The mammalian target of rapamycin (mTOR) is a conserved Ser/Thr kinase that forms two functionally distinct complexes important for nutrient and growth factor signaling. While mTOR complex 1 (mTORC1) regulates mRNA translation and ribosome biogenesis, mTORC2 plays an important role in the phosphorylation and subsequent activation of Akt. Interestingly, mTORC1 negatively regulates Akt activation, but whether mTORC1 signaling directly targets mTORC2 remains unknown. Here we show that growth factors promote the phosphorylation of Rictor (rapamycin-insensitive companion of mTOR), an essential subunit of mTORC2. We found that Rictor phosphorylation requires mTORC1 activity and, more specifically, the p70 ribosomal S6 kinase 1 (S6K1). We identified several phosphorylation sites in Rictor and found that Thr1135 is directly phosphorylated by S6K1 in vitro and in vivo, in a rapamycin-sensitive manner. Phosphorylation of Rictor on Thr1135 did not affect mTORC2 assembly, kinase activity, or cellular localization. However, cells expressing a Rictor T1135A mutant were found to have increased mTORC2-dependent phosphorylation of Akt. In addition, phosphorylation of the Akt substrates FoxO1/3a and glycogen synthase kinase 3␣/ (GSK3␣/) was found to be increased in these cells, indicating that S6K1-mediated phosphorylation of Rictor inhibits mTORC2 and Akt signaling. Together, our results uncover a new regulatory link between the two mTOR complexes, whereby Rictor integrates mTORC1-dependent signaling.
Disruption of NAD synthesis caused a deficiency of NAD and congenital malformations in humans and mice. Niacin supplementation during gestation prevented the malformations in mice. (Funded by the National Health and Medical Research Council of Australia and others.).
The Ras/mitogen-activated protein kinase (MAPK) pathway regulates a variety of cellular processes by activating specific transcriptional and translational programs. Ras/MAPK signaling promotes mRNA translation and protein synthesis, but the exact molecular mechanisms underlying this regulation remain poorly understood. Increasing evidence suggests that the mammalian target of rapamycin (mTOR) plays an essential role in this process. Here, we show that Raptor, an essential scaffolding protein of the mTOR complex 1 (mTORC1), becomes phosphorylated on proline-directed sites following activation of the Ras/MAPK pathway. We found that ERK1 and ERK2 interact with Raptor in cells and mediate its phosphorylation in vivo and in vitro. Using mass spectrometry and phosphospecific antibodies, we found three proline-directed residues within Raptor, Ser 8 , Ser 696 , and Ser 863 , which are directly phosphorylated by ERK1/2. Expression of phosphorylationdeficient alleles of Raptor revealed that phosphorylation of these sites by ERK1/2 normally promotes mTORC1 activity and signaling to downstream substrates, such as 4E-BP1. Our data provide a novel regulatory mechanism by which mitogenic and oncogenic activation of the Ras/MAPK pathway promotes mTOR signaling.The Ras/mitogen-activated protein kinase (MAPK) pathway regulates a variety of cellular processes, including cell growth, proliferation, survival, and motility, through the activation of specific transcriptional and translational programs (1-3). The first evidence linking Ras/MAPK signaling to the regulation of mRNA translation stemmed from the finding that inhibition of MEK1/2 blocks growth factor-stimulated global protein synthesis (4). Subsequently, the MAPK-activated protein kinases MNK1/2 were found to phosphorylate the translation initiation factor eIF4E (5, 6), providing a potential mechanism by which MAPK signaling regulates mRNA translation (reviewed in 7, 8). However, more recent evidence indicates that the MNKs are dispensable for global protein synthesis (9). Thus, the functional impact of eIF4E phosphorylation remains unclear and a matter of debate (8, 10).Increasing evidence suggests that Ras/MAPK signaling largely promotes protein synthesis through the regulation of the mammalian target of rapamycin (mTOR), 6 an important regulator of cell growth and proliferation (reviewed in 11-13). mTOR, a member of the phosphoinositide 3-kinase-related protein kinase family, senses and integrates signals from diverse environmental cues to regulate crucial biosynthetic processes involved in cell growth and proliferation (reviewed in 14 -16). mTOR associates with different protein partners to form two functionally distinct signaling complexes, the rapamycin-sensitive and rapamycin-insensitive mTOR complex (mTORC) 1 and 2, respectively (17-19). mTORC1 contains the catalytic subunit mTOR, mLST8/GL, PRAS40, and Raptor (17,[20][21][22][23][24]. Raptor directly binds mTOR and is thought to function as a scaffolding protein that recruits mTORC1 substrates through their TOR sig...
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