Environmental Effects on the Dynamism of Receptors of Lipophilic Signal Molecules in the Brain and its Functional Significance

环境对脑内亲脂信号分子受体动态的影响及其功能意义

基本信息

批准号：
16200026
负责人：
KAWATA Mitsuhiro
金额：
$ 31.62万
依托单位：
Kyoto Prefectural University of Medicine
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2006
项目状态：
已结题

项目摘要

The present research project was undertaken to investigate two types of steroid hormone receptors (glucocorticoids and sex steroids) dynamism by using real-time imaging with fluorescence recovery after photobleaching (FRAP) and fluorescent resonance energy transfer (FRET.) Glucocorticod receptor (GR) and mineralocorticoid receptor (MR) were localized in the cytoplasm in the absence of the ligand and they translocate to the nucleus after ligand binding. FRET demonstrated that importin a was involved in GR/MR translocation from the cytoplasm to the nucleus and GR and MR dimerize within the nucleus. FRAP showed that the movement of ligand-binded GR/MR was restricted in the nucleus. Upon estradiol treatment ERa and b were relocalized to show discrete pattern, and they were localized at the same discrete cluster. FRET clearly showed the interaction of ERa and ERb. In the presence of the estradiol, however, the discrete staining pattern of ERa and b were mostly overlapped with Brg-1, indicating that most of the ERs clusters are involved in the chromatin remodeling machinery. FRAP showed that nuclear ERa and b were dynamic and mobile in the absence of the ligand, but its mobility was slightly decreased after the ligand treatment. Nuclear matrix which was scaffolding sites within the nucleus was composed of actin and actin-related peptides. Treatment by detergent caused complete loss of ERa in the unliganded condition, but liganded ERa stick to the nuclear matrix. Nuclear matrix was an important factor to determine the dynamism of unliganded and liganded ERa.

实施本研究项目是为了研究两种类型的类固醇激素受体（糖皮质激素和性类固醇），通过使用实时成像在光漂白（FRAP）和荧光共振能量转移（FRET。）葡萄皮层受体（GR）和矿物质皮质受体（MRIG中）中的荧光（FRET。配体结合后的核。 FRET证明了Importin A参与了从细胞质到细胞核的GR/MR易位，并且在细胞核内二聚体。 FRAP表明，配体的GR/MR的运动在细胞核中受到限制。在雌二醇治疗时代和B后，将其重新定位以显示离散的模式，并且它们以相同的离散簇定位。 FRET清楚地显示了ERA和ERB的相互作用。但是，在雌二醇的存在下，ERA和B的离散染色模式主要与BRG-1重叠，表明大多数ERS簇都参与染色质重塑机械。 FRAP表明，在没有配体的情况下，核ERA和B是动态的，可移动，但在配体治疗后其迁移率略有下降。核基质中的脚手架位点由肌动蛋白和肌动蛋白相关的肽组成。洗涤剂的治疗在非配合条件下导致ERA完全丧失，但配体ERA坚持核基质。核基质是确定非配体和配体时代动态的重要因素。