T cell clones specific for hybrid I-A molecules. Discrimination with monoclonal anti-I-A (k) antibodies

Abstract

Alloreactive and soluble antigen-reactive, I-A-restricted T cell clones were examined for their ability to recognize hybrid I-A antigens. Several clones that recognized hybrid I-A(b)/I-A(k) molecules on (C57BL/6 x A/J)F(1) [(B6A)F(1)] spleen cells were studied. We were able to distinguish clones that recognized hybrid I-A molecules of the A(b)(a)A(k)(β) type from those that recognized A(k)(a)A(b)(β) molecules. We reached this conclusion by considering data from three independent types of experiments. (a) Monoclonal antibodies were used to inhibit T cell stimulation. Antibodies 10.2.16 and H116.32 distinguished two mutually exclusive "families" of T cell clones. One group of clones was inhibited by 10-2.16 and not H116.32, the other group exhibited reciprocal inhibition. (b) T cell proliferation was assayed using antigen-presenting cells from B6.C-H-2(bml2) (bml2) and [bml2 × B10.A(4R)]F(1) mice. Because the bml2 strain has a mutation that results in an altered A(b)(β) polypeptide chain (A(bm12)(β)), we reasoned that clones that could recognize the [bm12 × B 10.A(4R)]F(1) cells were recognizing A(b)(a)A(k)(β) molecules. Alternatively, clones not recognizing [bml2 × B10.A(4R)]F(1) cells had specificity for A(k)(a)A(b)(β) molecules. (c) I-A molecules immunoprecipitated from radiolabeled (B6A)F(1) splenocyte extracts were analyzed by two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These experiments confirmed an earlier report that antibody 10.2.16 recognized determinants on the A(k)(β) chain (12). Antibody H116.32 immunoprecipitated products consistent with recognition of A(k)(a) determinants. Taken together, these three types of results offer conclusive evidence that T cell clones recognizing "hybrid" I-A molecules use either A(b(k)A(k)(β) or A(k)(a)A(b)(β) molecules as recognition or restriction sites. Clones whose proliferation was supported by [bm 12 x B10.A(4R)]F(1) cells and blocked by anti-I-A(k) antibody 10-2.16 recognized A(b)(a)A(k)(β) B molecules. Clones that were blocked by antibody H116.32 and did not recognize [bml2 X B10.A(4R)]F(1) cells use a recognition site(s) on A(b)(a)A(k)(β) molecules. Thus, we can demonstrate both functionally and biochemically that hybrid F(1) I-A molecules of the structure A(k)(a)A(b)(β) and A(b)(a)A(k)(β) both exist on (B6A)F(1) splenocytes and that both configurations are used in immune recognition phenomena.

Loss in CD4 T‐cell responses to multiple epitopes in influenza due to expression of one additional MHC class II molecule in the host

“…Interestingly, responses to most of the HA-specific peptide epitopes, including major epitopes such as HA 22, were decreased to < 20% of that seen in the parental strain. Responses to epitopes derived from other influenza proteins were also significantly diminished, including the responses to NP 16 If one MHC class II molecule was highly favoured during thymic selection or peripheral CD4 T-cell activation, CD4 T-cell responses restricted to that MHC molecule might dominate the F 1 response. Although responses restricted to both the I-A b and I-A s molecules could be identified ( Fig.…”

Encephalitogenic T cell clones specific for myelin basic protein. An unusual bias in antigen recognition.

“…Ia Restriction Pattern of(PLSJ)FI Clones. Using mAb directed to specific I-A and I-E gene products, we have determined the class II molecule used by each clone for recognition of MBP (24,25). Clones in pattern I are inhibited from proliferating by mAb binding to I-A", (Table 1V).…”

Multiple functional sites on a single Ia molecule defined using T cell clones and antibodies with chain-determined specificity.

“…Blocking Assay of Cloned T Cell Proliferation. T cell clone proliferation assays and assays of T cell blocking of proliferation by anti-I-A k mAb in vitro were described previously (14). Briefly, 104 cloned T cells were cultured with 106 irradiated spleen cells (APC) and an appropriate dose of antigen.…”

“…Two-dimensional Get Electrophoresis. Methods of analysis of Ia molecules by two-dimensional gel electrophoresis (2-D gels) were as described previously (14). Spleen cells of strains C57BL6 × A/J (B6A)F1 or C57BL10 × BI0.A(4R)F1 mice were biosynthetically radiolabeled with pS]methionine and solubilized in buffer containing 0.5% Triton X-100.…”

“…The FI specific T cell clones are blocked by antibody 10-2.16 or H 116-32, but never both. Since previous biochemical studies showed that 10-2.16 reacts with A~ k (14,27,30) and H116-32 reacts with A~ k (14), it follows that FI restricted T cell clones that are blocked by 10-2. ability of a number of other anti-I-A k mAb to inhibit T cell proliferation utilizing clones that were blocked by either 10-2.16 or H116-32. These clones then act as defined reagents to characterize the reactivity of the new mAb for a or /3 chains.…”