Studies on molecular mechanism of mTOR signaling that controls protein synthesis, cell growth, and cell cycle in response to amino acid sufficiency

雷帕霉素的哺乳动物靶标(MTOR)是一种蛋白激酶，通过调节p70S6K(P70S6K)和真核细胞起始因子4E结合蛋白1(4B-BP1)的磷酸化，控制多种细胞功能，包括细胞生长，以响应氨基酸和生长因子。MTOR在体内调节p70S6K和4B-BP1的机制尚不完全清楚。我们发现了一个新的mTOR结合伙伴，Raptor(mTOR的调节相关蛋白)，它也能结合p70S6K和4E-BP1，在体内对TOR信号是必不可少的。我们还证明了Raptor通过各自的TOS(保守的TOR信号)基序与p70S6K和4E-BP1结合，这是一个短的保守片段，在体内对这些mTOR底物的氨基酸和mTOR依赖的调节是必需的。在p70S6K或4E-BP1的TOS基序中的一个点突变可以同时取消氨基酸和mTOR调节，选择性地取消它们与猛禽的结合。TOS基序的这种突变也消除了mTOR催化的4E-BP1在体外的所有磷酸化，并取消了mTOR催化的Vitrv中p70S6K磷酸化的猛禽依赖成分。Raptor不改变mTOR的内在催化活性，但似乎是一种mTOR支架蛋白，其与mTOR底物的TOS基序的结合是体内有效的mTOR催化的磷酸化所必需的，也可能是赋予其对雷帕霉素和氨基酸充足的敏感性所必需的。

The mammalian target of rapamycin (mTOR) is a protein kinase that controls multiple cellular functions, including cell growth, in response to amino acids and growth factors, in part by regulating the phosphorylation of p70 S6 kinase (p70S6k) and eukaryotic initiation factor 4E-binding protein 1 (4B-BP1). The mechanisms by which mTOR regulates p70S6k and 4B-BP1 in vivo remain incompletely understood. We identified a novel mTOR-binding partner, raptor (regulatory associated protein of mTOR) that also binds p70S6k and 4E-BP1 and is essential for TOR signaling in vivo. We also demonstrated that raptor binds to p70S6k and 4E-BP1 through their respective TOS (conserved TOR signaling) motife, a short conserved segment previously shown to be required for amino acid- and mTOR-dependent regulation of these mTOR substrates in vivo. A point mutation within the TOS motif of p70S6k or 4E-BP1 known to abolish both amino acid- and mTOR-regulation, selectively abolishes their binding to raptor. This mutation of the TOS motif also eliminates all in vitro mTOR-catalyzed 4E-BP1 phosphorylation and abolishes the raptor-dependent component of mTOR-catalyzed p70S6k phosphorylation in vitrv. Raptor does not alter mTOR's intrinsic catalytic activity, but appears to serve as an mTOR scaffold protein whose binding to the TOS motif of mTOR substrates is necessary for effective mTOR-catalyzed phosphorylation in vivo and perhaps for conferring their sensitivity to rapamycin and amino acid sufficiency.