Acute nonlymphocytic leukemia associated with the chromosomal translocation t(6;9)(p23;q34) is an entity that is frequently associated with basophilia, which it shares with chronic myelogenous leukemia. The breakpoint on chromosome 9, q34, appears to be cytogenetically identical in both malignancies and is the site of the cellular oncogene c-abl. We investigated the role of c-abl in cells from two patients with the t(6;9) using in situ chromosomal hybridization, Southern hybridization, and in vitro phosphorylation. We showed that c-abl is not translocated from chromosome 9, resulting in a breakpoint that is on the 3' side of this gene. The t(6;9) translocation does not appear to result in the production of an aberrantly sized protein product or in the acquisition of in vitro tyrosine kinase activity. This is in direct contrast to the findings in chronic myelogenous leukemia, in which c-abl is translocated, leading to the production of a structurally altered c-abl protein with activated tyrosine kinase. Lastly, we demonstrated that the cells of one patient contain sequences from chromosome 9 inserted at the junction of a reciprocal translocation between chromosomes 4 and 10 on the 4q+ chromosome. This insertion, which is at least 100 kilobase pairs in length, represents a duplication and translocation of the protein coding region of c-abl.