The expanding TOR signaling network

Cell growth (increase in cell mass or size) is tightly coupled to nutrient availability, growth factors and the energy status of the cell. The target of rapamycin (TOR) integrates all three inputs to control cell growth. The discovery of upstream regulators of TOR (AMPK, the TSC1-TSC2 complex and Rheb) has provided new insights into the mechanism by which TOR integrates its various inputs. A recent finding in flies reveals that TOR controls not only growth of the cell in which it resides (cell-autonomous growth) but also the growth of distant cells, thereby determining organ and organism size in addition to the size of isolated cells. In yeast and mammals, the identification of two structurally and functionally distinct multiprotein TOR complexes (TORC1 and TORC2) has provided a molecular basis for the complexity of TOR signaling. Furthermore, TOR has emerged as a regulator of growth-related processes such as development, aging and the response to hypoxia. Thus, TOR is part of an intra- and inter-cellular signaling network with a remarkably broad role in eukaryotic biology.     

雷帕霉素靶蛋白与自噬在衰老中的交互作用

衰老是一个非常复杂的过程,与细胞和组织中累积的各种大分子（DNA、蛋白质和脂质）损伤密不可分,并且是由细胞中不同的信号通道共同调控的结果,而雷帕霉素靶标途径就是其中的一种。该途径整合了各种来自细胞内外的信号以调控细胞的生长、增殖和代谢。越来越多证据表明,雷帕霉素靶蛋白（target of rapamycin,TOR）控制着细胞和组织老化的速度,影响着整个机体衰老过程。另外TOR参与调控自噬的发生,而自噬能使生物大分子和细胞器降解并回收重复利用。多种生物模型研究发现,衰老其实是与自噬的不足有关联。本文对TOR和自噬在衰老过程中的作用和相互关系进行综述,为发展与老年疾病相关的新型治疗方法提供思路。     

The TOR nutrient signalling pathway phosphorylates NPR1 and inhibits turnover of the tryptophan permease.

The Saccharomyces cerevisiae targets of rapamycin, TOR1 and TOR2, signal activation of cell growth in response to nutrient availability. Loss of TOR or rapamycin treatment causes yeast cells to arrest growth in early G1 and to express several other physiological properties of starved (G0) cells. As part of this starvation response, high affinity amino acid permeases such as the tryptophan permease TAT2 are targeted to the vacuole and degraded. Here we show that the TOR signalling pathway phosphorylates the Ser/Thr kinase NPR1 and thereby inhibits the starvation-induced turnover of TAT2. Overexpression of NPR1 inhibits growth and induces the degradation of TAT2, whereas loss of NPR1 confers resistance to rapamycin and to FK506, an inhibitor of amino acid import. NPR1 is controlled by TOR and the type 2A phosphatase-associated protein TAP42. First, overexpression of NPR1 is toxic only when TOR function is reduced. Secondly, NPR1 is rapidly dephosphorylated in the absence of TOR. Thirdly, NPR1 dephosphorylation does not occur in a rapamycin-resistant tap42 mutant. Thus, the TOR nutrient signalling pathway also controls growth by inhibiting a stationary phase (G0) programme. The control of NPR1 by TOR is analogous to the control of p70 s6 kinase and 4E-BP1 by mTOR in mammalian cells.     

酵母TOR信号转导途径

TOR(target of rapamycin)是真核细胞中一种高度保守的与磷脂酰肌醇激酶相关的蛋白激酶(PIKK),它是免疫抑制剂/抗癌药物雷帕霉素(rapamycin)的靶物质。TOR是细胞生长的中枢控制因子,外界营养因素通过TOR的作用控制酵母、果蝇和哺乳动物细胞的生长。TOR根据细胞环境的营养条件做出相应的应答,参与调控蛋白激酶和蛋白磷酸酯酶的活性,从而控制与蛋白质合成和基因转录相关基因的表达。现对酵母细胞中TOR信号转导途径的研究进行简明的阐述。     

The TOR and EGO protein complexes orchestrate microautophagy in yeast

The rapamycin-sensitive TOR signaling pathway in Saccharomyces cerevisiae positively controls cell growth in response to nutrient availability. Accordingly, TOR depletion or rapamycin treatment causes regulated entry of cells into a quiescent growth phase. Although this process has been elucidated in considerable detail, the transition from quiescence back to proliferation is poorly understood. Here, we describe the identification of a conserved member of the RagA subfamily of Ras-related GTPases, Gtr2, which acts in a vacuolar membrane-associated protein complex together with Ego1 and Ego3 to ensure proper exit from rapamycin-induced growth arrest. We demonstrate that the EGO complex, in conjunction with TOR, positively regulates microautophagy, thus counterbalancing the massive rapamycin-induced, macroautophagy-mediated membrane influx toward the vacuolar membrane. Moreover, large-scale genetic analyses of the EGO complex confirm the existence of a growth control mechanism originating at the vacuolar membrane and pinpoint the amino acid glutamine as a key metabolite in TOR signaling.     

Nutrient sensing and TOR signaling in yeast and mammals

Coordinating cell growth with nutrient availability is critical for cell survival. The evolutionarily conserved TOR (target of rapamycin) controls cell growth in response to nutrients, in particular amino acids. As a central controller of cell growth, mTOR (mammalian TOR) is implicated in several disorders, including cancer, obesity, and diabetes. Here, we review how nutrient availability is sensed and transduced to TOR in budding yeast and mammals. A better understanding of how nutrient availability is transduced to TOR may allow novel strategies in the treatment for mTOR‐related diseases.     

Regulation of the cell integrity pathway by rapamycin-sensitive TOR function in budding yeast

The TOR (target of rapamycin) pathway controls cell growth in response to nutrient availability in eukaryotic cells. Inactivation of TOR function by rapamycin or nutrient exhaustion is accompanied by triggering various cellular mechanisms aimed at overcoming the nutrient stress. Here we report that in Saccharomyces cerevisiae the protein kinase C (PKC)-mediated mitogen-activated protein kinase pathway is regulated by TOR function because upon specific Tor1 and Tor2 inhibition by rapamycin, Mpk1 is activated rapidly in a process mediated by Sit4 and Tap42. Osmotic stabilization of the plasma membrane prevents both Mpk1 activation by rapamycin and the growth defect that occurs upon the simultaneous absence of Tor1 and Mpk1 function, suggesting that, at least partially, TOR inhibition is sensed by the PKC pathway at the cell envelope. This process involves activation of cell surface sensors, Rom2, and downstream elements of the mitogen-activated protein kinase cascade. Rapamycin also induces depolarization of the actin cytoskeleton through the TOR proteins, Sit4 and Tap42, in an osmotically suppressible manner. Finally, we show that entry into stationary phase, a physiological situation of nutrient depletion, also leads to the activation of the PKC pathway, and we provide further evidence demonstrating that Mpk1 is essential for viability once cells enter G(0).     

The Arabidopsis THADA homologue modulates TOR activity and cold acclimation

• Low temperature is one of the most important environmental factors that affect global survival of humans and animals and equally importantly the distribution of plants and crop productivity. Survival of metazoan cells under cold stress requires regulation of the sensor‐kinase Target Of Rapamycin (TOR). TOR controls growth of eukaryotic cells by adjusting anabolic and catabolic metabolism. Previous studies identified the Thyroid Adenoma Associated (THADA) gene as the major effect locus by positive selection in the evolution of modern human adapted to cold. Here we investigate the role of THADA in TOR signaling and cold acclimation of plants.
• We applied BLAST searches and homology modeling to identify the AtTHADA (AT3G55160) in Arabidopsis thaliana as the highly probable orthologue protein. Reverse genetics approaches were combined with immunological detection of TOR activity and metabolite profiling to address the role of the TOR and THADA for growth regulation and cold acclimation.
• Depletion of the AtTHADA gene caused complete or partial loss of full‐length mRNA, respectively, and significant retardation of growth under non‐stressed conditions. Furthermore, depletion of AtTHADA caused hypersensitivity towards low‐temperatures. Atthada displayed a lowered energy charge. This went along with decreased TOR activity, which offers a molecular explanation for the slow growth phenotype of Atthada. Finally, we used TOR RNAi lines to identify the de‐regulation of TOR activity as one determinant for sensitivity towards low‐temperatures.
• Taken together our results provide evidence for a conserved function of THADA in cold acclimation of eukaryotes and suggest that cold acclimation in plants requires regulation of TOR.

     

Elucidating TOR signaling and rapamycin action: lessons from Saccharomyces cerevisiae.

TOR (target of rapamycin) is a phosphatidylinositol kinase-related protein kinase that controls cell growth in response to nutrients. Rapamycin is an immunosuppressive and anticancer drug that acts by inhibiting TOR. The modes of action of TOR and rapamycin are remarkably conserved from S. cerevisiae to humans. The current understanding of TOR and rapamycin is derived largely from studies with S. cerevisiae. In this review, we discuss the contributions made by S. cerevisiae to understanding rapamycin action and TOR function.     

Fission Yeast SCYL1/2 Homologue Ppk32: A Novel Regulator of TOR Signalling That Governs Survival during Brefeldin A Induced Stress to Protein Trafficking

Target of Rapamycin (TOR) signalling allows eukaryotic cells to adjust cell growth in response to changes in their nutritional and environmental context. The two distinct TOR complexes (TORC1/2) localise to the cell’s internal membrane compartments; the endoplasmic reticulum (ER), Golgi apparatus and lysosomes/vacuoles. Here, we show that Ppk32, a SCYL family pseudo-kinase, is a novel regulator of TOR signalling. The absence of ppk32 expression confers resistance to TOR inhibition. Ppk32 inhibition of TORC1 is critical for cell survival following Brefeldin A (BFA) induced stress. Treatment of wild type cells with either the TORC1 specific inhibitor rapamycin or the general TOR inhibitor Torin1 confirmed that a reduction in TORC1 activity promoted recovery from BFA induced stress. Phosphorylation of Ppk32 on two residues that are conserved within the SCYL pseudo-kinase family are required for this TOR inhibition. Phosphorylation on these sites controls Ppk32 protein levels and sensitivity to BFA. BFA induced ER stress does not account for the response to BFA that we report here, however BFA is also known to induce Golgi stress and impair traffic to lysosomes. In summary, Ppk32 reduce TOR signalling in response to BFA induced stress to support cell survival.     

Elucidating TOR Signaling and Rapamycin Action: Lessons from Saccharomyces cerevisiae

TOR (target of rapamycin) is a phosphatidylinositol kinase-related protein kinase that controls cell growth in response to nutrients. Rapamycin is an immunosuppressive and anticancer drug that acts by inhibiting TOR. The modes of action of TOR and rapamycin are remarkably conserved from S. cerevisiae to humans. The current understanding of TOR and rapamycin is derived largely from studies with S. cerevisiae. In this review, we discuss the contributions made by S. cerevisiae to understanding rapamycin action and TOR function.     

mTOR is essential for growth and proliferation in early mouse embryos and embryonic stem cells

TOR is a serine-threonine kinase that was originally identified as a target of rapamycin in Saccharomyces cerevisiae and then found to be highly conserved among eukaryotes. In Drosophila melanogaster, inactivation of TOR or its substrate, S6 kinase, results in reduced cell size and embryonic lethality, indicating a critical role for the TOR pathway in cell growth control. However, the in vivo functions of mammalian TOR (mTOR) remain unclear. In this study, we disrupted the kinase domain of mouse mTOR by homologous recombination. While heterozygous mutant mice were normal and fertile, homozygous mutant embryos died shortly after implantation due to impaired cell proliferation in both embryonic and extraembryonic compartments. Homozygous blastocysts looked normal, but their inner cell mass and trophoblast failed to proliferate in vitro. Deletion of the C-terminal six amino acids of mTOR, which are essential for kinase activity, resulted in reduced cell size and proliferation arrest in embryonic stem cells. These data show that mTOR controls both cell size and proliferation in early mouse embryos and embryonic stem cells.     

Target of rapamycin (TOR) signaling controls epithelial morphogenesis in the vertebrate intestine

The target of rapamycin (TOR) signaling pathway regulates cell growth and proliferation, however the extent to which TOR signaling mediates particular organogenesis programs remains to be determined. Here we report an examination of TOR signaling during zebrafish development, using a combination of small molecule treatment and morpholino-mediated gene knockdown. First, we amplified and sequenced the full-length cDNA for the zebrafish TOR ortholog (ztor). By in situ hybridization, we found that ztor is expressed ubiquitously in the early embryo, but displays a dynamic pattern in the gut between 48 and 72 h post-fertilization (hpf). Treatment of zebrafish embryos with rapamycin induced only a mild general developmental delay up to 72 hpf, but digestive tract development became arrested at the primitive gut tube stage. Rapamycin inhibited intestinal epithelial growth, morphogenesis and differentiation. Using morpholino-mediated gene knockdown of TOR pathway components, we show that this effect is mediated specifically by the rapamycin-sensitive TOR complex 1 (TORC1). Thus, in addition to regulating cell growth and proliferation, TOR signaling controls the developmental program guiding epithelial morphogenesis in the vertebrate intestine.     

Role and regulation of starvation-induced autophagy in the Drosophila fat body

In response to starvation, eukaryotic cells recover nutrients through autophagy, a lysosomal-mediated process of cytoplasmic degradation. Autophagy is known to be inhibited by TOR signaling, but the mechanisms of autophagy regulation and its role in TOR-mediated cell growth are unclear. Here, we show that signaling through TOR and its upstream regulators PI3K and Rheb is necessary and sufficient to suppress starvation-induced autophagy in the Drosophila fat body. In contrast, TOR's downstream effector S6K promotes rather than suppresses autophagy, suggesting S6K downregulation may limit autophagy during extended starvation. Despite the catabolic potential of autophagy, disruption of conserved components of the autophagic machinery, including ATG1 and ATG5, does not restore growth to TOR mutant cells. Instead, inhibition of autophagy enhances TOR mutant phenotypes, including reduced cell size, growth rate, and survival. Thus, in cells lacking TOR, autophagy plays a protective role that is dominant over its potential role as a growth suppressor.     

Inhibition of target of rapamycin signaling by rapamycin in the unicellular green alga Chlamydomonas reinhardtii

The macrolide rapamycin specifically binds the 12-kD FK506-binding protein (FKBP12), and this complex potently inhibits the target of rapamycin (TOR) kinase. The identification of TOR in Arabidopsis (Arabidopsis thaliana) revealed that TOR is conserved in photosynthetic eukaryotes. However, research on TOR signaling in plants has been hampered by the natural resistance of plants to rapamycin. Here, we report TOR inactivation by rapamycin treatment in a photosynthetic organism. We identified and characterized TOR and FKBP12 homologs in the unicellular green alga Chlamydomonas reinhardtii. Whereas growth of wild-type Chlamydomonas cells is sensitive to rapamycin, cells lacking FKBP12 are fully resistant to the drug, indicating that this protein mediates rapamycin action to inhibit cell growth. Unlike its plant homolog, Chlamydomonas FKBP12 exhibits high affinity to rapamycin in vivo, which was increased by mutation of conserved residues in the drug-binding pocket. Furthermore, pull-down assays demonstrated that TOR binds FKBP12 in the presence of rapamycin. Finally, rapamycin treatment resulted in a pronounced increase of vacuole size that resembled autophagic-like processes. Thus, our findings suggest that Chlamydomonas cell growth is positively controlled by a conserved TOR kinase and establish this unicellular alga as a useful model system for studying TOR signaling in photosynthetic eukaryotes.     

Signaling by target of rapamycin proteins in cell growth control.

Target of rapamycin (TOR) proteins are members of the phosphatidylinositol kinase-related kinase (PIKK) family and are highly conserved from yeast to mammals. TOR proteins integrate signals from growth factors, nutrients, stress, and cellular energy levels to control cell growth. The ribosomal S6 kinase 1 (S6K) and eukaryotic initiation factor 4E binding protein 1(4EBP1) are two cellular targets of TOR kinase activity and are known to mediate TOR function in translational control in mammalian cells. However, the precise molecular mechanism of TOR regulation is not completely understood. One of the recent breakthrough studies in TOR signaling resulted in the identification of the tuberous sclerosis complex gene products, TSC1 and TSC2, as negative regulators for TOR signaling. Furthermore, the discovery that the small GTPase Rheb is a direct downstream target of TSC1-TSC2 and a positive regulator of the TOR function has significantly advanced our understanding of the molecular mechanism of TOR activation. Here we review the current understanding of the regulation of TOR signaling and discuss its function as a signaling nexus to control cell growth during normal development and tumorigenesis.