HGF-induced Hnf4a binds to Foxa2 in Mgp^−/− lungs. (a) Expression of Hnf4a, Nkx2.1, and Foxa2 was determined in lungs by real-time PCR (n = 5). Liver of WT mice was used as a control. (b) Protein interactions in lung lysates were analyzed by immunoprecipitation (IP), followed by immunoblotting (IB) with antibodies as indicated (n = 6). Lysates from WT Mgp^+/+ liver were used as a positive control. (c–f) Hnf4a and Foxa2 bind to the albumin locus at the Hnf4a-binding site and alter the albumin transcription state in Mgp^−/− lungs, as detected by ChIP assays (n = 3). (g) Luciferase assay of pulmonary epithelial cells integrated with luciferase reporter driven by the albumin promoter and transfected with scrambled siRNA (SCR), Hnf4a siRNA, or Foxa2 siRNA (n = 3). (h) Expression of Hnf4a was determined in lungs of Cdh5^CreHgf^wt/wtMgp^+/+, Cdh5^CreHgf^flox/floxMgp^+/+, Cdh5^CreHgf^wt/wtMgp^−/−, and Cdh5^CreHgf^flox/floxMgp^−/− mice by immunoblotting. WT Mgp^+/+ liver was used as a control (n = 3). (i) Protein interactions of lung lysates were analyzed by immunoprecipitation, followed by immunoblotting with antibodies as indicated (n = 6). Lysates from WT Mgp^+/+ liver were used as a positive control. (j) Schematic diagram of the binding of Hnf4a and Foxa2 competing with that of Foxa2 and Nkx2.1 in Mgp^−/− lungs. Data in a and d–g were analyzed by two-sided t test. *, P < 0.05; ***, P < 0.001. Error bars are standard deviation. Data distribution was assumed to be normal, but this was not formally tested.