Study on the Mammalian target of rapamycin in thrombocytepoiesis, wound healing, and hyperlipemia

雷帕霉素在血小板生成、伤口愈合和高脂血症中的哺乳动物靶点研究

基本信息

批准号：
16390094
负责人：
YOSHINO Ken-ichi
金额：
$ 9.41万
依托单位：
Kobe University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
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 mammalian target of rapamycin (mTOR) is a Ser/Thr protein kinase that plays a crucial role in a nutrient-sensitive signalling pathway that regulates cell growth. TOR signalling is potently inhibited by rapamycin, through the direct binding of a FK506-binding protein 12 (FKBP12)/rapamycin complex to the TOR FRB domain, a segment amino terminal to the kinase catalytic domain. Raptor (regulatory associated protein of mTOR) is a recently identified mTOR binding partner that is essential for mTOR signalling in vivo, and whose binding to mTOR is critical for mTOR-catalyzed substrate phosphorylation in vitro.Here we investigated the stability of endogenous mTOR/raptor complex in response to rapamycin in vivo, and to the direct addition of a FKBP12/rapamycin complex in vitro. We demonstrated that rapamycin inhibits mTOR function, at least in part, by inhibiting the interaction of raptor with mTOR; this action uncouples mTOR from its substrates, and inhibits mTOR signalling without altering … More mTOR's intrinsic catalytic activity.We showed that heat shock protein 90 (Hsp90) is involved in the regulation of protein translation by facilitating the phosphorylation reaction of eukaryotic initiation factor-4E (eIF4E) binding protein 1 (4E-BP1) and p70 S6 kinase (S6K) catalyzed by the mTOR/raptor complex through the association with raptor, and that the mTOR signaling pathway is a novel target of geldanamycin.We found that the TOS motif has an essential function whereas the RAIP motif has an accessory role in the association with raptor and mTOR-mediated phosphorylation of 4E-BP1 to dissociate it from raptor and release eIF4E in response to amino acid stimulation leading to the control of cell size.The proline-rich Akt substrate of 40 kilodaltons (PRAS40) was identified as a raptor binding protein that is phosphorylated directly by mTOR complex (mTORC) 1 but not mTORC 2 in vitro, predominantly at PRAS40 (Serl 83). We establish PRAS40 as a physiological mTORCl substrate that contains a variant TOS motif Moreover, they indicate that the capacity or ability of raptor to bind endogenous substrates is limiting for the activity of mTORCl in vivo, and is therefore a potential locus of regulation, as suggested by others. Less

雷帕霉素的哺乳动物靶标 (mTOR) 是一种 Ser/Thr 蛋白激酶，在调节细胞生长的营养敏感信号通路中发挥着至关重要的作用，雷帕霉素通过直接结合 FK506 结合蛋白来有效抑制 TOR 信号通路。 12 (FKBP12)/雷帕霉素与 TOR FRB 结构域的复合物，TOR FRB 结构域是激酶催化结构域（Raptor 的调节相关蛋白）的氨基末端片段。 mTOR）是最近发现的一种 mTOR 结合伴侣，对于体内 mTOR 信号传导至关重要，并且其与 mTOR 的结合对于体外 mTOR 催化的底物磷酸化至关重要。在这里，我们研究了内源性 mTOR/raptor 复合物响应雷帕霉素的稳定性体内，以及体外直接添加 FKBP12/雷帕霉素复合物，我们证明雷帕霉素至少部分通过抑制 raptor 与 raptor 的相互作用来抑制 mTOR 功能。 mTOR；这种作用将 mTOR 与其底物解偶联，并抑制 mTOR 信号转导而不改变 mTOR 的内在催化活性。我们发现热休克蛋白 90 (Hsp90) 通过促进真核起始因子的磷酸化反应来参与蛋白质翻译的调节-4E (eIF4E) 结合蛋白 1 (4E-BP1) 和 p70 S6 激酶 (S6K) 催化mTOR/raptor复合物通过与raptor的关联，并且mTOR信号通路是格尔德霉素的新靶点。我们发现TOS基序具有重要功能，而RAIP基序在与raptor和mTOR介导的关联中具有辅助作用4E-BP1 磷酸化，使其与 raptor 分离并释放 eIF4E，以响应氨基酸刺激，从而控制细胞大小。 40 富含脯氨酸的 Akt 底物千道尔顿（PRAS40）被鉴定为猛禽结合蛋白，其在体外被mTOR复合物（mTORC）1而非mTORC 2直接磷酸化，主要在PRAS40（Serl 83）处我们将PRAS40建立为包含变体TOS的生理mTORC1底物。此外，它们表明raptor结合内源底物的能力或能力限制了mTORC1体内的活性，因此是一个潜在的基因座正如其他人所建议的那样，监管。