Dissociation of raptor from mTOR is a mechanism of rapamycin-induced inhibition of mTOR function
Dissociation of raptor from mTOR is a mechanism of rapamycin-induced inhibition of mTOR function
复制标题
DOI：
10.1111/j.1356-9597.2004.00727.x
复制
发表时间：
2004-04-01
期刊：
影响因子：
2.1
通讯作者：
Yonezawa, K
中科院分区：
文献类型：
--
作者：
Oshiro, N;Yoshino, K;Yonezawa, K
哺乳动物雷帕霉素靶蛋白 (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 FK506 结合蛋白 12 (FKBP12)/雷帕霉素复合物与 TOR FRB 结构域（激酶催化结构域的氨基末端片段）的直接结合，有效抑制 TOR 信号传导。 FKBP12/雷帕霉素抑制作用的分子基础仍不确定。 Raptor（mTOR 调节相关蛋白）是最近鉴定的 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 复合物在体内响应雷帕霉素的稳定性，以及在体外直接添加 FKBP12/雷帕霉素复合物的稳定性。雷帕霉素在体内减少了内源性 raptor 与内源性或重组 mTOR 的恢复；这种抑制需要 mTOR 结合 FKBP12/雷帕霉素复合物的能力，但与 mTOR 激酶活性无关。在 FKBP12 存在的情况下，雷帕霉素在体外直接抑制 raptor 与 mTOR 的结合，同时减少 mTOR 催化的 raptor 依赖性底物的磷酸化，但不减少 raptor 不依赖性底物的磷酸化； mTOR 自磷酸化未改变。这些观察结果表明，雷帕霉素至少部分通过抑制 raptor 与 mTOR 的相互作用来抑制 mTOR 功能；这一作用使 mTOR 与其底物解偶联，并抑制 mTOR 信号传导，而不改变 mTOR 的内在催化活性。
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. The molecular basis for the inhibitory action of FKBP12/rapamycin remains uncertain. 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-catalysed 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. Rapamycin diminished the recovery of endogenous raptor with endogenous or recombinant mTOR in vivo; this inhibition required the ability of mTOR to bind the FKBP12/rapamycin complex, but was independent of mTOR kinase activity. Rapamycin, in the presence of FKBP12, inhibited the association of raptor with mTOR directly in vitro, and concomitantly reduced the mTOR-catalysed phosphorylation of raptor-dependent, but not raptor-independent substrates; mTOR autophosphorylation was unaltered. These observations indicate 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 mTOR's intrinsic catalytic activity.