Serotonin Increases Phosphorylation of Synaptic 4EBP through TOR, but Eukaryotic Initiation Factor 4E Levels Do Not Limit Somatic Cap-Dependent Translation in <i>Aplysia</i> Neurons

Carroll, Matthew; Dyer, John; Sossin, Wayne S.

doi:10.1128/mcb.00955-06

Cited by 31 publications

(40 citation statements)

References 68 publications

(95 reference statements)

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“…However, the presence of a substantial amount of 4E-BP1 that is phosphorylated on Thr 37/46 , Ser 65 and Thr 70 but which remains of normal size in the presence of calyculin A (Figures 2, 4-6) suggests that, although phosphorylation may be necessary, it is not sufficient for polyubiquitination. It remains to be determined whether the effects of calyculin A are due to inhibition of the dephosphorylation of 4E-BP1 itself or to an ability to enhance the activity of mTOR (Carroll et al, 2006). Our data suggest that, under some circumstances, stimuli that promote the phosphorylation of 4E-BP1 Effects of calyculin A and MG132 on the stability of 4E-BP1 and conversion to high-molecular-weight forms.…”

Section: Phosphorylation Ubiquitination and Stability Of 4e-bp1mentioning

confidence: 82%

Effects of protein phosphorylation on ubiquitination and stability of the translational inhibitor protein 4E-BP1

2007

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The availability of the eukaryotic polypeptide chain initiation factor 4E (eIF4E) for protein synthesis is regulated by the 4E-binding proteins (4E-BPs), which act as inhibitors of cap-dependent mRNA translation. The ability of the 4E-BPs to sequester eIF4E is regulated by reversible phosphorylation at multiple sites. We show here that, in addition, 4E-BP1 is a substrate for polyubiquitination and that some forms of 4E-BP1 are simultaneously polyubiquitinated and phosphorylated. In Jurkat cells inhibition of proteasomal activity by MG132 enhances the level of hypophosphorylated, unmodified 4E-BP1 but only modestly increases the accumulation of high-molecularweight, phosphorylated forms of 4E-BP1. In contrast, inhibition of protein phosphatase activity with calyculin A reduces the level of unmodified 4E-BP1 but strongly enhances the amount of phosphorylated, high-molecularweight 4E-BP1. Turnover measurements in the presence of cycloheximide show that, whereas 4E-BP1 is normally a very stable protein, calyculin A decreases the apparent half-life of the normal-sized protein. Affinity chromatography on m 7 GTP-Sepharose indicates that the larger forms of 4E-BP1 bind very poorly to eIF4E. We suggest that the phosphorylation of 4E-BP1 may play a dual role in the regulation of protein synthesis, both reducing the affinity of 4E-BP1 for eIF4E and promoting the conversion of 4E-BP1 to alternative, polyubiquitinated forms.

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Section: Phosphorylation Ubiquitination and Stability Of 4e-bp1mentioning

confidence: 82%

Effects of protein phosphorylation on ubiquitination and stability of the translational inhibitor protein 4E-BP1

2007

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“…In this idealized model, the ability of 4E-BP to bind to eIF4E and prevent initiation of cap-dependent translation is inhibited by TORC1-mediated phosphorylation, while S6K is activated by TORC1-mediated phosphorylation. However, a previous study from our laboratory suggests that 4E-BP binding to eIF4E may not be rate-limiting for the initiation of translation in the Aplysia SN soma unless wild-type 4E-BP is overexpressed (11). In Aplysia, S6K has two identified downstream targets: eEF2K, through which it regulates translational elongation via disinhibition of eEF2, and the ribosomal protein S6.…”

Section: Resultsmentioning

confidence: 99%

“…We have previously characterized a form of Aplysia 4E-BP that lacks the carboxyl-terminal TOS motif (4E-BP(⌬TOS)), and expression of this construct is capable of inhibiting the initiation of cap-dependent translation, thus, providing a means of specifically interfering with TORC1 signaling through 4E-BP (Fig. 1C) (11). Here we use a similar strategy to generate a dominant negative S6K construct that should specifically disrupt the S6K pathway but not other TORC1 downstream effectors (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The dominant negative 4E-BP construct (4E-BP(⌬TOS)) has previously been described (11). To generate the dominant negative S6K construct (S6K(⌬TOS)-mRFP), S6K was amplified by PCR from BB4-S6K (9) using primers containing XbaI and BamHI sites.…”

Section: Generation Of Constructs and Cloning Of Aplysia Rheb-mentioning

confidence: 99%

“…Although 4E-BP regulates translation of all capped mRNAs, some mRNAs are particularly sensitive to levels of free eIF4E (4,16) and, therefore, should also be particularly sensitive to 4E-BP regulation. 4E-BP is phosphorylated by stimuli that induce late long-term potentiation (L-LTP), metabotropic glutamate receptor-dependent long-term depression (mGluR-LTD), and, in Aplysia, LTF (11,(17)(18)(19). Mice lacking the major isoform of 4E-BP in the brain have a decreased threshold for the induction of L-LTP (18) and display a form of mGluR-LTD that is no longer sensitive to rapamycin (17), suggesting that, in the absence of 4E-BP, proteins important for plasticity are translated in the absence of stimulation.…”

Section: Long-term Facilitation (Ltf) Inmentioning

confidence: 99%

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Ribosomal Protein S6 Kinase Is a Critical Downstream Effector of the Target of Rapamycin Complex 1 for Long-term Facilitation in Aplysia

Weatherill

Dyer

Sossin

2010

Journal of Biological Chemistry

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Long-term facilitation (LTF) inAplysia is a leading cellular model for elucidating the biochemical mechanisms of synaptic plasticity underlying learning. In Aplysia, LTF requires translational control downstream of the target of rapamycin (TOR) complex 1 (TORC1). The major known downstream targets of TORC1 are 4E binding protein (4E-BP) and S6 kinase (S6K). By removing the site within these regulators required for their interaction with TORC1, we have generated dominant negative proteins that disrupt specific pathways downstream of TORC1. Expression of dominant negative S6K, but not dominant negative 4E-BP, in Aplysia sensory neurons (SNs) blocked 24-h LTF. TORC1 is directly activated by the small GTP-binding protein, Ras homologue enriched in brain (Rheb). To determine the effects of TORC1 activation on translation in Aplysia neurons, we have examined the effects of expressing a constitutively active form of the Aplysia orthologue of Rheb, ApRheb (ApRheb(Q63L)). Expression of ApRheb(Q63L) increased 4E-BP phosphorylation and the level of general, cap-dependent translation within the SN cell soma in a rapamycin-sensitive manner. This increase in cap-dependent translation was blocked neither by dominant negative 4E-BP nor dominant negative S6K. Thus, we demonstrate that S6K is an important downstream target of TORC1 in Aplysia and that it is necessary for 24-h LTF, but not for TORC1-mediated increases in somatic cap-dependent translation.Memories, changes in synaptic strength within a neural network, are composed of mechanistically distinct phases. Whereas short-term memories rely on post-translational modification of pre-existing synaptic proteins, more persistent memories require the production of new proteins and likely involve synaptic growth (1). This de novo protein synthesis within neurons, underlying long-term synaptic plasticity, involves, not only transcriptional regulation, but translational regulation as well (2, 3). Moreover, the protein kinase complex, target of rapamycin (TOR) 3 complex 1 (TORC1), a major regulator of translation and growth in eukaryotic cells (4), has been shown to play an essential role in this process (5). In many cell types, TORC1 is activated when conditions are permissive for cell growth through the integration of signaling pathways that sense the presence of these permissive cues (growth factors, amino acids, and energy (ATP)) (4). In neurons, TORC1 is activated during both in vitro models of synaptic plasticity and in vivo models of memory formation and, similar to other cells, acts as a gatekeeper regulating neuronal growth and plasticity (6).The facilitation of neurotransmitter release at the sensoryto-motor neuron (SN-MN) synapse, in the mollusk, Aplysia californica, is a leading model system for the characterization of the biochemical basis of memory formation, because an increase in the strength of this synapse has been shown to contribute to behavioral sensitization of the reflex (7). In particular, whereas short-term facilitation (STF) is independent of protein synthe...

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mTOR kinase, a key player in the regulation of glial functions: Relevance for the therapy of multiple sclerosis

Russo¹,

Lisi²,

Feinstein

et al. 2012

Glia

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The mammalian target of rapamycin (mTOR) is a serine/threonine protein kinase with a central role in the regulation of cell growth and proliferation, and several intracellular processes, such as mRNA transcription and translation, autophagy and cytoskeletal organization. The relevance of this pathway in the regulation of the immune system is well characterized. mTOR is essential for the proper activation and proliferation of effector T cells, restricts the development of regulatory T cells, and downregulates innate immune responses. Recently, a direct role of mTOR in the modulation of glial functions has also been recognized. Data from our group and others support the notion that mTOR is involved in microglial proinflammatory activation. The kinase regulates several intracellular processes in astrocytes, among which the rate of mRNA degradation of the inducible form of NO synthase. Therefore, the inhibition of mTOR kinase activity in glial cells results in anti-inflammatory actions, suggesting possible beneficial effects of mTOR inhibitors (like rapamycin) in the treatment of inflammatory-based pathologies of the central nervous system. In contrast, mTOR plays an important role in the regulation of oligodendrocyte development and myelination process as well as several neuronal functions, which may limit this therapeutic approach. Nevertheless, as reviewed here, there is robust evidence that rapamycin ameliorates the clinical course of both the relapsing-remitting and the chronic experimental autoimmune encephalomyelitis (EAE), and significantly reduces the hyperalgesia observed before clinical development of EAE. These findings may have important clinical implications for the therapy of multiple sclerosis.

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Serotonin Increases Phosphorylation of Synaptic 4EBP through TOR, but Eukaryotic Initiation Factor 4E Levels Do Not Limit Somatic Cap-Dependent Translation in Aplysia Neurons

Cited by 31 publications

References 68 publications

Effects of protein phosphorylation on ubiquitination and stability of the translational inhibitor protein 4E-BP1

Effects of protein phosphorylation on ubiquitination and stability of the translational inhibitor protein 4E-BP1

Ribosomal Protein S6 Kinase Is a Critical Downstream Effector of the Target of Rapamycin Complex 1 for Long-term Facilitation in Aplysia

mTOR kinase, a key player in the regulation of glial functions: Relevance for the therapy of multiple sclerosis

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