FIG. 6.

Transcriptional activation of the COX-2 promoter by VacA in AZ-521 cells. (A) Schematic representation of the 5′ regulatory region of the COX-2 gene with mutations (26, 27). Rectangles indicate the location of the NF-κB, NF-IL-6, and CRE sites. The luciferase reporter driven by the COX-2 promoter region (−327/+59) was mutated at the NF-κB, NF-IL-6, or CRE site. The mutated sequences are as follows: NF-κB site (−233/−214), changed from GGGACTACCC to cccgggACCC; NF-IL-6 site (−132/−124), changed from TTACGCAAT to TTggtaccT; CRE site (−59/−53), changed from TTCGTCA to TTgagCt. Promoter activity is not contained in the short type of reporter vector containing the 5′-flanking region of the COX-2 gene promoter (−52/+59). Distances are given as nucleotide positions relative to the transcriptional start site, which is +1. (B) AZ-521 cells were transiently transfected with COX-2 promoter-luciferase reporter plasmids with the −327/+59, CRM, ILM, KBM, or −52/+59 promoters. Cells were either untreated or treated with 120 nM VacA (0 or 6 h) at 37°C. Relative changes in luciferase expression were measured. Open bars, activities without VacA; solid bars, activities with VacA incubation. Data are means ± standard deviations of values from three independent experiments with assays in duplicate. *, P < 0.02.