Differentiation of CD4+ T cells into T helper (Th) 1 or Th2 cells requires the cytokines interleukin (IL)-12 and IL-4, respectively. However, transcription factors that regulate expression of Th1 or Th2 cell-specific genes remain largely unclear. In the present study, a new Th1-specific transcription factor, named Tbt-1 (T-box protein expressed in T lymphocytes), was identified. Tbt-1 is a novel member of the T-box family, which is characterized by a conserved T-box DNA-binding domain. Unlike other known T-box proteins that regulate embryo development and organogenesis, Tbt-1 expression is restricted to adult lymphoid organs. Tbt-1 mRNA is only detected in peripheral lymphoid tissues such as spleen, lymph nodes, and blood leukocytes, but not in thymus or bone marrow. Tbt-1 mRNA is not detected in resting T cells but is strongly induced in differentiating Thl cells and CD8+ cytotoxic effector cells. In contrast, Tbt-1 expression was not observed in the entire process of Th2 cell differentiation. In addition, phylogenetic analyses indicate that Tbt-1 co-evolved with adaptive immune responses. Thus, Tbt-1 is the first T-box transcription factor shown to be specific for Th1 cell differentiation.