Novel Models for Epigenetic Screening Mitchell Turker Oregon Health & Science University Portland, OR 97239 A relation between environmental exposure and epigenetic change is increasingly accepted, but underlying mechanisms remain to be elucidated. A variety of environmental exposure can decrease expression of target genes and we have hypothesized that one trigger for epigenetic silencing is prolonged gene repression. Two model systems were developed to this hypothesis. The first model targeted the tet repressible promoter (TRE) for silencing. This promoter is expressed at high levels when bound to the tTA fusion protein (tTA), which contains a viral transcriptional activator. Expression of the tet promoter is reduced significantly when cells are exposed to tetracycline or doxycycline because these drugs bind the tTA activator protein and thereby prevent it from binding and activating the tet promoter. To test our hypothesis, the TRE promoter was used to express the selectable HPRT gene in a mouse cell line. Prolonged repression of the TRE-HPRT construct with doxycycline led to a time dependent increase in HPRT deficient cells. A molecular analysis demonstrated that loss of HPRT expression was due to epigenetic silencing of the TRE promoter and revealed a potential pathway for sequential epigenetic changes. The second model extended the TRE work using the BRCA1 promoter, which is a disease relevant mammalian promoter that is repressible with an environmental exposure, hypoxia. A BRCA1-HPRT construct was made, introduced into the same mouse cell line used above, and hypoxic conditions were found to induce epigenetic silencing. Additional work showed specific histone changes that occurred at the BRCA1 promoter under the hypoxic conditions and after promoter silencing had occurred. In sum, we have shown that promoter repression causes epigenetic silencing in mammalian cells. These observations suggest potential pathways for environmentally-induced epigenetic change.