Am. J. Respir. Cell Mol. Biol., Volume 19, Number 2, August, 1998 316-323

Egr-1 and Sp1 Interact Functionally with the 5-Lipoxygenase Promoter and Its Naturally Occurring Mutants

Eric S. Silverman, Jing Du, George T. De Sanctis, Olof Rådmark, Bengt Samuelsson, Jeffrey M. Drazen, and Tucker Collins

Pulmonary and Critical Care Division, Department of Medicine, and Vascular Research Division, Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts; and Department of Medical Biochemistry and Biophysics, Division of Physiological Chemistry II, Karolinska Institutet, Stockholm, Sweden

5-Lipoxygenase (5-LO), an enzyme essential for the formation of leukotrienes, is functionally modulated by a number of mechanisms, including transcriptional controls. The 5-LO promoter has a unique G+C-rich sequence, located between 176 and 147 base pairs upstream of the ATG translation start site, which contains five tandem Sp1 (a zinc-finger transcription factor) consensus binding sites overlapping five tandem early growth response protein 1 (Egr-1), a zinc-finger transcription factor, consensus binding sites. A family of naturally occurring mutations has been identified that consists of additions or deletions of these binding sites. The role of these overlapping Sp1/Egr-1 sites in the regulation of 5-LO transcription and the effects of these mutations on transcriptional regulatory mechanisms are unknown. We now show that Sp1 and Egr-1 bind specifically to the G+C-rich promoter sequence using in vitro deoxyribonuclease I footprinting. Both Sp1 and Egr-1 activate 5-LO promoter-reporter constructs in a minimally active drosophila SL2 cotransfection system, and the G+C-rich sequence is involved in this process. Moreover, studies comparing mutant promoter function indicate that both Sp1 and Egr-1 trans-activation are proportional to the number of Sp1/Egr-1 consensus binding sites within the G+C-rich sequence. It is possible that basal and inducible 5-LO gene transcriptions are mediated by an interplay of Sp1, Egr-1, and other transcription factors within the G+C-rich promoter region, and the naturally occurring mutations alter transcription by modifying their trans-activation potential.

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