15-deoxy-Δ12,14-prostaglandin J2 inhibits transcriptional activity of estrogen receptor-α via covalent modification of DNA-binding domain
15-deoxy-Δ12,14-prostaglandin J2 inhibits transcriptional activity of estrogen receptor-α via covalent modification of DNA-binding domain
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DOI：
10.1158/0008-5472.can-06-3043
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发表时间：
2007-03-15
期刊：
影响因子：
11.2
通讯作者：
Farrar, William L.
中科院分区：
文献类型：
--
作者：
Kim, Han-Jong;Kim, Joon-Young;Farrar, William L.
环戊烯酮 15-脱氧-Delta(12,14)-prostagiandin J(2) (15d-PGJ(2)) 通过过氧化物酶体增殖物激活受体 γ (PPAR γ) 依赖性和 PPAR γ 独立机制抑制癌细胞（包括乳腺癌）的增殖。然而，人们对其对雌激素受体α（ERα）转录活性的影响知之甚少，而雌激素受体α在乳腺癌的生长中起着至关重要的作用。在这里，我们发现 15d-PGJ2 通过 PPAR γ 独立机制抑制 17 β-雌二醇 (E-2) 依赖性和 E-2 独立 ER α 转录活性。此外，15d-PGJ(2) 通过其反应性环戊烯酮部分直接修饰 ER α 蛋白，这一点可通过生物素化的 15d-PGJ(2) 在体外和体内掺入 ER α 来证明。纳流反相液相色谱串联质谱分析确定 ER α DNA 结合结构域 (DBD) 的 COOH 末端锌指内的两个半胱氨酸（Cys(227) 和 Cys(240)）作为 15d-PGJ2 共价修饰的靶标。凝胶迁移率变化和染色质免疫沉淀测定表明，15d-PGJ(2) 抑制 ER α 的 DNA 结合，并随后抑制 ER α 靶基因的表达，例如 pS2 和 c-Myc。因此，我们的结果表明，15d-PGJ(2) 可以通过共价修饰 ER α DBD 脆弱的 COOH 末端锌指内的半胱氨酸残基来阻断 ER α 功能，从而从根本上抑制激素依赖性和激素非依赖性 ER α 转录活性。
The cyclopentenone 15-deoxy-Delta(12,14)-prostagiandin J(2) (15d-PGJ(2)) inhibits proliferation of cancer cells, including breast cancers, by peroxisome proliferator-activated receptor-gamma (PPAR gamma)-dependent and PPAR gamma-independent mechanisms. However, little is known about its effect on the transcriptional activity of estrogen receptor-alpha (ER alpha) that plays vital roles in the growth of breast cancers. Here, we show that 15d-PGJ2 inhibits both 17 beta-estradiol (E-2)-dependent and E-2-independent ER alpha transcriptional activity by PPAR gamma-independent mechanism. In addition, 15d-PGJ(2) directly modifies ER alpha protein via its reactive cyclopentenone moiety, evidenced by incorporation of biotinylated 15d-PGJ(2) into ER alpha, both in vitro and in vivo. Nanoflow reverse-phase liquid chromatography tandem mass spectrometry analysis identifies two cysteines (Cys(227) and Cys(240)) within the COOH-terminal zinc finger of ER alpha DNA-binding domain (DBD) as targets for covalent modification by 15d-PGJ2. Gel mobility shift and chromatin immunoprecipitation assays show that 15d-PGJ(2) inhibits DNA binding of ER alpha and subsequent repression of ER alpha target gene expression, such as pS2 and c-Myc. Therefore, our results suggest that 15d-PGJ(2) can block ER alpha function by covalent modification of cysteine residues within the vulnerable COOH-terminal zinc finger of ER alpha DBD, resulting in fundamental inhibition of both hormone-dependent and hormone-independent ER alpha transcriptional activity.