Suppression of the mTOR-raptor signaling pathway by the inhibitor of heat shock protein 90 geldanamycin
Suppression of the mTOR-raptor signaling pathway by the inhibitor of heat shock protein 90 geldanamycin
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DOI：
10.1093/jb/mvj008
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发表时间：
2006-01-01
影响因子：
2.7
通讯作者：
Yonezawa, K
中科院分区：
文献类型：
--
作者：
Ohji, G;Hidayat, S;Yonezawa, K
将热休克蛋白90（Hsp 90）与raptor免疫共沉淀，raptor是来自HEK 293细胞的雷帕霉素的哺乳动物靶标（mTOR）的结合伴侣。通过使用抗Hsp 90抗体的免疫印迹分析，在从细胞提取物制备的抗猛禽抗体免疫沉淀物中检测到Hsp 90，并且通过来自共表达Hsp 90和猛禽作为表位标记分子的细胞的免疫沉淀，确认这两种蛋白质的缔合。Geldanamycin是一种有效的Hsp 90抑制剂，在体内破坏Hsp 90与raptor的结合，而不影响raptor与mTOR的结合，并抑制mTOR对下游翻译调节因子p70 S6激酶（S6 K）和真核起始因子4 E-binding protein 1（4 E-BP 1）的磷酸化。在格尔德霉素处理的细胞中，S6 K的蛋白激酶活性以及底物40 S核糖体蛋白S6的磷酸化降低。这些结果表明，Hsp 90通过与raptor的结合促进mTOR/raptor复合物催化的4 E-BP 1和S6 K的磷酸化反应，参与蛋白质翻译的调控，mTOR信号通路是格尔德霉素的一个新靶点。
Heat shock protein 90 (Hsp90) was co-immunoprecipitated with raptor, the binding partner of the mammalian target of rapamycin (mTOR) from HEK293 cells. Hsp90 was detected in the anti-raptor antibody immunoprecipitates prepared from the cell extract by immunoblot analysis using the anti-Hsp90 antibody, and the association of these two proteins was confirmed by immunoprecipitation from the cells co-expressing Hsp90 and raptor as epitope-tagged molecules. Geldanamycin, a potent inhibitor of Hsp90, disrupted the in vivo binding of Hsp90 to raptor without affecting the association of raptor and mTOR, and suppressed the phosphorylation by mTOR of the downstream translational regulators p70 S6 kinase (S6K) and eukaryotic initiation factor 4E-binding protein 1 (4E-BP1). The protein kinase activity of S6K as well as the phosphorylation of the substrate, 40S ribosomal protein S6, were lowered in the geldanamycin-treated cells. These results indicate that Hsp90 is involved in the regulation of protein translation by facilitating the phosphorylation reaction of 4E-BP1 and S6K catalyzed by the mTOR/raptor complex through the association with raptor, and that the mTOR signaling pathway is a novel target of geldanamycin.