Figure 4. Prdx6-/- LECs, a model for aging, bore an enhanced Sumoylated form of Sp1, and levels were further increased with exposure to oxidative stress. (A) Nuclear extracts were prepared from Prdx6+/+ and Prdx6−/− mLECs and submitted to Sp1 Sandwich/Sumo1-ELISA assays to examine the total and Sumoylated forms of Sp1 protein. Sumoylated Sp1 protein was subtracted from total Sp1 protein, and results are presented as deSumoylated (gray bars) and Sumoylated (black bars) forms of Sp1. The data represent mean ± SD from three independent experiments. Prdx6+/+ vs Prdx6−/− ; *p<0.001. (B-E) Prdx6-/- LECs displayed increased levels of Sumoylated Sp1 in response to increased oxidative loads. Prdx6-/- LECs were exposed to H2O2 (B and C) or UVB (D and E) in different concentrations and for different time intervals as indicated. Nuclear extracts were prepared and used to perform Sp1 sandwich/Sumo1-ELISA specific to Sp1. Sumoylated Sp1 protein was subtracted from total Sp1 protein, presented as deSumoylated Sp1 (gray bars) and Sumoylated Sp1 (black bars). B-E: Data represent mean ± SD from three independent experiments. 0µM vs 25, 50 and 75µM (H2O2), 0 min vs 15 , 30 and 90 min of H2O2 exposure and 0 J/m2 vs 30, 60 and 90 J/m2 (UVB) and 0h vs 3, 6 and 12h UVB exposure; **p<0.05; *p<0.001.