Friend murine leukemia virus (F-MuLV) induces erythroleukemia when inoculated into newborn BALB/c or NIH/Swiss mice. We have molecularly cloned F-MuLV host cell DNA junction fragments from an erythroleukemia cell line induced by F-MuLV to identify cellular genes involved in the leukemogenic process. One particular proviral integration site, Fli-1, is rearranged in 75% (9/12) of independently isolated erythroleukemia cell lines derived from either BALB/c or NIH/Swiss mice inoculated at birth with F-MuLV. Other hematopoietic neoplasms induced by F-MuLV, including myeloid (granulocytic) and lymphoid tumors, did not show rearrangements of the Fli-1 locus. Similarly, none of 35 erythroleukemia cell lines induced by the Friend virus complexes (FV-A and FV-P) was rearranged at the Fli-1 locus. In contrast, no rearrangements were detected at the Sfpi-1 locus, a preferred site of integration in either FV-P- or FV-A-induced leukemias. Using recombinant inbred mice, the Fli-1 locus was situated on mouse chromosome 9 close to the cellular protooncogene c-ets-1. DNA and RNA analysis suggests, however, that Fli-1 is different from ets-1. Thus, Fli-1 appears to define a distinct locus specifically involved in the induction of erythroid leukemias by F-MuLV.