Lin Zhihong, Reierstad Scott, Huang Chiang-Ching, Bulun Serdar E
Department of Obstetrics and Gynecology, Feinberg School of Medicine, Northwestern University, Chicago, Illinois 60611, USA.
Cancer Res. 2007 May 15;67(10):5017-24. doi: 10.1158/0008-5472.CAN-06-3696.
Estrogen receptor-alpha (ERalpha) and its ligand estradiol play critical roles in breast cancer growth and are important therapeutic targets for this disease. Using chromatin immunoprecipitation (ChIP)-on-chip, ligand-bound ERalpha was recently found to function as a master transcriptional regulator via binding to many cis-acting sites genome-wide. Here, we used an alternative technology (ChIP cloning) and identified 94 ERalpha target loci in breast cancer cells. The ERalpha-binding sites contained both classic estrogen response elements and nonclassic binding sequences, showed specific transcriptional activity in reporter gene assay, and interacted with the key transcriptional regulators, including RNA polymerase II and nuclear receptor coactivator-3. The great majority of the binding sites were located in either introns or far distant to coding regions of genes. Forty-three percent of the genes that lie within 50 kb to an ERalpha-binding site were regulated by estradiol. Most of these genes are novel estradiol targets encoding receptors, signaling messengers, and ion binders/transporters. mRNA profiling in estradiol-treated breast cancer cell lines and tissues revealed that these genes are highly ERalpha responsive both in vitro and in vivo. Among estradiol-induced genes, Wnt11 was found to increase cell survival by significantly reducing apoptosis in breast cancer cells. Taken together, we showed novel genomic binding sites of ERalpha that regulate a novel set of genes in response to estradiol in breast cancer. Our findings suggest that at least a subset of these genes, including Wnt11, may play important in vivo and in vitro biological roles in breast cancer.
雌激素受体α（ERα）及其配体雌二醇在乳腺癌生长中起关键作用，是该疾病重要的治疗靶点。最近通过染色质免疫沉淀芯片技术（ChIP-on-chip）发现，与配体结合的ERα通过全基因组范围内与许多顺式作用位点结合，发挥主要转录调节因子的功能。在此，我们使用了另一种技术（ChIP克隆），并在乳腺癌细胞中鉴定出94个ERα靶基因座。ERα结合位点既包含经典的雌激素反应元件，也包含非经典的结合序列，在报告基因检测中显示出特定的转录活性，并与关键转录调节因子相互作用，包括RNA聚合酶II和核受体共激活因子3。绝大多数结合位点位于基因的内含子中或与编码区相距甚远。位于ERα结合位点50 kb范围内的基因中有43%受雌二醇调控。这些基因中的大多数是编码受体、信号信使和离子结合蛋白/转运蛋白的新型雌二醇靶点。对经雌二醇处理的乳腺癌细胞系和组织进行的mRNA分析表明，这些基因在体外和体内均对ERα高度敏感。在雌二醇诱导的基因中，发现Wnt11通过显著减少乳腺癌细胞凋亡来提高细胞存活率。综上所述，我们展示了ERα的新型基因组结合位点，这些位点在乳腺癌中响应雌二醇调节一组新的基因。我们的研究结果表明，这些基因中至少有一部分，包括Wnt11，可能在乳腺癌的体内和体外生物学作用中发挥重要作用。
