FUNCTIONAL ANTIOXIDANT RESPONSIVE ELEMENTS

Exposure of human and rodent cells to a wide variety of chemoprotectiv e compounds confers resistance against a broad set of carcinogens, For a subset of the chemoprotective compounds, protection is generated by an increase in the abundance of protective enzymes like glutathione S -transferases (GST), Antioxidant responsive elements (AREs) mediate th e transcriptional induction of a battery of genes which comprise much of this chemoprotective response system. Past studies identified a nec essary ARE ''core'' sequence of RTGACnnnGC, but this sequence alone is insufficient to mediate induction, In this study, the additional sequ ences necessary to define a sufficient, functional ARE are identified through systematic mutational analysis of the murine GST Ya ARE, Intro duction of the newly identified necessary nucleotides into the regions flanking a nonresponsive, ARE-like, GST-Mu promoter sequence produced an inducible element. A screen of the GenBank database with the newly identified ARE consensus identified 16 genes which contained the func tional ARE consensus sequence in their promoters. Included within this group,vas an ARE sequence from the murine ferritin-L promoter that me diated induction when tested. In an electrophoretic mobility-shift ass ay, the ferritin-L ARE was bound by ARE-binding protein 1, a protein p reviously identified as the likely mediator of the chemoprotective res ponse. A three level ARE classification system is presented to account for the distinct induction strengths observed in our mutagenesis stud ies, A model of the ARE as a composite regulatory site, where multiple transcription factors interact, is presented to account for the compl ex characteristics of ARE-mediated chemoprotective gene expression.