Figure 2. High sensitivity of TCRβ1⁺ TCR for MOG_35-55.

Seven TCRα chains derived from non-TCRβ1 TCR were cloned into the MSCV-I-GFP retroviral vector with TCRβ1. CD4⁺ 4G4 TCRαβ⁻ hybridoma cells were transduced and TCRαβ⁺ cells were sorted and stimulated with 100 μg/ml MOG_35-55 or anti-CD3. IL-2 production was measured by ELISA at 24 h. (A) TCRα chains that conferred MOG_35-55 responsiveness when paired with TCRβ1 and (B) TCRα chains that did not confer MOG_35-55 responsiveness. Graphs are labeled with the TCRα name (top), TRAV and TRAJ segment use (middle), and the CDR3α sequence (bottom). (C) Five TCRα chains isolated from CNS-infiltrating TCRβ1⁺ TCR were linked to TCRβ1 in retroviral constructs as above (TCRβ1-342α, TCRβ1-307α, TCRβ1-308α, TCRβ1-323α, and TCRβ1-355α). Alternatively, retroviral constructs incorporating six private MOG_35-55 -reactive TCRαβ (1MOG213, 1MOG203, 1MOG202, 5MOG113, 1MOG9, and 1MOG23) were similarly generated. Constructs were transduced into CD4⁺ 4G4 TCRαβ⁻ hybridoma cells and sorted for GFP-positivity and similar levels of TCR. Cell lines were stimulated with the indicated concentration of MOG_35-55 or anti-CD3, and IL-2 production was measured at 24 h by ELISA. Private (non-TCRβ1) TCR are shown in the left panel and CNS-infiltrating TCRβ1⁺ TCR are shown in the right panel. Private TCR 1MOG9 is included in both panels to illustrate enhanced sensitivity of TCRβ1⁺ TCR relative to those from private MOG-reactive TCR. Data are representative of at least 3 independent experiments performed in duplicate.