Regulation of SRC-3 intercompartmental dynamics by estrogen receptor and phosphorylation
Regulation of SRC-3 intercompartmental dynamics by estrogen receptor and phosphorylation
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DOI：
10.1128/mcb.01695-06
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发表时间：
2007-10-01
影响因子：
5.3
通讯作者：
Mancini, Michael A.
中科院分区：
文献类型：
--
作者：
Amazit, Larbi;Pasini, Luigi;Mancini, Michael A.
类固醇受体共激活因子3基因（SRC-3）（AIBI/ACTR/pCIP/RAC 3/TRAM 1）是p160家族转录共激活因子，并且是已知的癌基因。尽管其重要性，SRC-3的功能调节在细胞环境中仍然知之甚少。使用活细胞，高通量和荧光显微镜的新组合，我们报告SRC-3是一个核质穿梭蛋白，其细胞内的流动性，溶解度和细胞定位的磷酸化和雌激素受体α（ER α）的相互作用进行调节。我们发现，化学抑制和小干扰RNA减少丝裂原活化蛋白激酶/细胞外信号调节激酶1/2（MEK 1/2）途径诱导SRC-3定位的细胞质移位，而表皮生长因子信号刺激通过诱导T24，S857和S860磷酸化增强其核定位，已知参与调节SRC-3活性的磷酸密码子。因此，非磷酸化SRC-3突变体的细胞质定位进一步支持了这些结果。在ER α存在的情况下，U 0126还显著减少（i）SRC-3和ER α的配体依赖性共定位，（ii）细胞裂解物中ER-SRC-3复合物的形成，以及（iii）SRC-3靶向可见的、ER α占据和调节的催乳素启动子阵列。总之，这些结果表明，磷酸化协调SRC-3辅激活因子的功能，通过连接的概率形成的瞬时核受体辅激活因子复合物与其分子动力学和细胞区室化。从技术和概念上讲，这些发现对评估基因调节剂在细胞系统水平上的作用机制具有新的和广泛的影响。
The steroid receptor coactivator 3 gene (SRC-3) (AIBI/ACTR/pCIP/RAC3/TRAM1) is a p160 family transcription coactivator and a known oncogene. Despite its importance, the functional regulation of SRC-3 remains poorly understood within a cellular context. Using a novel combination of live-cell, high-throughput, and fluorescent microscopy, we report SRC-3 to be a nucleocytoplasmic shuttling protein whose intracellular mobility, solubility, and cellular localization are regulated by phosphorylation and estrogen receptor alpha (ER alpha) interactions. We show that both chemical inhibition and small interfering RNA reduction of the mitogen activated protein kinase/extracellular signal-regulated kinase 1/2 (MEK1/2) pathway induce a cytoplasmic shift in SRC-3 localization, whereas stimulation by epidermal growth factor signaling enhances its nuclear localization by inducing phosphorylation at T24, S857, and S860, known participants in the phosphocode that regulates SRC-3 activity. Accordingly, the cytoplasmic localization of a nonphosphorylatable SRC-3 mutant further supported these results. In the presence of ER alpha, U0126 also dramatically reduces (i) ligand-dependent colocalization of SRC-3 and ER alpha, (ii) the formation of ER-SRC-3 complexes in cell lysates, and (iii) SRC-3 targeting to a visible, ER alpha-occupied and -regulated prolactin promoter array. Taken together, these results indicate that phosphorylation coordinates SRC-3 coactivator function by linking the probabilistic formation of transient nuclear receptor-coactivator complexes with its molecular dynamics and cellular compartmentalization. Technically and conceptually, these findings have a new and broad impact upon evaluating mechanisms of action of gene regulators at a cellular system level.