Lithium Can Relieve Translational Repression of TOP mRNAs Elicited by Various Blocks along the Cell Cycle in a Glycogen Synthase Kinase-3- and S6-Kinase-independent Manner

112

The mTOR signaling complex integrates signals from growth factors and nutrient availability to control cell growth and proliferation, in part through effects on the protein-synthetic machinery. Protein synthesis rates fluctuate throughout the cell cycle but diminish significantly during the G 2 /M transition. The fate of the mTOR complex and its role in coordinating cell growth and proliferation signals with protein synthesis during mitosis remain unknown. Here we demonstrate that the mTOR complex 1 (mTORC1) pathway, which stimulates protein synthesis, is actually hyperactive during mitosis despite decreased protein synthesis and reduced activity of mTORC1 upstream activators. We describe previously unknown G 2 /M-specific phosphorylation of a component of mTORC1, the protein raptor, and demonstrate that mitotic raptor phosphorylation alters mTORC1 function during mitosis. Phosphopeptide mapping and mutational analysis demonstrate that mitotic phosphorylation of raptor facilitates cell cycle transit through G 2 /M. Phosphorylation-deficient mutants of raptor cause cells to delay in G 2 /M, whereas depletion of raptor causes cells to accumulate in G 1 . We identify cyclindependent kinase 1 (cdk1 [cdc2]) and glycogen synthase kinase 3 (GSK3) pathways as two probable mitosisregulated protein kinase pathways involved in mitosis-specific raptor phosphorylation and altered mTORC1 activity. In addition, mitotic raptor promotes translation by internal ribosome entry sites (IRES) on mRNA during mitosis and is demonstrated to be associated with rapamycin resistance. These data suggest that this pathway may play a role in increased IRES-dependent mRNA translation during mitosis and in rapamycin insensitivity.

Section: Resultsmentioning

confidence: 56%

Mitotic Raptor Promotes mTORC1 Activity, G₂/M Cell Cycle Progression, and Internal Ribosome Entry Site-Mediated mRNA Translation

Ramírez‐Valle

Badura

Braunstein

et al. 2010

112

“…8B). Similar observations have been reported for nonneuronal cells and suggest that neither S6K1 nor S6K2 is required for the induction of S6 phosphorylation or the synthesis of 5ЈTOP-encoded proteins (36,46,48). Also important to consider is that we were not able to measure LTD-associated increases in S6 phosphorylation and EF1A levels in S6K1/S6K2 double knockout mice.…”

Section: Discussionmentioning

confidence: 57%

mGluR-Dependent Long-Term Depression Is Associated with Increased Phosphorylation of S6 and Synthesis of Elongation Factor 1A but Remains Expressed in S6K-Deficient Mice

Antion¹,

Hou

Wong

et al. 2008

103

102

Metabotropic glutamate receptor-dependent long-term depression (mGluR-LTD) in the hippocampus requires rapid protein synthesis, which suggests that mGluR activation is coupled to signaling pathways that regulate translation. Herein, we have investigated the signaling pathways that couple group I mGluRs to ribosomal S6 protein phosphorylation and 5oligopyrimidine tract (5TOP)-encoded protein synthesis during mGluR-LTD. We found that mGluR-LTD was associated with increased phosphorylation of p70S6 kinase (S6K1) and S6, as well as the synthesis of the 5TOP-encoded protein elongation factor 1A (EF1A). Moreover, we found that LTD-associated increases in S6K1 phosphorylation, S6 phosphorylation, and levels of EF1A were sensitive to inhibitors of phosphoinositide 3-kinase (PI3K), mammalian target of rapamycin (mTOR), and extracellular signal-regulated kinase (ERK). However, mGluR-LTD was normal in S6K1 knockout mice and enhanced in both S6K2 knockout mice and S6K1/S6K2 double knockout mice. In addition, we observed that LTD-associated increases in S6 phosphorylation were still increased in S6K1-and S6K2-deficient mice, whereas basal levels of EF1A were abnormally elevated. Taken together, these findings indicate that mGluR-LTD is associated with PI3K-, mTOR-, and ERK-dependent alterations in the phosphorylation of S6 and S6K. Our data also suggest that S6Ks are not required for the expression of mGluR-LTD and that the synthesis of 5TOP-encoded proteins is independent of S6Ks during mGluR-LTD.The activation of group I metabotropic glutamate receptors (mGluRs) is intimately involved in the regulation of protein synthesis in neurons. Brief application of the mGluR I agonist [3,5-RS] dihydroxyphenylhydrazine (DHPG) has been reported to enhance de novo protein synthesis in hippocampal slices (37) and synaptoneurosomes (12), alter mRNA granule distribution (3), enhance polyribosome formation in synaptoneurosomes (12, 52), and promote rapid dendritic protein synthesis during mGluRdependent long-term depression (mGluR-LTD) (19). These findings strongly suggest that mGluR-mediated signaling is directly coupled to the regulation of translation initiation in neurons.Multiple signaling pathways critical for regulating protein synthesis have been reported to be required for mGluR-LTD. Both mammalian target of rapamycin (mTOR) and extracellular signal-regulated kinase (ERK) are activated during mGluR-LTD, and inhibitors of these kinases, as well as inhibitors of phosphoinositide-3-kinase (PI3K), have been shown to block mGluR-LTD (4, 9, 16). Substrates of mTOR and ERK that are known to be involved in translational control during mGluR-LTD include eukaryotic initiation factor 4E and its repressor 4E-binding protein (4). S6 kinases also are known to integrate mTOR, PI3K, and ERK signaling to influence protein synthesis; however, the requirement of S6 kinases for the expression of mGluR-LTD has not been examined.In mammals, S6 kinases are encoded by two separate genes, S6K1 and S6K2, which are approximately 80% homologous (8). One of t...

“…The fact that the translation of TOP mRNAs is repressed upon mitotic arrest by any means (48) and that rapamycin can completely block progression through the cell cycle, at least in some cell types (1,38,45), suggested a causal relationship between the effect of rapamycin on cell cycle progression and on TOP mRNAs. However, the lack of any correlation between the effects of rapamycin on the mitogenic activity of HEK 293 and 3T3-L1 cells (data not shown) on the one hand and on the translation efficiency of TOP mRNAs in these cell lines on the other hand ( Fig.…”

Section: Resultsmentioning

confidence: 99%

“…TOP mRNAs are characterized by an oligopyrimidine tract at the 5Ј terminus and encode ribosomal proteins, elongation factors, and several other proteins associated with the assembly or function of the translational apparatus (34). The translation of these mRNAs is selectively repressed when proliferation of vertebrate cells is blocked at various phases of the cell cycle by a wide variety of physiological signals and pharmacological treatments or when they are amino acid starved (48). It has previously been shown that translational activation of TOP mRNAs is strictly dependent on the integrity of the phosphatidylinositol 3-kinase (PI3K) pathway (49,50).…”

mentioning

confidence: 99%

The TSC-mTOR Pathway Mediates Translational Activation of TOP mRNAs by Insulin Largely in a Raptor- or Rictor-Independent Manner

Patursky-Polischuk

Stolovich-Rain

Hausner-Hanochi

et al. 2009

Self Cite

114

The stimulatory effect of insulin on protein synthesis is due to its ability to activate various translation factors. We now show that insulin can increase protein synthesis capacity also by translational activation of TOP mRNAs encoding various components of the translation machinery. This translational activation involves the tuberous sclerosis complex (TSC), as the knockout of TSC1 or TSC2 rescues TOP mRNAs from translational repression in mitotically arrested cells. Similar results were obtained upon overexpression of Rheb, an immediate TSC1-TSC2 target. The role of mTOR, a downstream effector of Rheb, in translational control of TOP mRNAs has been extensively studied, albeit with conflicting results. Even though rapamycin fully blocks mTOR complex 1 (mTORC1) kinase activity, the response of TOP mRNAs to this drug varies from complete resistance to high sensitivity. Here we show that mTOR knockdown blunts the translation efficiency of TOP mRNAs in insulin-treated cells, thus unequivocally establishing a role for mTOR in this mode of regulation. However, knockout of the raptor or rictor gene has only a slight effect on the translation efficiency of these mRNAs, implying that mTOR exerts its effect on TOP mRNAs through a novel pathway with a minor, if any, contribution of the canonical mTOR complexes mTORC1 and mTORC2. This conclusion is further supported by the observation that raptor knockout renders the translation of TOP mRNAs rapamycin hypersensitive.