Identification of R2BP as a Pax-5.

(A) Putative binding sites of transcription factors in the RAG-2 promoter. The asterisk denotes a nucleotide identical with consensus binding sequences (c-Myb, 5′-GNCNGTT-3′; Ikaros, 5′-^T/_CGGGA^A/_T-3′; Pax-5, 5′-CTTG^A/_GTCAA^A/_TGCAG^T/_CG^T/_G^G/_AAC^G/_C^G/_ATAGC-3′27), and the minus sign shows a nucleotide nonidentical with consensus binding sequences. (B) Competition of R2BP binding with the consensus sequence of Pax-5. EMSA was performed as in Figure 4 using nuclear extract prepared from 18.8.1 cells and the −80/−17 nt RAG-2 promoter region as a probe. The consensus binding sequence for the c-Myb, Ikaros, Pax-5, or Pax-5 binding sequence mutant (Pax-5m) was used as a competitor. Histograms indicate the relative density of the C2 complex shown on the bottom. The density of the C2 complex in the absence of the competitor is denoted as 100%. (C) The effect of anti–Pax-5 antibody on EMSA. EMSA was performed as described above in the absence or the presence of anti–Pax-5 or anti–c-Myb antibody. The asterisk denotes the band shifted with the anti–Pax-5 antibody.