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The mTORC1 complex has been implicated in tumorigenesis owing partially to its ability to increase protein translation; now, mTORC1 activity in the mouse intestine is shown not to be required for normal homeostasis but to be necessary for the triggering of tumorigenesis by APC mutations, suggesting that it could be a good target for the prevention of colorectal cancer in high-risk patients. How mTORC sustains tumour growth The mTORC1 complex, a protein kinase complex found in all eukaryotic cells, has been implicated in tumorigenesis because it is known to stimulate protein translation. The main effector pathway downstream of mTORC1 is thought to be 4EBP1, which promotes initiation of translation. William Faller et al . now show that in the mouse intestine, mTORC1 activity is not required for normal homeostasis, but is absolutely required for intestinal tumour formation triggered by APC tumour suppressor gene mutations. The authors identify increased translational elongation downstream of S6 kinase via the elongation factor eEF2 as a requirement for proliferation in APC-deficient but not normal cells. This suggests that translational elongation, rather than initiation, is limiting to cancer cell proliferation in vivo . These findings raise the possibility that targeting mTORC1 signalling may be beneficial in prevention of colorectal cancers in high-risk patients. Inactivation of APC is a strongly predisposing event in the development of colorectal cancer 1 , 2 , prompting the search for vulnerabilities specific to cells that have lost APC function. Signalling through the mTOR pathway is known to be required for epithelial cell proliferation and tumour growth 3 , 4 , 5 , and the current paradigm suggests that a critical function of mTOR activity is to upregulate translational initiation through phosphorylation of 4EBP1 (refs 6 , 7 ). This model predicts that the mTOR inhibitor rapamycin, which does not efficiently inhibit 4EBP1 (ref. 8 ), would be ineffective in limiting cancer progression in APC-deficient lesions. Here we show in mice that mTOR complex 1 (mTORC1) activity is absolutely required for the proliferation of Apc -deficient (but not wild-type) enterocytes, revealing an unexpected opportunity for therapeutic intervention. Although APC-deficient cells show the expected increases in protein synthesis, our study reveals that it is translation elongation, and not initiation, which is the rate-limiting component. Mechanistically, mTORC1-mediated inhibition of eEF2 kinase is required for the proliferation of APC-deficient cells. Importantly, treatment of established APC-deficient adenomas with rapamycin (which can target eEF2 through the mTORC1–S6K–eEF2K axis) causes tumour cells to undergo growth arrest and differentiation. Taken together, our data suggest that inhibition of translation elongation using existing, clinically approved drugs, such as the rapalogs, would provide clear therapeutic benefit for patients at high risk of developing colorectal cancer.

Vascular integrity helps maintain brain microenvironment homeostasis, which is critical for the normal development and function of the central nervous system. It is known that neural cells can regulate brain vascular integrity. However, due to the high complexity of neurovascular interactions involved, understanding of the neural regulation of brain vascular integrity is still rudimentary. Using intact zebrafish larvae and cultured rodent brain cells, we find that neurons transfer miR-132 , a highly conserved and neuron-enriched microRNA, via secreting exosomes to endothelial cells (ECs) to maintain brain vascular integrity. Following translocation to ECs through exosome internalization, miR-132 regulates the expression of vascular endothelial cadherin (VE-cadherin), an important adherens junction protein, by directly targeting eukaryotic elongation factor 2 kinase ( eef2k ). Disruption of neuronal miR-132 expression or exosome secretion, or overexpression of vascular eef2k impairs VE-cadherin expression and brain vascular integrity. Our study indicates that miR-132 acts as an intercellular signal mediating neural regulation of the brain vascular integrity and suggests that the neuronal exosome is a novel avenue for neurovascular communication.

Nutrition is an important co-factor in exercise-induced training adaptations in muscle. We compared the effect of 6 weeks endurance training (3 days/week, 1–2 h at 75% VO 2peak ) in either the fasted state (F; n  = 10) or in the high carbohydrate state (CHO, n  = 10), on Ca 2+ -dependent intramyocellular signalling in young male volunteers. Subjects in CHO received a carbohydrate-rich breakfast before each training session, as well as ingested carbohydrates during exercise. Before ( pretest ) and after ( posttest ) the training period, subjects performed a 2 h constant-load exercise bout (~70% of pretest VO 2peak ) while ingesting carbohydrates (1 g/kg h −1 ). A muscle biopsy was taken from m. vastus lateralis immediately before and after the test, and after 4 h of recovery. Compared with pretest , in the posttest basal eukaryotic elongation factor 2 (eEF2) phosphorylation was elevated in CHO ( P  < 0.05), but not in F. In the pretest , exercise increased the degree of eEF2 phosphorylation about twofold ( P  < 0.05), and values returned to baseline within the 4 h recovery period in each group. However, in the posttest dephosphorylation of eEF2 was negated after recovery in CHO, but not in F. Independent of the dietary condition training enhanced the basal phosphorylation status of Phospholamban at Thr 17 , 5′-AMP-activated protein kinase α (AMPKα), and Acetyl CoA carboxylase β (ACCβ), and abolished the exercise-induced increase of AMPKα and ACCβ ( P  < 0.05). In conclusion, training in the fasted state, compared with identical training with ample carbohydrate intake, facilitates post-exercise dephosphorylation of eEF2. This may contribute to rapid re-activation of muscle protein translation following endurance exercise.

Molecular signaling mechanisms underlying Alzheimer's disease (AD) remain unclear. Maintenance of memory and synaptic plasticity depend on de novo protein synthesis, dysregulation of which is implicated in AD. Recent studies showed AD-associated hyperphosphorylation of mRNA translation factor eukaryotic elongation factor 2 (eEF2), which results in inhibition of protein synthesis. We tested to determine whether suppression of eEF2 phosphorylation could improve protein synthesis capacity and AD-associated cognitive and synaptic impairments. Genetic reduction of the eEF2 kinase (eEF2K) in 2 AD mouse models suppressed AD-associated eEF2 hyperphosphorylation and improved memory deficits and hippocampal long-term potentiation (LTP) impairments without altering brain amyloid β (Aβ) pathology. Furthermore, eEF2K reduction alleviated AD-associated defects in dendritic spine morphology, postsynaptic density formation, de novo protein synthesis, and dendritic polyribosome assembly. Our results link eEF2K/eEF2 signaling dysregulation to AD pathophysiology and therefore offer a feasible therapeutic target.

Gene expression is rapidly remodeled by infection and inflammation in part via transcription factor NF-κB activation and regulated protein synthesis. While protein synthesis is largely controlled by mRNA translation initiation, whether cellular translation elongation factors are responsive to inflammation and infection remains poorly understood. Here, we reveal a surprising mechanism whereby NF-κB restricts phosphorylation of the critical translation elongation factor eEF2, which catalyzes the protein synthesis translocation step. Upon exposure to NF-κB–activating stimuli, including TNFα, human cytomegalovirus infection, or double-stranded DNA, eEF2 phosphorylation on Thr56, which slows elongation to limit protein synthesis, and the overall abundance of eEF2 kinase (eEF2K) are reduced. Significantly, this reflected a p65 NF-κB subunit-dependent reduction in eEF2K pre-mRNA, indicating that NF-κB activation represses eEF2K transcription to decrease eEF2K protein levels. Finally, we demonstrate that reducing eEF2K abundance regulates protein synthesis in response to a bacterial toxin that inactivates eEF2. This establishes that NF-κB activation by diverse physiological effectors controls eEF2 activity via a transcriptional repression mechanism that reduces eEF2K polypeptide abundance to preclude eEF2 phosphorylation, thereby stimulating translation elongation and protein synthesis. Moreover, it illustrates how nuclear transcription regulation shapes translation elongation factor activity and exposes how eEF2 is integrated into innate immune response networks orchestrated by NF-κB.

INTRODUCTION Cognitive impairment is a core feature of Down syndrome (DS), and the underlying neurobiological mechanisms remain unclear. Translation dysregulation is linked to multiple neurological disorders characterized by cognitive impairments. Phosphorylation of the translational factor eukaryotic elongation factor 2 (eEF2) by its kinase eEF2K results in inhibition of general protein synthesis. METHODS We used genetic and pharmacological methods to suppress eEF2K in two lines of DS mouse models. We further applied multiple approaches to evaluate the effects of eEF2K inhibition on DS pathophysiology. RESULTS We found that eEF2K signaling was overactive in the brain of patients with DS and DS mouse models. Inhibition of eEF2 phosphorylation through suppression of eEF2K in DS model mice improved multiple aspects of DS‐associated pathophysiology including de novo protein synthesis deficiency, synaptic morphological defects, long‐term synaptic plasticity failure, and cognitive impairments. DISCUSSION Our data suggested that eEF2K signaling dysregulation mediates DS‐associated synaptic and cognitive impairments. Highlights Phosphorylation of the translational factor eukaryotic elongation factor 2 (eEF2) is increased in the Down syndrome (DS) brain. Suppression of the eEF2 kinase (eEF2K) alleviates cognitive deficits in DS models. Suppression of eEF2K improves synaptic dysregulation in DS models. Cognitive and synaptic impairments in DS models are rescued by eEF2K inhibitors.

Processing bodies (p-bodies) are a prototypical phase-separated RNA-containing granule. Their abundance is highly dynamic and has been linked to translation. Yet, the molecular mechanisms responsible for coordinate control of the two processes are unclear. Here, we uncover key roles for eEF2 kinase (eEF2K) in the control of ribosome availability and p-body abundance. eEF2K acts on a sole known substrate, eEF2, to inhibit translation. We find that the eEF2K agonist nelfinavir abolishes p-bodies in sensory neurons and impairs translation. To probe the latter, we used cryo-electron microscopy. Nelfinavir stabilizes vacant 80S ribosomes. They contain SERBP1 in place of mRNA and eEF2 in the acceptor site. Phosphorylated eEF2 associates with inactive ribosomes that resist splitting in vitro. Collectively, the data suggest that eEF2K defines a population of inactive ribosomes resistant to recycling and protected from degradation. Thus, eEF2K activity is central to both p-body abundance and ribosome availability in sensory neurons. Processing bodies are phase separated compartments enriched in translationally repressed mRNAs. Here, Smith et al. show that, in sensory neurons, eukaryotic elongation factor 2 kinase (eEF2K) plays key roles in the regulation of processing body abundance and the formation of translationally inactive ribosomes.

Route to fast antidepressants? Antidepressants such as selective serotonin re-uptake inhibitors can take months to take effect, but small doses of ketamine, a glutamatergic N-methyl-D-aspartate receptor (NMDAR) agonist, can have antidepressant effects within hours. The antidepressant mechanism of ketamine is not well understood. Work in mice shows that antidepressant-like effects of ketamine depend on rapid synthesis of brain-derived neurotrophic factor (BDNF). Ketamine-mediated NMDAR blockade deactivates eukaryotic elongation factor 2 (eEF2) kinase, resulting in reduced eEF2 phosphorylation and de-suppression of BDNF translation. These findings raise the possibility of this pathway as a therapeutic target for fast-acting antidepressants. Clinical studies consistently demonstrate that a single sub-psychomimetic dose of ketamine, an ionotropic glutamatergic NMDAR ( N -methyl- D -aspartate receptor) antagonist, produces fast-acting antidepressant responses in patients suffering from major depressive disorder, although the underlying mechanism is unclear 1 , 2 , 3 . Depressed patients report the alleviation of major depressive disorder symptoms within two hours of a single, low-dose intravenous infusion of ketamine, with effects lasting up to two weeks 1 , 2 , 3 , unlike traditional antidepressants (serotonin re-uptake inhibitors), which take weeks to reach efficacy. This delay is a major drawback to current therapies for major depressive disorder and faster-acting antidepressants are needed, particularly for suicide-risk patients 3 . The ability of ketamine to produce rapidly acting, long-lasting antidepressant responses in depressed patients provides a unique opportunity to investigate underlying cellular mechanisms. Here we show that ketamine and other NMDAR antagonists produce fast-acting behavioural antidepressant-like effects in mouse models, and that these effects depend on the rapid synthesis of brain-derived neurotrophic factor. We find that the ketamine-mediated blockade of NMDAR at rest deactivates eukaryotic elongation factor 2 (eEF2) kinase (also called CaMKIII), resulting in reduced eEF2 phosphorylation and de-suppression of translation of brain-derived neurotrophic factor. Furthermore, we find that inhibitors of eEF2 kinase induce fast-acting behavioural antidepressant-like effects. Our findings indicate that the regulation of protein synthesis by spontaneous neurotransmission may serve as a viable therapeutic target for the development of fast-acting antidepressants.

Eukaryotic elongation factor-2 kinase (eEF-2K), a negative regulator of protein synthesis, has been shown to play an important role in modulating autophagy and apoptosis in tumor cells under various stresses. In this study, we investigated the regulatory role of eEF-2K in pyroptosis (a new form of programmed necrosis) in doxorubicin-treated human melanoma cells. We found that doxorubicin (0.5–5 μmol/L) induced pyroptosis in melanoma cell lines SK-MEL-5, SK-MEL-28, and A-375 with high expression of DFNA5, but not in human breast cancer cell line MCF-7 with little expression of DFNA5. On the other hand, doxorubicin treatment activated autophagy in the melanoma cells; inhibition of autophagy by transfecting the cells with siRNA targeting Beclin1 or by pretreatment with chloroquine (20 μmol/L) significantly augmented pyroptosis, thus sensitizing the melanoma cells to doxorubicin. We further demonstrated that doxorubicin treatment activated eEF-2K in the melanoma cells, and silencing of eEF-2K blunted autophagic responses, but promoted doxorubicin-induced pyroptotic cell death. Taken together, the above results demonstrate that eEF-2K dictates the cross-talk between pyroptosis and autophagy in doxorubicin-treated human melanoma cells; suppression of eEF-2K results in inhibiting autophagy and augmenting pyroptosis, thus modulating the sensitivity of melanoma cells to doxorubicin, suggesting that targeting eEF-2K may reinforce the antitumor efficacy of doxorubicin, offering a new insight into tumor chemotherapy.

Cells and organisms frequently experience starvation. To survive, they mount an evolutionarily conserved stress response. A vital component in the mammalian starvation response is eukaryotic elongation factor 2 (eEF2) kinase (eEF2K), which suppresses translation in starvation by phosphorylating and inactivating the translation elongation driver eEF2. C. elegans EFK-1/eEF2K phosphorylates EEF-2/eEF2 on a conserved residue and is required for starvation survival, but how it promotes survival remains unclear. Surprisingly, we found that eEF2 phosphorylation is unchanged in starved C. elegans and EFK-1’s kinase activity is dispensable for starvation survival, suggesting that efk-1 promotes survival via a noncanonical pathway. We show that efk-1 upregulates transcription of DNA repair pathways, nucleotide excision repair (NER) and base excision repair (BER), to promote starvation survival. Furthermore, efk-1 suppresses oxygen consumption and ROS production in starvation to prevent oxidative stress. Thus, efk-1 enables starvation survival by protecting animals from starvation-induced oxidative damage through an EEF-2-independent pathway. Cells survive starvation by activating protective pathways. Here, the authors describe that kinase EFK-1/eEF2K defends against oxidative damage in long term starvation in C. elegans independently from its kinase activity and translation regulation.