The BCL6 gene, mapped at the chromosomal band 3q27, encodes a POZ/Zinc finger transcription repressor protein. It is frequently activated in Non-Hodgkin's lymphomas (NHL) by translocations with breakpoints clustering in the 5' major breakpoint region (MBR) as well as by mutations in the same region. The translocations lead to BCL6 activation by substitution of promoters of rearranging genes derived from the reciprocal chromosomal partners such as IG. We report the molecular genetic analysis of a novel t(2;3)(q21;q27) translocation subset in NHL comprising three cases without apparent BCL6 involvement in the translocation. Southern blot analysis of tumor DNAs utilizing BCL6 MBR probes revealed no rearrangement in two cases. Two rearranged bands in the third case resulted from a deletion in one allele and a mutation in the other allele. Southern blot analysis of DNA from one of the two tumors without BCL6 rearrangement, using a probe derived from the recently identified alternative breakpoint region (ABR), showed a rearrangement. The ABR is located 200-270 kb telomeric to MBR. Mutations were identified in the previously reported hypermutable region of BCL6 in all three tumors. In one, the mutant allele alone was found to be expressed by RT-PCR analysis of RNA. These results demonstrate the presence of 3q27 translocation breakpoints at a distance from BCL6 suggesting distant breaks that deregulate the gene or involvement of other genes that may be subject to rearrangement.