NF-KAPPA-B SITES FUNCTION AS POSITIVE REGULATORS OF EXPRESSION OF THETRANSLOCATED C-MYC ALLELE IN BURKITTS-LYMPHOMA

An in vivo footprint over a potential NF-kappa B site in the first exo n of the c-myc gene has been identified on the translocated allele in the Ramos Burkitt's lymphoma cell line. The potential NF-kappa B site in the 5' flanking sequence of c-myc was found to be occupied on the t ranslocated allele ire the Raji Burkitt's cell line. Electrophoretic m obility shift assays with each of these sequences demonstrated complex es with mobilities identical to those of the NF-kappa B site from the kappa light-chain gene. A supershift was obtained with anti-p50 antibo dy with the exon site. The upstream-site shift complex disappeared wit h the addition of anti-p50 antibody. Binding of NF-kappa B proteins to the c-myc exon and upstream sites was demonstrated by induction of bi nding upon differentiation of pre-B 70Z/3 cells to B cells. UV cross-l inking experiments revealed that a protein with a molecular mass of 50 kDa bound to the exon and upstream sites. Transfection experiments wi th Raji cells demonstrated that both sites functioned as positive regu latory regions, with a drop in activity level when either site was mut ated. Access to these sites is blocked in the silent normal c-myc alle le in Burkitt's lymphoma cells, while Rel family proteins bind to thes e sites in the translocated allele. We conclude that the two NF-kappa B sites function as positive regulatory regions for the translocated c -myc gene in Burkitt's lymphoma.