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. Author manuscript; available in PMC: 2008 Jul 1.
Published in final edited form as: Immunity. 2007 Jul 19;27(1):89–99. doi: 10.1016/j.immuni.2007.05.021

Figure 5. The upstream Gata3 promoter contains a bona fide conserved RBP-J binding element.

Figure 5

(a) diagram of the Gata3 locus showing upstream and downstream Gata3 promoters each driving expression of a unique exon 1, which is spliced to the common exon 2. Underneath, the consensus RBP-J binding element is shown and its conservation between mouse and human. (b) EMSA using radiolabeled double strands probe containing the RBPG3P1a and lysates from CHO cells transfected with vector alone (1st lane), RBP-J (2nd lane and 4th to 10th lane) or RBP-J R218H (3rd lane). A probe carrying a point mutation mRBPG3P1a was used as specificity control (10th lane). The specific RBP-J-RBPG3P1a complex was competed away by addition of (10-fold or 3-fold) excess unlabeled RBPG3P1a (lanes 4 and 5) or the previously described RBP-J binding element RE (lanes 8 and 9), but not by the mutant probe mRBPG3P1a (lanes 6 and 7). (c) EMSA using lysates from CD4+ T cells (lanes 2 to 5) and radiolabeled RBPG3P1a, (lanes 1,2 and 4,5) or mRBPG3P1a(lane 3). Antibody to RBP-J (lane 5) or control mouse Ig (lane 4) was added to the reaction to supershift the RBP-J- RBPG3P1a complex.