Developing T cells undergo distinct selection processes that determine the TCR repertoire. Positive selection involves the differentiation of immature thymocytes capable of recognizing antigens complexed with self-MHC molecules to mature T cells. Besides the central role of TCR engagement by MHC in triggering selection; the interaction of CD8 and CD4 with MHC class I and class II, respectively; is thought to be important in regulating the selection process. To study potential mechanisms involved in positive selection of CD8+ cells, we have analyzed mice expressing a unique transgenic TCR. The transgenic receptor recognizes the HY male Ag in the context of the MHC class I molecule, H2-Db. We describe that CD8 and the TCR are selectively associated in thymocytes of mice expressing the restricting MHC, but not in thymocytes of mice expressing a nonrestricting MHC. pp56lck and pp59fyn, the tyrosine kinases associated with CD8 and TCR, respectively, were found to be present in this complex in an activated form. No comparable TCR-CD4 complex formation was found in thymuses undergoing positive selection to CD8+ cells. The formation of a multimolecular complex between CD8 and TCR, in which pp56lck and pp59fyn are activated, may initiate specific signaling programs involved in the maturation of CD8+ cells.