THY-1 TRIGGERS MOUSE THYMOCYTE APOPTOSIS THROUGH A BCL-2-RESISTANT MECHANISM

Programmed cell death plays an important role during thymocyte develop ment, since a vast majority (97%) of mouse cortical thymocytes die in thymus, whereas only 3% of these cells are rescued from cell death and positively selected. Although it seems well established that thymocyt e fate depends upon appropriate surface-expressed T cell receptor, lit tle is known about the molecular mechanism(s) responsible for the mass ive thymocyte elimination that occurs in the thymus. We report here th at Thy-1 is capable of triggering mouse thymocyte death in vitro throu gh a bcl-2-resistant mechanism. We have previously shown that Thy-1 is involved in mouse thymocyte adhesion to thymic stroma through interac tion with an epithelial cell ligand. To examine the Thy-1 signaling fu nction in thymocytes, we have mimicked its interaction with stromal ce lls by culturing mouse thymocytes onto tissue culture plates coated wi th monoclonal antibodies (mAb) directed at distinct Thy-1 epitope regi ons. mAb recognizing determinants in a defined Thy-1 structural domain , but not others, were found to induce marked thymocyte apoptosis as e videnced by morphological and biochemical data. Use of a quantitative DNA dot blot assay indicated that Thy-1-mediated thymocyte apoptosis w as not blocked by RNA or protein synthesis inhibitors, EGTA, or by cyc losporin A, and differed, therefore, from ''activation-driven cell dea th''. Moreover, Thy-1(+)-transfected, but not wild-type AKR1 (Thy-1(-) d) thymoma cells underwent apoptosis after ligation with apoptosis-ind ucing, Thy-1-specific mAb. In contrast to thymocytes, the latter event was inhibitable by RNA and protein synthesis inhibitors, an indicatio n that thymocytes, but not thymoma cells, contain the molecular compon ents necessary for Thy-1-driven apoptosis. We further showed that Thy- 1-triggered thymocyte death is a developmentally regulated process ope rative in fetal thymocytes from day 17 of gestation, but not in periph eral T cells. Indeed, the target of apoptosis by anti-Thy-1 was found to reside mainly within the CD4(+)8(+)3(-) and CD4(+)8(+)3(lo) double positive immature thymocyte subsets. Finally, it is of major interest that Thy-1-mediated apoptosis, which was found to be readily detectabl e in thymocytes from bcl-2-transgenic mice, represents a thus far uniq ue experimental system for studying bcl-2-resistant thymocyte death me chanism(s).