Estrogen induces c-myc gene expression via an upstream enhancer activated by the estrogen receptor and the AP-1 transcription factor

c-myc oncogene is implicated in tumorigenesis of many cancers, including breast cancer. Although c-myc is a well-known estrogen-induced gene, its promoter has no estrogen-response element, and the underlying mechanism by which estrogen induces its expression remains obscure. Recent genome-wide studies by us and others suggested that distant elements may mediate estrogen induction of gene expression. In this study, we investigated the molecular mechanism by which estrogen induces c-myc expression with a focus on these distal elements. Estrogen rapidly induced c-myc expression in estrogen receptor (ER)-positive breast cancer cells. Although estrogen had little effect on c-myc proximal promoter activity, it did stimulate the activity of a luciferase reporter containing a distal 67-kb enhancer. Estrogen induction of this luciferase reporter was dependent upon both a half-estrogen response element and an activator protein 1 (AP-1) site within this enhancer, which are conserved across 11 different mammalian species. Small interfering RNA experiments and chromatin immunoprecipitation assays demonstrated the necessity of ER and AP-1 cross talk for estrogen to induce c-myc expression. TAM67, the AP-1 dominant negative, partially inhibited estrogen induction of c-myc expression and suppressed estrogen-induced cell cycle progression. Together, these results demonstrate a novel pathway of estrogen regulation of gene expression by cooperation between ER and AP-1 at the distal enhancer element and that AP-1 is involved in estrogen induction of the c-myc oncogene. These results solve the long-standing question in the field of endocrinology of how estrogen induces c-myc expression.     

Pax6 and Pdx1 form a functional complex on the rat somatostatin gene upstream enhancer

The somatostatin upstream enhancer (SMS-UE) is a highly complex enhancer element. The distal A-element contains overlapping Pdx1 and Pbx binding sites. However, a point mutation in the A-element that abolishes both Pdxl and Pbx binding does not impair promoter activity. In contrast, a point mutation that selectively eliminates Pdx1 binding to a proximal B-element reduces the promoter activity. The B-element completely overlaps with a Pax6 binding site, the C-element. A point mutation in the C-element demonstrates that Pax6 binding is essential for promoter activity. Interestingly, a block mutation in the A-element reduces both Pax6 binding and promoter activity. In heterologous cells, Pdx1 potentiated Pax6 mediated activation of a somatostatin reporter. We conclude that the beta/delta-cell-specific activity of the SMS-UE is achieved through simultaneous binding of Pdx1 and Pax6 to the B- and C-elements, respectively. Furthermore, the A-element appears to stabilise Pax6 binding.