Biochemical characterization of antigen-specific glycosylation-inhibiting factor from antigen-specific suppressor T cells: II. The 55-kDa glycosylation-inhibiting factor peptide is a derivative of TCR α-chain and a subunit of antigen-specific glycosylation-inhibiting factor

We isolated a unique TCR α-chain cDNA derived from the OVA-specific Ts hybridoma, 231F₁. The TCR α-chain consists of Vα11.3, a unique Jα and a complete sequence of Cα region. Transfection of the TCR-α cDNA into a TCR-α^-, TCR-β⁺ T cell line, 175.2, resulted in the expression of TCR-αβ, and the transfectant contained a 35-kDa peptide having the TCR-α-specific antigenic determinant. However, the stable transfectant failed to release a peptide with the TCR-α determinant upon stimulation with anti-CD3. In contrast, overexpression of the cDNA in the 231F₁ cells markedly increased the formation of the 55-kDa peptide, which reacted with both anti-glycosylation-inhibiting factor (GIF) and the mAb H28-710. Definitive evidence for the relationship between the 55-kDa peptide and the TCR α-chain was obtained by transfection of the cDNA of the TCR α-chain with histidine tag into the 231F₁ cells. The 55-kDa GIF peptide formed by stable tranfectants of the TCR-α-tag cDNA bound to Ni⁺-nitrilotriacetic acid-agarose. Upon stimulation with anti-CD3, a stable transfectant of the TCR-α cDNA formed OVA-specific GIF which contained the 55-kDa GIF peptide, and bound not only to anti-TCR-α column but also to anti-TCR-β column. The results indicate that the OVA-specific GIF consists of the TCR-α⁺ 55-kDa GIF and another peptide with TCR-β determinant. It was found that association of the TCR-β⁺ peptide with the 55-kDa GIF is required for binding of the factor to OVA, but not essential for the formation and release of the latter peptide.