Figure 8. Effect of dominant-negative A-USF expression on HO-1 reporter and mRNA expression.

(A) HK-2 cells were co-transfected with pHOGH/4.5 and either empty vector (pCMV566) or A-USF as described in the Experimental section and subsequently treated with vehicle (V), cadmium (Cd, 10 μM) or haemin (H; 5 μM) for 12 h. Total RNA was isolated and subjected to Northern-blot analysis with ³²P-labelled cDNA probes specific for hGH or GAPDH. The numbers at the bottom refer to the relative band intensities as determined by densitometry and controlled for GAPDH from three independent experiments. (B) Effect of A-USF expression on luciferase reporter activity of pHOGL3/4.5. Results are expressed as means±S.E.M. *P<0.05 versus vector; **P<0.05 versus vehicle. (C) Northern-blot analysis of endogenous HO-1 gene induction with and without A-USF expression. HK-2 cells were co-transfected with either empty vector or A-USF and treated with V, Cd or H as in (A). Total RNA was isolated and subjected to Northern-blot analysis with ³²P-labelled cDNA probes specific for HO-1 or GAPDH. A densitometric analysis of the autoradiogram corrected for the internal control (GAPDH) is shown in the graph below.