mourier
G
G
Léonetti
M
M
Leroy
S
S
Ménez
A
A
D000595:Amino Acid Sequence; D000818:Animals; D000938:Antigen-Presenting Cells / Q000502:physiology; D003039:Cobra Neurotoxin Proteins / Q000276:immunology*; D000939:Epitopes; D005260:Female; D006825:Hybridomas / Q000276:immunology; D008213:Lymphocyte Activation; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000276:immunology; D013438:Sulfhydryl Compounds / Q000502:physiology*; D013601:T-Lymphocytes / Q000276:immunology*
D003039:Cobra Neurotoxin Proteins; D000939:Epitopes; D010446:Peptide Fragments; D013438:Sulfhydryl Compounds; C025154:alpha-cobratoxin