Abstract
Thymocyte β-selection and positive-selection depend on TCR signaling via the immune adaptors SLP-76 and LAT. Gads bridges the recruitment of SLP-76 to LAT, yet is not required for the maturation of single positive (SP) thymocytes. To illuminate this paradox, we performed tamoxifen-induced ablation of Gads (GadsiKO), accompanied by the expression of tdTomato, and compared the development of Gads-expressing (Tom-) and Gads-ablated (Tom+) thymocytes within the same mouse. GadsiKO (Tom+) thymocytes exhibited impaired β- and positive-selection, yet δ-selection was not affected. While susceptible to apoptosis ex vivo, the marked accumulation of self-MHC nonresponding (CD5-) GadsiKO DP thymocytes suggested the possibility of impaired death by neglect in situ. Further supporting this notion, GadsiKO CD5lo DP thymocytes exhibited reduced apoptosis in situ and reduced CD8-induced apoptosis ex vivo. Most GadsiKO CD4 SP thymocytes were positively selected, yet a distinct population of unselected (CD5- TCRβneg/low CCR7lo CD24hi) CD4 SP thymocytes was seen only in the absence of Gads. This unselected population did not include Treg or TCRγδ subsets; rather, it encompassed CD44lo CD25+ cells, resembling pre-β-selection thymocytes. Our results suggest that Gads promotes passage through key TCR-driven developmental checkpoints while repressing the progression of unselected DN and DP thymocytes.