CD2 Is Specifically Involved In CD3e-mediated T Cell Receptor Trafficking: Combined Modulation Of CD2 And CD3e Impairs TCR Recovery After Stimulation

O Yang; L Qin; Y Ding and JS Bromberg, Departments of Surgery, Microbiology and Immunology, University of Michigan, Ann Arbor, Michigan

Ligation of the TCR/CD3 complex by alternate ligands can result in distinct patterns of receptor trafficking. In this study, we investigated the effect of CD2 co-receptor ligation on the mechanism of T cell receptor trafficking induced by ligation of TCRß and CD3e.

Methods: Nylon wool purified native splenic murine T lymphocytes were incubated with anti-TCRß or anti-CD3e in culture and assayed for TCRß, CD3e and CDz expression by fluorescence flow cytometry and immunoblotting. In parallel, control and treated cells were washed, re-stimulated and measured for secondary proliferation.

Results: Anti-TCRß had no effect on the surface expression of TCRß itself, but caused moderate yet prolonged down-modulation of CD3e and CD3z. Conversely, anti-CD3e substantially down-modulated TCRß, CD3e and CD3z after 24 hours of culture, and these subunits were all re-expressed after 72 hours. No alteration in TCR/CD3 expression was observed with ligation of CD2 alone, and anti-CD2 plus anti-TCRß did not differ from anti-TCRß alone. However, combined ligation of both CD2 and CD3e resulted in TCR/CD3 down modulation after one day of culture and significantly prohibited the reexpression of TCR/CD3 surface markers after 3 days. In correlation, cells treated with anti-CD2 plus anti-CD3e were anergized and their proliferation was markedly reduced in response to secondary stimulation.

Conclusions: 1) Physical dissociation between TCRß and the CD3 complex takes place in response to TCR ligation and is maintained over several days. 2) Ligation of CD2 specifically changes the impact of CD3e-mediated modulation by delaying TCR recovery and inhibiting secondary proliferation. The effect of CD2 ligation is likely related to the molecular mechanism by which combined anti-CD2 plus anti-CD3e mAbs induce antigen specific tolerance in vivo.