FIGURE 2.

Effect of sumoylation on MTF-1 transcriptional activity. A, MTF-1 or its K627R mutant were co-transfected with the SUMO-1 and 5×MREd-luciferase reporter plasmids into CHO K1 cells. The cells were treated with or without 100 μm zinc for 6 h, and luciferase activity was measured. B, SUMO-1 and MTF-1-His₆ or MTF-1-SUMO-His₆ plasmids were co-transfected into CHO K1 cells. The expressed proteins were immunoprecipitated (IP) and then subjected to Western blotting with SUMO-1 antibodies. C, MTF-1-His₆, MTF-1-SUMO-His₆, or MTF-1-GFP-His₆ plasmids were co-transfected with 5×MREd-luciferase reporter plasmids into CHO K1 cells. Relative luciferase activity was determined after adding 100 μm zinc to the transfected cells for 6 h. D, HEK293 cells were transfected with MTF-1-His₆ or MTF-1-SUMO-His₆ encoding plasmids and treated with 100 μm zinc for 6 h. RNA was extracted and the relative MTIIA mRNA level was determined by quantitative real-time PCR. The MTIIA mRNA level in cells transfected with the MTF-1 plasmid with zinc treatment was designated as 100%. Each value represents a mean ± S.D. of three independent experiments. Asterisks denote significant differences (p < 0.05) from metal-treated cells transfected with MTF-1 (A, C, and D).