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While studies have indicated a role for TCR clustering in effector functions, there has been no analysis of whether T cell receptor organization and subsequent changes in the TCR avidity are modulated in different physiologic states. Here we examined the influence of activation on the ability of T cells to bind dimeric peptide MHC complexes. We use dimeric complexes as an idealized representation of an antigen presenting cell. Using these complexes in a fluorescence based binding assay, we found that increased binding of peptide/MHC dimers to activated T cells compared to naive cells was apparent at both 4C and 37C. The TCR, on activated T cells, was readily crosslinked by soluble dimeric MHC I molecules, increasing the overall binding affinity (avidity) of the TCR for peptide-MHC complex.

The increased number of crosslinks observed on activated T cells provided a means for enhancing there sensitivity to low density of antigen. To block this TCR reorganization we used anti-TCR mAb that sterically separates the TCRs without affecting its antigen recognition site. Furthermore, we showed that TCR reorganization could be affected by changes in the cholesterol content of the cellmembrane. Increasing the cholesterol content of the membrane was shown to induce early activation signals.

In addition to the TCR the role of CD8 in TCR reorganization on the cell surface was investigated. We found that in naive T cells, the presence of CD8 stabilized the interaction of SIYKb-Ig to 2C T cells but not the interaction of QL9Ld-Ig with 2C T cells. Second, while the equilibrium binding profiles of QL9Ld-Ig was CD8-independent on naive 2C T cells the binding kinetics were very different. Finally, preliminary data showed that the binding of QL9Ld-Ig on activated 2C T cells is CD8-dependent showing higher avidities than for activatedCD8-T cells. These results highlighted the physiological state of the T cell as a variable in the organization and the function of CD8 as well as the TCR.