Interaction and dissociation by ligands of estrogen receptor and Hsp90: The antiestrogen RU 58668 induces a protein synthesis-dependent clustering of the receptor in the cytoplasm
Interaction and dissociation by ligands of estrogen receptor and Hsp90: The antiestrogen RU 58668 induces a protein synthesis-dependent clustering of the receptor in the cytoplasm
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DOI：
10.1210/me.12.6.842
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发表时间：
1998-06-01
影响因子：
--
通讯作者：
Catelli, MG
中科院分区：
文献类型：
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作者：
Devin-Leclerc, J;Meng, X;Catelli, MG
雌激素受体（ER）和热休克蛋白90的体内相互作用，在激素的情况下，通过细胞质热休克蛋白90与核受体的核共易位试验证明，被破坏的激动剂和拮抗剂配体，解离后的热休克蛋白90，允许伴侣蛋白被重新定位在细胞质中。纯抗雌激素RU 58668（RU），这是无法刺激雌激素依赖的报告基因，并完全抑制其雌二醇诱导的活性也深刻地修改了亚细胞分布的ER在一个特定的时间和剂量依赖性的方式; ER出现斑点荧光团主要位于细胞质的核周区域。在存在或不存在放线菌酮的情况下，RU依赖性ER错误定位的出现和逆转的动力学表明：1）这种作用被RU停药或雌二醇（E-2）处理逆转; 2）放线菌酮与RU抑制并逆转了RU单独诱导的ER胞质错误定位。这些结果表明，在这种抗雌激素的作用机制中存在蛋白质合成依赖性步骤。RU处理后，在细胞质的颗粒部分中发现了大部分ER。然而，共聚焦和电子显微镜分析表明，ER集群不与特定的细胞器或车厢。使用ER突变体，发现配体结合结构域足以使RU产生受体错误定位，而组成性核定位信号是不稳定的。我们提出RU的抗雌激素性质主要是由于诱导了一种聚集倾向的受体构象，这种构象不能承担受体的组成性和配体诱导的核定位功能，因为它通过快速翻转蛋白质而被隔离在细胞质中。我们预测，能够阻断ER核定位的抗雌激素将表现为纯抗激素，并将抑制激动性配体或配体非依赖性机制（如生长因子刺激）引起的ER的所有核作用。
The in vivo interaction of estrogen receptor (ER) and Hsp90, demonstrated in the absence of hormone by a nuclear cotranslocation assay of the cytoplasmic Hsp90 with the karyophilic receptor, was disrupted by agonist and antagonist ligands, which, after dissociating the Hsp90, allowed the chaperone protein to be relocalized in the cytoplasm. The pure antiestrogen RU 58668 (RU), which was unable to stimulate an estrogen-dependent reporter gene and completely inhibited its estradiol-induced activity also profoundly modified the subcellular distribution of ER in a specific time- and dose-dependent manner; ER appeared as speckled fluorescent clusters mainly located in the perinuclear region of the cytoplasm. The kinetics of appearance and reversal of the RU-dependent ER mislocalization in the presence or absence of cycloheximide demonstrated 1) that this effect was reversed by RU withdrawal or estradiol (E-2) treatment, and 2) that cycloheximide with RU Inhibited and reversed the ER cytoplasmic mislocalization induced by RU alone. These results point to a protein synthesis-dependent step in the mechanism of action of this antiestrogen. After RU treatment, a large portion of ER was found in the particulate fraction of the cytoplasm. However, confocal and electron microscopic analysis showed that ER clusters were not associated with specific cytoplasmic organelles or compartments. using ER mutants, it was found that the ligand binding domain was sufficient for RU to produce receptor mislocalization, while the constitutive nuclear localization signals were dispensable. We propose that the antiestrogenic properties of RU are primarily due to the induction of an aggregation-prone receptor conformation that cannot undertake the constitutive and the ligand-induced nuclear localization function of the receptor because it is sequestered in the cytoplasm by fast turning over protein(s). We predict that antiestrogens able to block ER nuclear localization will behave as pure antihormones and will inhibit all the nuclear action of ER elicited by agonistic ligands or by ligand-independent mechanisms such as growth factor stimulation.