Molecular events involved in specification of early hematopoietic system are not well known. In Xenopus, a paired-box homeodomain family (Mix.1-4) has been implicated in this process. Although Mix-like homeobox genes have been isolated from chicken (CMIX) and mice (Mml/MIXL1), isolation of a human Mix-like gene has remained elusive. We have recently isolated and characterized a novel human Mix-like homeobox gene with a predicted open reading frame of 232 amino acids designated the Mix.1 homeobox (Xenopus laevis)-like gene (MIXL). The overall identity of this novel protein to CMIX and Mml/MIXL1 is 41% and 69%, respectively. However, the identity in the homeodomain is 66% to that of Xenopus Mix.1, 79% to that of CMIX, and 94% to that of Mml/MIXL1. In normal hematopoiesis, MIXL expression appears to be restricted to immature B- and T-lymphoid cells. Several acute leukemic cell lines of B, T, and myeloid lineage express MIXL suggesting a survival/block in differentiation advantage. Furthermore, Xenopus animal cap assay revealed that MIXL could induce expression of the alpha-globin gene, suggesting a functional conservation of the homeodomain. Isolation of the MIXL gene is the first step toward understanding novel regulatory circuits in early hematopoietic differentiation and malignant transformation.