Survival and Developmental Progression of Unselected Thymocytes in the Absence of the T-Cell Adaptor Gads

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 (Gads^iKO), accompanied by the expression of tdTomato, and compared the development of Gads-expressing (Tom⁻) and Gads-ablated (Tom⁺) thymocytes within the same mouse. Gads^iKO (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⁻) Gads^iKO DP thymocytes suggested the possibility of impaired death by neglect in situ. Further supporting this notion, Gads^iKO CD5^lo DP thymocytes exhibited reduced apoptosis in situ and reduced CD8-induced apoptosis ex vivo. Most Gads^iKO CD4 SP thymocytes were positively selected, yet a distinct population of unselected (CD5⁻ TCRβ^neg/low CCR7^lo CD24^hi) CD4 SP thymocytes was seen only in the absence of Gads. This unselected population did not include Treg or TCRγδ subsets; rather, it encompassed CD44^lo 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.