Virtually all mature T cells are CD4(+)CD8(-) or CD4(-)CD8(+) and this not only is their most important surface-phenotype distinction but also has crucial functional consequences for the entire immune response. Both subsets arise from double-positive thymocytes, and much has been learned about the molecular events that govern this lineage bifurcation process. As detailed in this review, the signaling pathways and specific molecules that control this process are now being discovered. In particular, the transcription factors ThPOK (T-helper inducing POZ-Kruppel factor) and Runx3 have emerged as the crucial regulators of helper lineage commitment and the cytotoxic lineage, respectively. This article describes their antagonistic interaction that is an important mechanism of the lineage specification, as well as the hierarchy and importance of several other transcription factors and cytokine signals in the network of pathways that govern thymocyte helper/cytotoxic lineage commitment.