Effect of 3 PU.1 sites onp40^phox promoter function in HL-60 cells.

(A) Mutational analysis of the −1599 to +104 region of thep40^phox promoter. The 3 PU.1 sites (open boxes) present in the pGL3-p40-1599 construct were mutated (hatched boxes) singly or in combination, and the resultant constructs were assayed for reporter gene activity in HL-60 cells as in Figure 1. The arrow indicates the reported transcription start site. (B) Deletion analysis of the contribution of each PU.1 site to the overall transactivation activity of the proximal promoter (−106 to +104 bp) of the p40^phox gene. Deletion constructs were prepared and assayed for reporter gene activity in HL-60 cells as in Figure 1. Data shown in both panels are means (± SE) of 8 independent experiments. Analysis of variance showed that differences in luciferase activity among the constructs were significant (P < .01), and t test demonstrated that the luciferase activities of the mutation and deletion constructs were significantly lower (P < .01 except for the PU.1c mutation construct, where P = .044) than those from wild-type counterparts.