Structure of S6 kinase 1 determines whether raptor-mTOR or rictor-mTOR phosphorylates its hydrophobic motif site

Whitehead Institute for Biomedical Research and Massachusetts Institute of Technology, Department of Biology, Cambridge, 02142, USA.
Journal of Biological Chemistry (Impact Factor: 4.57). 06/2005; 280(20):19445-8. DOI: 10.1074/jbc.C500125200
Source: PubMed

ABSTRACT The mTOR protein kinase is the target of the immunosuppressive and anti-cancer drug rapamycin and is increasingly recognized as a key regulator of cell growth in mammals. S6 kinase 1 (S6K1) is the best characterized effector of mTOR, and its regulation serves as a model for mTOR signaling. Nutrients and growth factors activate S6K1 by inducing the phosphorylation of threonine 389 in the hydrophobic motif of S6K1. As phosphorylation of Thr(389) is rapamycin sensitive and mTOR can phosphorylate the same site in vitro, it has been suggested that mTOR is the physiological Thr(389) kinase. This proposal is not supported, however, by the existence of mutants of S6K1 that are phosphorylated in vivo on Thr(389) in a rapamycin-resistant fashion. Here, we demonstrate that the raptor-mTOR complex phosphorylates the rapamycin-sensitive forms of S6K1, while the distinct rictor-mTOR complex phosphorylates the rapamycin-resistant mutants of S6K1. Phosphorylation of Thr(389) by rictor-mTOR is independent of the TOR signaling motif and depends on removal of the carboxyl terminal domain of S6K1. Because many members of the AGC family of kinases lack an analogous domain, rictor-mTOR may phosphorylate the hydrophobic motifs of other kinases.

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    • "Here, we report that in human cells S6K1 has two alternatively spliced short isoforms that are overproduced in breast cancer cell lines and tumors. Furthermore, all of S6K1 short splicing variants lack an autoinhibitory C terminus domain (Ali and Sabatini, 2005) and half of the kinase domain and do not exhibit kinase activity, at least on the known S6K1 substrate rpS6 (Figures 2A and S2E–S2G). Overexpression of mouse or human S6K1 short isoforms enhanced transformation, anchorage-independent growth, cell motility, and growth-factorindependent 3D acinus formation of human breast epithelial cells (Figures 2, 3A–3C, S2C, and S3A). "
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    ABSTRACT: Ribosomal S6 kinase 1 (S6K1) is a major mTOR downstream signaling molecule that regulates cell size and translation efficiency. Here, we report that short isoforms of S6K1 are overproduced in breast cancer cell lines and tumors. Overexpression of S6K1 short isoforms induces transformation of human breast epithelial cells. The long S6K1 variant (Iso-1) induced opposite effects. It inhibits Ras-induced transformation and tumor formation, while its knockdown or knockout induces transformation, suggesting that Iso-1 has a tumor-suppressor activity. Furthermore, we found that S6K1 short isoforms bind and activate mTORC1, elevating 4E-BP1 phosphorylation, cap-dependent translation, and Mcl-1 protein levels. Both a phosphorylation-defective 4E-BP1 mutant and the mTORC1 inhibitor rapamycin partially blocked the oncogenic effects of S6K1 short isoforms, suggesting that these are mediated by mTORC1 and 4E-BP1. Thus, alternative splicing of S6K1 acts as a molecular switch in breast cancer cells, elevating oncogenic isoforms that activate mTORC1.
    Cell Reports 12/2012; 3(1). DOI:10.1016/j.celrep.2012.11.020 · 8.36 Impact Factor
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    • "Efficacy of mTOR inhibition was assessed by measuring the phosphorylation of the ribosomal protein S6, which is phosphorylated by the mTORC1 effector S6 kinase (Yang and Guan, 2007). Both rapamycin and mTOR depletion strongly reduced S6 phosphorylation, whereas raptor and rictor depletion each partially affected S6 phosphorylation (Fig. 4B), consistent with roles for both mTORC1 and mTORC2 in regulating S6 kinase (Ali and Sabatini, 2005). Wortmannin did not alter S6 phosphorylation , probably because at the concentration used (100 nM), it does not effectively block mTOR activity (Ballou and Lin, 2008). "
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    • "Both complexes have specific effects on distinct cellular functions, such as controlling mRNA translation, ribosome biogenesis, autophagy and metabolism [4] [5] [6]. mTORC2 phosphorylates AGC kinases such as AKT, serum-and glucocorticoid-induced protein kinase-1 (SGK1) and protein kinase C-alpha (PKCa) [7] [8] [9]. AKT is one of the best-known downstream effectors of phosphatidyli- nositol-3 kinase (PI3K). "
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