Deb Santanu, Zhou Jianfeng, Amin Sanober A, Imir Ayse Gonca, Yilmaz Mehmet Bertan, Lin Zhihong, Bulun Serdar E
Division of Reproductive Biology Research, Northwestern University, Chicago, Illinois 60611, USA.
J Biol Chem. 2006 Feb 3;281(5):2585-97. doi: 10.1074/jbc.M508498200. Epub 2005 Nov 21.
The aromatase gene encodes the key enzyme for estrogen formation. Aromatase enzyme inhibitors eliminate total body estrogen production and are highly effective therapeutics for postmenopausal breast cancer. A distal promoter (I.4) regulates low levels of aromatase expression in tumor-free breast adipose tissue. Two proximal promoters (I.3/II) strikingly induce in vivo aromatase expression in breast fibroblasts surrounding malignant cells. Treatment of breast fibroblasts with medium conditioned with malignant breast epithelial cells (MCM) or a surrogate hormonal mixture (dibutyryl (Bt2)cAMP plus phorbol diacetate (PDA)) induces promoters I.3/II. The mechanism of promoter-selective expression, however, is not clear. Here we reported that sodium butyrate profoundly decreased MCM- or Bt2cAMP + PDA-induced promoter I.3/II-specific aromatase mRNA. MCM, Bt2cAMP + PDA, or sodium butyrate regulated aromatase mRNA or activity only via promoters I.3/II but not promoters I.1 or I.4 in breast, ovarian, placental, and hepatic cells. Mechanistically, recruitment of phosphorylated ATF-2 by a CRE (-211/-199, promoter I.3/II) conferred inductions by MCM or Bt2cAMP + PDA. Chromatin immunoprecipitation-PCR and immunoprecipitation-immunoblotting assays indicated that MCM or Bt2cAMP + PDA stabilized a complex composed of phosphorylated ATF-2, C/EBPbeta, and cAMP-response element-binding protein (CREB)-binding protein in the common regulatory region of promoters I.3/II. Overall, histone acetylation patterns of promoters I.3/II did not correlate with sodium butyrate-dependent silencing of promoters I.3/II. Sodium butyrate, however, consistently disrupted the activating complex composed of phosphorylated ATF-2, C/EBPbeta, and CREB-binding protein. This was mediated, in part, by decreased ATF-2 phosphorylation. Together, these findings represent a novel mechanism of sodium butyrate action and provide evidence that aromatase activity can be ablated in a signaling pathway- and cell-specific fashion.
芳香化酶基因编码雌激素形成的关键酶。芳香化酶抑制剂可消除全身雌激素的产生，是绝经后乳腺癌的高效治疗药物。一个远端启动子（I.4）调节无肿瘤乳腺脂肪组织中低水平的芳香化酶表达。两个近端启动子（I.3/II）显著诱导恶性细胞周围乳腺成纤维细胞中的体内芳香化酶表达。用恶性乳腺上皮细胞条件培养基（MCM）或替代激素混合物（二丁酰（Bt2）环磷酸腺苷（cAMP）加佛波酯二乙酸酯（PDA））处理乳腺成纤维细胞可诱导启动子I.3/II。然而，启动子选择性表达的机制尚不清楚。在此，我们报道丁酸钠显著降低MCM或Bt2cAMP + PDA诱导的启动子I.3/II特异性芳香化酶mRNA。MCM、Bt2cAMP + PDA或丁酸钠仅通过启动子I.3/II调节乳腺、卵巢、胎盘和肝细胞中的芳香化酶mRNA或活性，而不通过启动子I.1或I.4。从机制上讲，CRE（-211/-199，启动子I.3/II）招募磷酸化的ATF-2介导了MCM或Bt2cAMP + PDA的诱导作用。染色质免疫沉淀-PCR和免疫沉淀-免疫印迹分析表明，MCM或Bt2cAMP + PDA在启动子I.3/II的共同调节区域稳定了由磷酸化的ATF-2、C/EBPβ和cAMP反应元件结合蛋白（CREB）结合蛋白组成的复合物。总体而言，启动子I.3/II的组蛋白乙酰化模式与丁酸钠依赖性启动子I.3/II沉默无关。然而，丁酸钠持续破坏由磷酸化的ATF-2、C/EBPβ和CREB结合蛋白组成的激活复合物。这部分是由ATF-2磷酸化减少介导的。总之，这些发现代表了丁酸钠作用的一种新机制，并提供了证据表明芳香化酶活性可以通过信号通路和细胞特异性方式被消除。
