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. 2004 Nov;24(21):9601–9609. doi: 10.1128/MCB.24.21.9601-9609.2004

FIG. 3.

FIG. 3.

FoxA proteins bind directly to site 1 and site 2 in the E1D fragment in vitro. (A) EMSAs were performed using the E1D-a fragment (site 1) as a radiolabeled probe with no extract (free probe [F.P.], lane 1), Hep3B nuclear extracts (lanes 2 to 8), or nuclear extracts from HeLa cells that were transfected with an EV (lane 9) or expression vectors for FoxA1 (lane 10) or FoxA2 (lane 11). Cold competitions were performed with the Hep3B extracts by using a 100-fold molar excess of either no competitor (“-, ” lane 2), E1D-a fragment (self, lane 4), the E1D-g fragment (lane 5), or the albumin promoter C/EBP site (lane 6). Supershift experiments were performed with antibodies to FoxA2 (lane 7) or C/EBPδ (lane 8). (B) EMSAs performed using the E1D-f fragment (site 2) as the radiolabeled probe. Cold competitions and supershift experiments were performed as described for panel A. (C) EMSA with the TTR-FoxA site (TTR-FoxA) as a radiolabeled probe and nuclear extracts from HeLa cells transfected with a control EV (lane 2), FoxA1 expression vector (lane 3), or FoxA2 expression vector (lane 4) demonstrating equal amounts of FoxA1 and FoxA2 proteins in the HeLa nuclear extracts. Lane 1, free probe (F.P.).