Chromosome band 11q23 is the site of recurring translocations with a variety of partner chromosomes in myeloid and lymphoid acute leukemias, infant leukemias, and treatment-induced secondary acute myelogenous leukemia. The translocation breakpoints cluster in a restricted region of the HRX gene resulting in fusion genes that encode an N-terminal portion of Hrx fused to various partner proteins. We have characterized the transcriptional transactivation properties of Enl, a protein that is fused to Hrx in t(11;19) leukemias. Enl is a nuclear protein that is capable of activating transcription from synthetic reporter genes in both lymphoid and myeloid cells, as well as in yeast. Deletion mutagenesis demonstrated that the minimal portion of Enl required for activation of transcription was localized to its C-terminal 90 amino acids. This region is highly conserved between Enl and the t(9;11) fusion partner Af-9 and is retained in all Hrx-Enl and Hrx-Af9 fusion proteins. Thus, the leukemogenic contribution and transcriptional activation potential of Enl colocalize to its highly conserved carboxy terminus, suggesting that Hrx-Enl chimeric proteins mediate alterations in the transcription program of t(11;19)-bearing cells.