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. 2009 Jun 1;119(7):1888–1898. doi: 10.1172/JCI37028

Figure 5. Pdx1 regulates Ngn3, at least in part, via its direct occupancy of a conserved upstream enhancer of Ngn3.

Figure 5

(A) Ngn3 mRNA levels in total pancreas of E13.5 animals (n = 5; **P = 0.005). (B) Expression of Ngn3 in PDCs transduced with pBABE or pBABE-Pdx1 retroviruses. Pdx1 and Ngn3 mRNA levels are expressed compared with empty vector (n = 6; ‡P < 0.007). (C) Colocalization of Pdx1 and Ngn3 during pancreas development. Double immunofluorescence for Ngn3 (green) and Pdx1 (red) in wild-type pancreata. Arrows indicate double-positive cells and arrowheads mark single-positive Ngn3 cells. Scale bar: 10 μm. (D) Diagram depicting conserved TT/AAT sites in the mouse Ngn3 promoter (black and red boxes). The region homologous to the previously described human Ngn3 Cluster 1 enhancer that contains binding sites for Hnf1, Foxa2, and Hnf6 is indicated in red. (E) ChIP with Pdx1 antisera performed on at least 65 pooled wild-type E13.5 pancreata per experiment (n = 3; *P < 0.05). (F) Promoter reporter assays were performed in HepG2 cells. Cells were transfected with expression vectors for Pdx1, Pdx1ΔC or empty vector, and the promoter reporter Ngn3(–3379 to –3227)-tkluc (n = 3; ††P < 0.05 versus empty vector, †P < 0.05 versus full-length Pdx1).