Burkitt's lymphoma is invariably associated with chromosomal translocations that juxtapose the c-myc proto-oncogene with regulatory elements of the immunoglobulin heavy (IgH) or light chain loci resulting in the deregulation of c-myc expression. However, the enhancer elements mediating c-myc deregulation in vivo remain largely unidentified. To investigate the role of the IgH 3' enhancers in c-myc deregulation, we used gene targeting to generate knock-in mice in which four DNase I hypersensitive regions (HS1243) from the murine IgH 3' region were integrated into the 5' region of the c-myc locus. The IgH 3' enhancers induced the upregulation of c-myc expression specifically in B cells of IgH-3'E-myc mice. After approximately 10 months, the mice developed a Burkitt-like B-cell lymphoma with the phenotype of B220+, IgM+ and IgDlow. Analysis of immunoglobulin gene rearrangements indicated that the lymphoma cells were of clonal origin. The presence of a rapidly expanding population of B cells in the spleen and bone marrow of young knock-in mice at 2-4 months of age was observed. Premalignant splenic B cells of knock-in mice showed higher spontaneous and induced apoptosis, however, malignant B cells were more resistant to apoptosis. The p53-ARF-Mdm2 pathway was disabled in half of the lymphomas examined, in most cases through Mdm2 overexpression. Although c-myc expression was increased in premalignant B cells, the promoter shift from P2 to P1 was observed only in malignant B cells. Our studies demonstrate that the IgH 3' enhancers play an important role in c-myc deregulation and B cell lymphomagenesis in vivo.