Exposure of mammalian cells to redox active xenobiotics, such as quinoid compounds, leads to considerable structural damage as well as carcinogenesis. A certain degree of protection against such substances is offered by induction of phase II xenobiotic metabolizing enzymes such as quinone reductase (QR) and glutathione S-transferase (GST). This induction occurs at the transcriptional level and is mediated through a common cis acting element present in the promoters of both the QR and GST-Ya subunit genes, termed either ARE (antioxidant response element) of ERE (electrophile response element). The ARE/ERE sequence shows considerable similarity to the consensus sequence recognized by AP-1, a sequence specific transcription factor composed of Jun and Fos proteins. In fact, AP-1 binds to the various ARE/ERE sequences and its activity is induced in response to oxidative stress. We propose that AP-1 serves a second prototype of xenobiotic """"""""receptor"""""""". Unlike the classical Ah receptor, AP-1 activity is not regulated by direct binding of xenobiotics. A likely mechanism that controls AP-1 activity involves a signal transduction cascade that is initiated by free radicals that are produced as a result of exposure to redox active xenobiotics. to test this hypothesis and determine the mechanisms by which electrophilic xenobiotics stimulate AP-1 activity, we propose to: 1) determine the composition of the AP-1 complex that interacts with the ARE/EREs; 2) determine whether redox active xenobiotics stimulate AP-1 activity by transcriptional or post-translational mechanisms; 3) analyze changes in Jun protein phosphorylation in response to xenobiotic exposure; 4) examine the induction of c-jun transcription by xenobiotics and determine the xenobiotic response element in the c-jun promoter; 5) examine the involvement of tyrosine kinases, Ha-Ras and Raf-1 proteins in the signal transduction pathway leading to gene induction through ARE/ERE and increased AP-1 activity; 6) examine the effect of xenobiotics on tyrosine phosphorylation and the activity of tyrosine kinases; 7) examine whether xenobiotics lead to inhibition of tyrosine phosphatases via free radical production. These studies should provide a detailed molecular understanding of the mechanisms by which redox active xenobiotics affect gene expression and act as tumor promoters.
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