Phosphorylation of elongation factor-2 kinase on serine 499 by cAMP-dependent protein kinase induces Ca2+/calmodulin-independent activity

Diggle, Tricia A.; Subkhankulova, Tatiana; Lilley, Kathryn S.; Shikotra, Nita; Willis, Anne E.; Redpath, Nicholas T.

doi:10.1042/0264-6021:3530621

Cited by 52 publications

(60 citation statements)

References 18 publications

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“…From the literature, 4E-BP1 activation seems to be complex and the effect of insulin could involve multiple phosphorylation events, which have been described as rapamycin sensitive and insensitive. 20,31,41 We show that a rapamycin-sensitive pathway is involved in 4E-BP1 phosphorylation in response to insulin. This regulation could account for the permissive role of insulin on hepatocyte proliferation.…”

Section: Discussionmentioning

confidence: 79%

PI3K-FRAP/mTOR pathway is critical for hepatocyte proliferation whereas MEK/ERK supports both proliferation and survival

Coutant¹,

Rescan²,

Gilot³

et al. 2002

Hepatology

119

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Growth factors are known to favor both proliferation and survival of hepatocytes. In this work, we investigated the role of 2 main signaling pathways, phosphoinositide 3-kinase (PI3K) and mitogen-activated protein kinase (MEK)/extracellular signal-regulated kinase (ERK), in these processes. First, evidence was provided that the PI3K cascade as well as the MEK/ERK cascade is a key transduction pathway controlling hepatocyte proliferation, as ascertained by arrest of DNA synthesis in the presence of LY294002, a specific PI3K inhibitor. Inhibition of FRAP/mTOR by rapamycin also abrogated DNA replication and protein synthesis induced by growth factor. We showed that expression of cyclin D1 at messenger RNA (mRNA) and protein levels was regulated by this pathway. We highlighted that 4E-BP1 phosphorylation was not activated by epidermal growth factor (EGF) but was under an insulin-regulation mechanism through a PI3K-FRAP/mTOR activation that could account for the permissive role of insulin on hepatocyte proliferation. No interference between the MEK/ERK pathway and 4E-BP1 phosphorylation was detected, whereas p70S6K phosphorylation induced by EGF was under a U0126-sensitive regulation. Last, we established that the antiapoptotic function of EGF was dependent on MEK, whereas LY294002 and rapamycin had no direct effect on cell survival. Taken together, these data highlight the regulation and the role of 2 pathways that mediate growth-related response by acting onto distinct steps. In conclusion, hepatocyte progression in late G1 phase induced by EGF generates survival signals depending on MEK activation, whereas PI3K and MEK/ERK cascades are both necessary for hepatocyte replication. (HEPATOLOGY 2002;36:1079-1088 C ell cycle progression through G1 phase requires the integration of signals from the extracellular environment such as growth factors, cytokines, and extracellular matrix proteins mediated by different transduction cascades. The ability to induce cellular proliferation is often correlated with the ability to promote cell survival. Two signaling cascades have emerged as major players in the mitogenic and antiapoptotic response in many cells: the mitogen-activated protein kinase (MEK)/ extracellular signal-regulated kinase (ERK) and the phosphoinositide 3-kinase (PI3K) pathways.Growth factors as survival factors bind to cell surface receptors and trigger the activation of several kinases, including PI3K, which seems to be implicated in the control of growth and apoptosis of a wide range of cell types. 1 This kinase activates PDK1 that in turn leads to the activation of a serine/threonine kinase termed AKT or PKB, which plays a central role in promoting the survival of many cell types. PDK1 also phosphorylates and activates FRAP/mTOR (target of rapamycin). p70s6k, which regulates the ribosomal S6 subunit phosphorylation in response to mitogens, is controlled by FRAP/mTOR and plays an essential role in the translation machinery. 2-4 FRAP/mTOR also controls the phosphorylation of 4E-BP1, a key protein involved in t...

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Section: Discussionmentioning

confidence: 79%

PI3K-FRAP/mTOR pathway is critical for hepatocyte proliferation whereas MEK/ERK supports both proliferation and survival

Coutant¹,

Rescan²,

Gilot³

et al. 2002

Hepatology

119

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“…Furthermore, pharmacological inducers of ER stress such as Tm and thapsigargin (Tg), an inhibitor of the ER-resident Ca 2 þ -dependent ATPase, induced eEF-2 phosphorylation with kinetics similar to those of sal (Supplementary Figure 3). EEF-2 phosphorylation has been observed in response to several stimuli, [26][27][28][29][30][31][32][33] but never in the context of ER stress. Our observations suggest that eEF-2 might be regulated by phosphorylation during ESR.…”

Section: Resultsmentioning

confidence: 99%

“…[22][23][24] The phosphorylation of eEF-2 is catalyzed by eEF-2 kinase (eEF-2K), an unusual calcium/calmodulin-dependent enzyme belonging to the alpha kinase family of atypical protein kinases. 25 EEF-2 phosphorylation has been shown to play an important role in coupling protein synthesis to energy metabolism in response to calcium flux, 26 cyclic adenosine monophosphate (AMP)/protein kinase A and AMP-activated protein kinase signaling, [27][28][29][30] amino acid or glucose limitation, 31,32 and cytoplasmic pH changes, 33 but no role for eEF-2 phosphorylation in ER stress has been reported.…”

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confidence: 99%

A pharmacoproteomic approach implicates eukaryotic elongation factor 2 kinase in ER stress-induced cell death

Boyce

Ryazanov

et al. 2008

Cell Death Differ

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Apoptosis triggered by endoplasmic reticulum (ER) stress has been implicated in many diseases but its cellular regulation remains poorly understood. Previously, we identified salubrinal (sal), a small molecule that protects cells from ER stressinduced apoptosis by selectively activating a subset of endogenous ER stress-signaling events. Here, we use sal as a probe in a proteomic approach to discover new information about the endogenous cellular response to ER stress. We show that sal induces phosphorylation of the translation elongation factor eukaryotic translation elongation factor 2 (eEF-2), an event that depends on eEF-2 kinase (eEF-2K). ER stress itself also induces eEF-2K-dependent eEF-2 phosphorylation, and this pathway promotes translational arrest and cell death in this context, identifying eEF-2K as a hitherto unknown regulator of ER stress-induced apoptosis. Finally, we use both sal and ER stress models to show that eEF-2 phosphorylation can be activated by at least two signaling mechanisms. Our work identifies eEF-2K as a new component of the ER stress response and underlines the utility of novel small molecules in discovering new cell biology. Cell Death and Differentiation (2008) 15, 589-599; doi:10.1038/sj.cdd.4402296; published online 11 January 2008 The endoplasmic reticulum (ER) serves as the primary processing site for membrane and secreted proteins. The ER recruits translating ribosomes, translocates newly synthesized polypeptides into its lumen, and promotes a variety of post-translational modifications and chaperone-facilitated protein folding. 1,2 Proper ER function is critical for numerous aspects of cell physiology, including vesicle trafficking, lipid and membrane biogenesis, and protein targeting and secretion. Accordingly, cells react rapidly to various forms of ER dysfunction -including the accumulation of unfolded, misfolded or excessive protein, ER lipid or glycolipid imbalances, or changes in the redox or ionic conditions of the ER lumenthrough a set of adaptive pathways known collectively as the ER stress response (ESR). [2][3][4][5] The ESR promotes cell survival both by increasing the capacity of the ER to fold and process client proteins and by reducing the amount of protein inside the ER. These effects are achieved through three major pathways: (1) the unfolded protein response, a transcription-dependent induction of ER lumenal chaperone proteins and many other components of the secretory apparatus, which augments the polypeptide processing capacity of the ER; 5,6 (2) the activation of proteasome-dependent ER-associated degradation to remove proteins from the ER; 7,8 and (3) the control of protein translation to modulate the polypeptide traffic into the ER. 9,10 Normally, this suite of responses succeeds in restoring ER homeostasis. However, in metazoans, persistent or intense ER stress can also trigger apoptosis. [11][12][13][14] ER stress and the apoptotic program coupled to it have been implicated in many important pathologies, including diabetes, obesity, neurodegenerativ...

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“…1), which plays a role in the ubiquitin-mediated degradation of eEF-2K (29,30). Ser-500, a major autophosphorylation site (31) and also a substrate for PKA (32), lies at the N terminus of a putative helix. The C-terminal domain is predicted to be highly helical, containing three proposed SEL1-like domains (starting at residues 576, 610, and 665) that are thought to be important for protein-protein interactions (25).…”

Section: Eef-2kmentioning

confidence: 99%

“…The C terminus is essential for eEF-2 binding to eEF-2K (25). Several phosphorylation sites associated with eEF-2K regulation are indicated: Ser-78 (which is phosphorylated by an unknown mTOR-regulated kinase (25)), Thr-348 (an autophosphorylation site (31, 57)), Ser-359 (phosphorylated by p38␦ (58) and the Cdc2-cyclin B complex (53)), Ser-366 (phosphorylated by p70 S6 kinase (59) and p90 RSK1 (59)), Ser-398 (phosphorylated by AMPK (55)), Ser-441 (phosphorylated by an unknown kinase (30,60)), Ser-445 (an autophosphorylation site (30,31,57)), and Ser-500 (phosphorylated by PKA (32) and an autophosphorylation site (31)). Binding of phospho-Thr-348 to ABP facilitates full activation of eEF-2K.…”

Section: Reagents Plasmids Strains and Equipmentmentioning

confidence: 99%

The Molecular Mechanism of Eukaryotic Elongation Factor 2 Kinase Activation

Tavares¹,

Ferguson

Giles

et al. 2014

Journal of Biological Chemistry

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Background: Eukaryotic elongation factor 2 kinase (eEF-2K) regulates protein translation elongation rates. Results: eEF-2K activation involves a two-step process of calmodulin binding and rapid Thr-348 autophosphorylation. Conclusion: Activation of eEF-2K involves two distinct allosteric steps, both of which potentially induce a conformational change. Significance: This mechanism provides a framework for understanding how eEF-2K integrates inputs from multiple upstream signaling pathways.

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Phosphorylation of elongation factor-2 kinase on serine 499 by cAMP-dependent protein kinase induces Ca2+/calmodulin-independent activity

Cited by 52 publications

References 18 publications

PI3K-FRAP/mTOR pathway is critical for hepatocyte proliferation whereas MEK/ERK supports both proliferation and survival

PI3K-FRAP/mTOR pathway is critical for hepatocyte proliferation whereas MEK/ERK supports both proliferation and survival

A pharmacoproteomic approach implicates eukaryotic elongation factor 2 kinase in ER stress-induced cell death

The Molecular Mechanism of Eukaryotic Elongation Factor 2 Kinase Activation

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