Fig 2.

BCL-6 exon 1 contains two functional BCL-6 binding sites that are targeted by activating mutations in lymphomas. (A) Sequences surrounding the transcriptional start site of the major promoter (Pb) (2), showing the locations of the two BCL-6 binding sites (SRB and SR20), and the four activating mutations from three lymphoma cases 1682, 1652, and Ly1. The two alleles of 1682 each carry a mutation in this region. Arrows underlie the 9-bp core sequences of the motifs. The consensus B6BS is also given for comparison. (B) Gel shift analysis of the SRB and SR20 motifs. The ³²P-labeled B6BS probe was used with nuclear extract from the BCL-6-positive Ly1 cells. Specificity of the major complex, indicated by an arrow, was verified by supershift assay with the anti-BCL-6 serum, 73-6. An excess amount of cold B6BS, SRB, and SR20 oligos was used to assess their relative affinity for BCL-6. (C) Reporter assay of Pab-Luc construct in 3T3 cells. Pab-Luc and Pab-dm1-Luc were transfected with increasing amounts of a BCL-6 expression plasmid. (D and E) Gel shift analysis of the mutated SRB and SR20 sites. Labels are used as described for B.