scholarly journals eIF2B as a Target for Viral Evasion of PKR-Mediated Translation Inhibition

mBio ◽  
2020 ◽  
Vol 11 (4) ◽  
Author(s):  
Jennifer Deborah Wuerth ◽  
Matthias Habjan ◽  
Markus Kainulainen ◽  
Besim Berisha ◽  
Damien Bertheloot ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Protein Kinase ◽  
Nonstructural Protein ◽  
Mrna Translation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Mammalian Host ◽  
Eukaryotic Initiation Factor ◽  
Α Subunit ◽  
Immune Factor

ABSTRACT RNA-activated protein kinase (PKR) is a major innate immune factor that senses viral double-stranded RNA (dsRNA) and phosphorylates eukaryotic initiation factor (eIF) 2α. Phosphorylation of the α subunit converts the eIF2αβγ complex into a stoichiometric inhibitor of eukaryotic initiation factor eIF2B, thus halting mRNA translation. To escape this protein synthesis shutoff, viruses have evolved countermechanisms such as dsRNA sequestration, eIF-independent translation by an internal ribosome binding site, degradation of PKR, or dephosphorylation of PKR or of phospho-eIF2α. Here, we report that sandfly fever Sicilian phlebovirus (SFSV) confers such a resistance without interfering with PKR activation or eIF2α phosphorylation. Rather, SFSV expresses a nonstructural protein termed NSs that strongly binds to eIF2B. Although NSs still allows phospho-eIF2α binding to eIF2B, protein synthesis and virus replication are unhindered. Hence, SFSV encodes a unique PKR antagonist that acts by rendering eIF2B resistant to the inhibitory action of bound phospho-eIF2α. IMPORTANCE RNA-activated protein kinase (PKR) is one of the most powerful antiviral defense factors of the mammalian host. PKR acts by phosphorylating mRNA translation initiation factor eIF2α, thereby converting it from a cofactor to an inhibitor of mRNA translation that strongly binds to initiation factor eIF2B. To sustain synthesis of their proteins, viruses are known to counteract this on the level of PKR or eIF2α or by circumventing initiation factor-dependent translation altogether. Here, we report a different PKR escape strategy executed by sandfly fever Sicilian virus (SFSV), a member of the increasingly important group of phleboviruses. We found that the nonstructural protein NSs of SFSV binds to eIF2B and protects it from inactivation by PKR-generated phospho-eIF2α. Protein synthesis is hence maintained and the virus can replicate despite ongoing full-fledged PKR signaling in the infected cells. Thus, SFSV has evolved a unique strategy to escape the powerful antiviral PKR.

scholarly journals Regulation of mRNA Translation and Cellular Signaling by Hepatitis C Virus Nonstructural Protein NS5A

2001 ◽  
Vol 75 (11) ◽  
pp. 5090-5098 ◽  
Author(s):  
Yupeng He ◽  
Seng-Lai Tan ◽  
Semih U. Tareen ◽  
Sangeetha Vijaysri ◽  
Jeffrey O. Langland ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Protein Kinase ◽  
Virus Infection ◽  
Nonstructural Protein ◽  
Mrna Translation ◽  
Mitogen Activated Protein Kinase ◽  
Initiation Factor ◽  
Translation Rate ◽  
Α Subunit

ABSTRACT The NS5A nonstructural protein of hepatitis C virus (HCV) has been shown to inhibit the cellular interferon (IFN)-induced protein kinase R (PKR). PKR mediates the host IFN-induced antiviral response at least in part by inhibiting mRNA translation initiation through phosphorylation of the α subunit of eukaryotic initiation factor 2 (eIF2α). We thus examined the effect of NS5A inhibition of PKR on mRNA translation within the context of virus infection by using a recombinant vaccinia virus (VV)-based assay. The VV E3L protein is a potent inhibitor of PKR. Accordingly, infection of IFN-pretreated HeLa S3 cells with an E3L-deficient VV (VVΔE3L) resulted in increased phosphorylation levels of both PKR and eIF2α. IFN-pretreated cells infected with VV in which the E3L locus was replaced with theNS5A gene (VVNS5A) displayed diminished phosphorylation of PKR and eIF2α in a transient manner. We also observed an increase in activation of p38 mitogen-activated protein kinase in IFN-pretreated cells infected with VVΔE3L, consistent with reports that p38 lies downstream of the PKR pathway. Furthermore, these cells exhibited increased phosphorylation of the cap-binding initiation factor 4E (eIF4E), which is downstream of the p38 pathway. Importantly, these effects were reduced in cells infected with VVNS5A. NS5A was also found to inhibit activation of the p38-eIF4E pathway in epidermal growth factor-treated cells stably expressing NS5A. NS5A-induced inhibition of eIF2α and eIF4E phosphorylation may exert counteracting effects on mRNA translation. Indeed, IFN-pretreated cells infected with VVNS5A exhibited a partial and transient restoration of cellular and viral mRNA translation compared with IFN-pretreated cells infected with VVΔE3L. Taken together, these results support the role of NS5A as a PKR inhibitor and suggest a potential mechanism by which HCV might maintain global mRNA translation rate during early virus infection while favoring cap-independent translation of HCV mRNA during late infection.

scholarly journals Defects in Translational Regulation Mediated by the α Subunit of Eukaryotic Initiation Factor 2 Inhibit Antiviral Activity and Facilitate the Malignant Transformation of Human Fibroblasts

2004 ◽  
Vol 24 (5) ◽  
pp. 2025-2040 ◽  
Author(s):  
Darren J. Perkins ◽  
Glen N. Barber
Keyword(s):  
Protein Synthesis ◽  
Translation Initiation ◽  
T Antigen ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Eukaryotic Initiation Factor ◽  
Α Subunit ◽  
Eukaryotic Initiation Factor 2 ◽  
Initiation Factor 2

ABSTRACT Suppression of protein synthesis through phosphorylation of the translation initiation factor α subunit of eukaryotic initiation factor 2 (eIF2α) is known to occur in response to many forms of cellular stress. To further study this, we have developed novel cell lines that inducibly express FLAG-tagged versions of either the phosphomimetic eIF2α variant, eIF2α-S51D, or the phosphorylation-insensitive eIF2α-S51A. These variants showed authentic subcellular localization, were incorporated into endogenous ternary complexes, and were able to modulate overall rates of protein synthesis as well as influence cell division. However, phosphorylation of eIF2α failed to induce cell death or sensitize cells to killing by proapoptotic stimuli, though it was able to inhibit viral replication, confirming the role of eIF2α in host defense. Further, although the eIF2α-S51A variant has been shown to transform NIH 3T3 cells, it was unable to transform the murine fibroblast 3T3 L1 cell line. To therefore clarify this issue, we explored the role of eIF2α in growth control and demonstrated that the eIF2α-S51A variant is capable of collaborating with hTERT and the simian virus 40 large T antigen in the transformation of primary human kidney cells. Thus, dysregulation of translation initiation is indeed sufficient to cooperate with defined oncogenic elements and participate in the tumorigenesis of human tissue.

A critical review of translation initiation factor eIF2α kinases in plants - regulating protein synthesis during stress

2012 ◽  
Vol 39 (9) ◽  
pp. 717 ◽  
Author(s):  
Tracey M. Immanuel ◽  
David R. Greenwood ◽  
Robin M. MacDiarmid
Keyword(s):  
Protein Synthesis ◽  
Translation Initiation ◽  
Current Knowledge ◽  
Mrna Translation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Α Subunit ◽  
Eif2α Phosphorylation ◽  
Phosphorylation Mechanism ◽  
Eif2α Kinase

Eukaryotic cells must cope with environmental stress. One type of general stress response is the downregulation of protein synthesis in order to conserve cellular resources. Protein synthesis is mainly regulated at the level of mRNA translation initiation and when the α subunit of eukaryotic translation initiation factor 2 (eIF2) is phosphorylated, protein synthesis is downregulated. Although eIF2 has the same translation initiation function in all eukaryotes, it is not known whether plants downregulate protein synthesis via eIF2α phosphorylation. Similarly, although there is evidence that plants possess eIF2α kinases, it is not known whether they operate in a similar manner to the well characterised mammalian and yeast eIF2α kinases. Two types of eIF2α kinases have been reported in plants, yet the full understanding of the plant eIF2α phosphorylation mechanism is still lacking. Here we review the current knowledge of the eIF2α phosphorylation mechanism within plants and discuss plant eIF2α, plant eIF2α kinase GCN2 and the data supporting and contradicting the hypothesis that a functional orthologue for the eIF2α kinase PKR, is present and functional in plants.

scholarly journals Correction of eIF2-dependent defects in brain protein synthesis, synaptic plasticity, and memory in mouse models of Alzheimer’s disease

2021 ◽  
Vol 14 (668) ◽  
pp. eabc5429
Author(s):  
Mauricio M. Oliveira ◽  
Mychael V. Lourenco ◽  
Francesco Longo ◽  
Nicole P. Kasica ◽  
Wenzhong Yang ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Protein Synthesis ◽  
Synaptic Plasticity ◽  
Translation Initiation ◽  
Mrna Translation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Postmortem Brain Tissue ◽  
Phosphorylated Eif2α

Neuronal protein synthesis is essential for long-term memory consolidation, and its dysregulation is implicated in various neurodegenerative disorders, including Alzheimer’s disease (AD). Cellular stress triggers the activation of protein kinases that converge on the phosphorylation of eukaryotic translation initiation factor 2α (eIF2α), which attenuates mRNA translation. This translational inhibition is one aspect of the integrated stress response (ISR). We found that postmortem brain tissue from AD patients showed increased phosphorylation of eIF2α and reduced abundance of eIF2B, another key component of the translation initiation complex. Systemic administration of the small-molecule compound ISRIB (which blocks the ISR downstream of phosphorylated eIF2α) rescued protein synthesis in the hippocampus, measures of synaptic plasticity, and performance on memory-associated behavior tests in wild-type mice cotreated with salubrinal (which inhibits translation by inducing eIF2α phosphorylation) and in both β-amyloid-treated and transgenic AD model mice. Thus, attenuating the ISR downstream of phosphorylated eIF2α may restore hippocampal protein synthesis and delay cognitive decline in AD patients.

scholarly journals Caspase-mediated cleavage of eukaryotic translation initiation factor subunit 2α

1999 ◽  
Vol 342 (1) ◽  
pp. 65-70 ◽  
Author(s):  
Shinya SATOH ◽  
Makoto HIJIKATA ◽  
Hiroshi HANDA ◽  
Kunitada SHIMOTOHNO
Keyword(s):  
Protein Synthesis ◽  
Translation Initiation ◽  
Caspase 3 ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Eukaryotic Translation Initiation Factor ◽  
Α Subunit ◽  
Eukaryotic Translation ◽  
Initiation Of Translation ◽  
Eukaryotic Translation Initiation

Eukaryotic translation initiation factor 2α (eIF-2α), a target molecule of the interferon-inducible double-stranded-RNA-dependent protein kinase (PKR), was cleaved in apoptotic Saos-2 cells on treatment with poly(I)˙poly(C) or tumour necrosis factor α. This cleavage occurred with a time course similar to that of poly(ADP-ribose) polymerase, a well-known caspase substrate. In addition, eIF-2α was cleaved by recombinant active caspase-3 in vitro. By site-directed mutagenesis, the cleavage site was mapped to an Ala-Glu-Val-Asp300 ↓ Gly301 sequence located in the C-terminal portion of eIF-2α. PKR phosphorylates eIF-2α on Ser51, resulting in the suppression of protein synthesis. PKR-mediated translational suppression was repressed when the C-terminally cleaved product of eIF-2α was overexpressed in Saos-2 cells, even though PKR can phosphorylate this cleaved product. These results suggest that caspase-3 or related protease(s) can modulate the efficiency of protein synthesis by cleaving the α subunit of eIF-2, a key component in the initiation of translation.

scholarly journals Severe Acute Respiratory Syndrome Coronavirus Triggers Apoptosis via Protein Kinase R but Is Resistant to Its Antiviral Activity

2008 ◽  
Vol 83 (5) ◽  
pp. 2298-2309 ◽  
Author(s):  
Verena Krähling ◽  
David A. Stein ◽  
Martin Spiegel ◽  
Friedemann Weber ◽  
Elke Mühlberger
Keyword(s):  
Protein Kinase ◽  
Severe Acute Respiratory Syndrome ◽  
Chromatin Condensation ◽  
Mrna Translation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Protein Kinase R ◽  
Beta Interferon ◽  
Reverse Transcription Pcr ◽  
Eif2α Phosphorylation

ABSTRACT In this study, infection of 293/ACE2 cells with severe acute respiratory syndrome coronavirus (SARS-CoV) activated several apoptosis-associated events, namely, cleavage of caspase-3, caspase-8, and poly(ADP-ribose) polymerase 1 (PARP), and chromatin condensation and the phosphorylation and hence inactivation of the eukaryotic translation initiation factor 2α (eIF2α). In addition, two of the three cellular eIF2α kinases known to be virus induced, protein kinase R (PKR) and PKR-like endoplasmic reticulum kinase (PERK), were activated by SARS-CoV. The third kinase, general control nonderepressible-2 kinase (GCN2), was not activated, but late in infection the level of GCN2 protein was significantly reduced. Reverse transcription-PCR analyses revealed that the reduction of GCN2 protein was not due to decreased transcription or stability of GCN2 mRNA. The specific reduction of PKR protein expression by antisense peptide-conjugated phosphorodiamidate morpholino oligomers strongly reduced cleavage of PARP in infected cells. Surprisingly, the knockdown of PKR neither enhanced SARS-CoV replication nor abrogated SARS-CoV-induced eIF2α phosphorylation. Pretreatment of cells with beta interferon prior to SARS-CoV infection led to a significant decrease in PERK activation, eIF2α phosphorylation, and SARS-CoV replication. The various effects of beta interferon treatment were found to function independently on the expression of PKR. Our results show that SARS-CoV infection activates PKR and PERK, leading to sustained eIF2α phosphorylation. However, virus replication was not impaired by these events, suggesting that SARS-CoV possesses a mechanism to overcome the inhibitory effects of phosphorylated eIF2α on viral mRNA translation. Furthermore, our data suggest that viral activation of PKR can lead to apoptosis via a pathway that is independent of eIF2α phosphorylation.

scholarly journals Activation of mRNA translation in rat cardiac myocytes by insulin involves multiple rapamycin-sensitive steps

2000 ◽  
Vol 278 (4) ◽  
pp. H1056-H1068 ◽  
Author(s):  
Lijun Wang ◽  
Xuemin Wang ◽  
Christopher G. Proud
Keyword(s):  
Protein Synthesis ◽  
Protein Kinase ◽  
Signaling Pathways ◽  
Glycogen Synthase ◽  
Mrna Translation ◽  
Mitogen Activated Protein Kinase ◽  
Initiation Factor ◽  
Translation Factors ◽  
P70 S6k ◽  
Eef2 Kinase

Insulin acutely activates protein synthesis in ventricular cardiomyocytes from adult rats. In this study, we have established the methodology for studying the regulation of the signaling pathways and translation factors that may be involved in this response and have examined the effects of acute insulin treatment on them. Insulin rapidly activated the 70-kDa ribosomal S6 kinase (p70 S6k), and this effect was inhibited both by rapamycin and by inhibitors of phosphatidylinositol 3-kinase. The activation of p70 S6k is mediated by a signaling pathway involving the mammalian target of rapamycin (mTOR), which also modulates other translation factors. These include the eukaryotic initiation factor (eIF) 4E binding proteins (4E-BPs) and eukaryotic elongation factor 2 (eEF2). Insulin caused phosphorylation of 4E-BP1 and induced its dissociation from eIF4E, and these effects were also blocked by rapamycin. Concomitant with this, insulin increased the binding of eIF4E to eIF4G. Insulin also activated protein kinase B (PKB), which may lie upstream of p70 S6k and 4E-BP1, with the activation of the different isoforms being in the order α>β>γ. Insulin also caused inhibition of glycogen synthase kinase 3, which lies downstream of PKB, and of eEF2 kinase. The phosphorylation of eEF2 itself was also decreased by insulin, and this effect and the inactivation of eEF2 kinase were attenuated by rapamycin. The activation of overall protein synthesis by insulin in cardiomyocytes was substantially inhibited by rapamycin (but not by inhibitors of other specific signaling pathways, e.g., mitogen-activated protein kinase), showing that signaling events linked to mTOR play a major role in the control of translation by insulin in this cell type.

scholarly journals eIF-Three to Tango: emerging functions of translation initiation factor eIF3 in protein synthesis and disease

2020 ◽  
Vol 12 (6) ◽  
pp. 403-409 ◽  
Author(s):  
Dieter A Wolf ◽  
Yingying Lin ◽  
Haoran Duan ◽  
Yabin Cheng
Keyword(s):  
Protein Synthesis ◽  
Messenger Rna ◽  
Protein Production ◽  
Mrna Translation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Protein Homeostasis ◽  
Subcellular Targeting ◽  
Post Translational Modifications ◽  
The Past

Abstract Studies over the past three years have substantially expanded the involvements of eukaryotic initiation factor 3 (eIF3) in messenger RNA (mRNA) translation. It now appears that this multi-subunit complex is involved in every possible form of mRNA translation, controlling every step of protein synthesis from initiation to elongation, termination, and quality control in positive as well as negative fashion. Through the study of eIF3, we are beginning to appreciate protein synthesis as a highly integrated process coordinating protein production with protein folding, subcellular targeting, and degradation. At the same time, eIF3 subunits appear to have specific functions that probably vary between different tissues and individual cells. Considering the broad functions of eIF3 in protein homeostasis, it comes as little surprise that eIF3 is increasingly implicated in major human diseases and first attempts at therapeutically targeting eIF3 have been undertaken. Much remains to be learned, however, about subunit- and tissue-specific functions of eIF3 in protein synthesis and disease and their regulation by environmental conditions and post-translational modifications.

Physiological rise in plasma leucine stimulates muscle protein synthesis in neonatal pigs by enhancing translation initiation factor activation

2005 ◽  
Vol 288 (5) ◽  
pp. E914-E921 ◽  
Author(s):  
Jeffery Escobar ◽  
Jason W. Frank ◽  
Agus Suryawan ◽  
Hanh V. Nguyen ◽  
Scot R. Kimball ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Muscle Protein ◽  
Muscle Protein Synthesis ◽  
Mrna Translation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Neonatal Pigs ◽  
Plasma Leucine

Protein synthesis in skeletal muscle of adult rats increases in response to oral gavage of supraphysiological doses of leucine. However, the effect on protein synthesis of a physiological rise in plasma leucine has not been investigated in neonates, an anabolic population highly sensitive to amino acids and insulin. Therefore, in the current study, fasted pigs were infused intra-arterially with leucine (0, 200, or 400 μmol·kg−1·h−1), and protein synthesis was measured after 60 or 120 min. Protein synthesis was increased in muscle, but not in liver, at 60 min. At 120 min, however, protein synthesis returned to baseline levels in muscle but was reduced below baseline values in liver. The increase in protein synthesis in muscle was associated with increased plasma leucine of 1.5- to 3-fold and no change in plasma insulin. Leucine infusion for 120 min reduced plasma essential amino acid levels. Phosphorylation of eukaryotic initiation factor (eIF)-4E-binding protein-1 (4E-BP1), ribosomal protein (rp) S6 kinase, and rpS6 was increased, and the amount of eIF4E associated with its repressor 4E-BP1 was reduced after 60 and 120 min of leucine infusion. No change in these biomarkers of mRNA translation was observed in liver. Thus a physiological increase in plasma leucine stimulates protein synthesis in skeletal muscle of neonatal pigs in association with increased eIF4E availability for eIF4F assembly. This response appears to be insulin independent, substrate dependent, and tissue specific. The results suggest that the branched-chain amino acid leucine can act as a nutrient signal to stimulate protein synthesis in skeletal muscle of neonates.

scholarly journals Phosphorylation of Eukaryotic Initiation Factor 2 by Heme-Regulated Inhibitor Kinase-Related Protein Kinases in Schizosaccharomyces pombe Is Important for Resistance to Environmental Stresses

2002 ◽  
Vol 22 (20) ◽  
pp. 7134-7146 ◽  
Author(s):  
Ke Zhan ◽  
Krishna M. Vattem ◽  
Bettina N. Bauer ◽  
Thomas E. Dever ◽  
Jane-Jane Chen ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Schizosaccharomyces Pombe ◽  
Environmental Stresses ◽  
Initiation Factor ◽  
Eukaryotic Initiation Factor ◽  
Α Subunit ◽  
Eukaryotic Initiation Factor 2 ◽  
Initiation Factor 2 ◽  
Eif2α Kinase

ABSTRACT Protein synthesis is regulated by the phosphorylation of the α subunit of eukaryotic initiation factor 2 (eIF2α) in response to different environmental stresses. One member of the eIF2α kinase family, heme-regulated inhibitor kinase (HRI), is activated under heme-deficient conditions and blocks protein synthesis, principally globin, in mammalian erythroid cells. We identified two HRI-related kinases from Schizosaccharomyces pombe which have full-length homology with mammalian HRI. The two HRI-related kinases, named Hri1p and Hri2p, exhibit autokinase and kinase activity specific for Ser-51 of eIF2α, and both activities were inhibited in vitro by hemin, as previously described for mammalian HRI. Overexpression of Hri1p, Hri2p, or the human eIF2α kinase, double-stranded-RNA-dependent protein kinase (PKR), impeded growth of S. pombe due to elevated phosphorylation of eIF2α. Cells from strains with deletions of the hri1+ and hri2+ genes, individually or in combination, exhibited a reduced growth rate when exposed to heat shock or to arsenic compounds. Measurements of in vivo phosphorylation of eIF2α suggest that Hri1p and Hri2p differentially phosphorylate eIF2α in response to these stress conditions. These results demonstrate that HRI-related enzymes are not unique to vertebrates and suggest that these eIF2α kinases are important participants in diverse stress response pathways in some lower eukaryotes.

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