scholarly journals Translation suppression promotes stress granule formation and cell survival in response to cold shock

2012 ◽  
Vol 23 (19) ◽  
pp. 3786-3800 ◽  
Author(s):  
Sarah Hofmann ◽  
Valeria Cherkasova ◽  
Peter Bankhead ◽  
Bernd Bukau ◽  
Georg Stoecklin
Keyword(s):  
Protein Synthesis ◽  
Mammalian Cells ◽  
Cold Shock ◽  
Optimal Growth ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Dependent Manner ◽  
Granule Formation ◽  
Cellular Survival ◽  
Compound C

Cells respond to different types of stress by inhibition of protein synthesis and subsequent assembly of stress granules (SGs), cytoplasmic aggregates that contain stalled translation preinitiation complexes. Global translation is regulated through the translation initiation factor eukaryotic initiation factor 2α (eIF2α) and the mTOR pathway. Here we identify cold shock as a novel trigger of SG assembly in yeast and mammals. Whereas cold shock–induced SGs take hours to form, they dissolve within minutes when cells are returned to optimal growth temperatures. Cold shock causes eIF2α phosphorylation through the kinase PERK in mammalian cells, yet this pathway is not alone responsible for translation arrest and SG formation. In addition, cold shock leads to reduced mitochondrial function, energy depletion, concomitant activation of AMP-activated protein kinase (AMPK), and inhibition of mTOR signaling. Compound C, a pharmacological inhibitor of AMPK, prevents the formation of SGs and strongly reduces cellular survival in a translation-dependent manner. Our results demonstrate that cells actively suppress protein synthesis by parallel pathways, which induce SG formation and ensure cellular survival during hypothermia.

scholarly journals Measles Virus N Protein Inhibits Host Translation by Binding to eIF3-p40

2007 ◽  
Vol 81 (21) ◽  
pp. 11569-11576 ◽  
Author(s):  
Hiroki Sato ◽  
Munemitsu Masuda ◽  
Moeko Kanai ◽  
Kyoko Tsukiyama-Kohara ◽  
Misako Yoneda ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Mammalian Cells ◽  
Measles Virus ◽  
Encephalomyocarditis Virus ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Translation System ◽  
Dependent Manner ◽  
Initiation Factors ◽  
Human T Cell

ABSTRACT The nonsegmented, negative-sense RNA genome of measles virus (MV) is encapsidated by the virus-encoded nucleocapsid protein (N). In this study, we searched for N-binding cellular proteins by using MV-N as bait and screening the human T-cell cDNA library by yeast two-hybrid assay and isolated the p40 subunit of eukaryotic initiation factor 3 (eIF3-p40) as a binding partner. The interaction between MV-N and eIF3-p40 in mammalian cells was confirmed by coimmunoprecipitation. Since eIF3-p40 is a translation initiation factor, we analyzed the potential inhibitory effect of MV-N on protein synthesis. Glutathione S-transferase (GST)-fused MV-N (GST-N) inhibited translation of reporter mRNAs in rabbit reticulocyte lysate translation system in a dose-dependent manner. Encephalomyocarditis virus internal ribosomal entry site-mediated translation, which requires canonical initiation factors to initiate translation, was also inhibited by GST-N. In contrast, a unique form of translation mediated by the intergenic region of Plautia stali intestine virus, which can assemble 80S ribosomes in the absence of canonical initiation factors, was scarcely affected by GST-N. In vivo expression of MV-N induced by the Cre/loxP switching system inhibited the synthesis of a transfected reporter protein, as well as overall protein synthesis. These results suggest that MV-N targets eIF3-p40 and may be involved in inhibiting MV-induced host translation.

scholarly journals IRE1α-Dependent Decay of CReP/Ppp1r15b mRNA Increases Eukaryotic Initiation Factor 2α Phosphorylation and Suppresses Protein Synthesis

2015 ◽  
Vol 35 (16) ◽  
pp. 2761-2770 ◽  
Author(s):  
Jae-Seon So ◽  
Sungyun Cho ◽  
Sang-Hyun Min ◽  
Scot R. Kimball ◽  
Ann-Hwee Lee
Keyword(s):  
Protein Synthesis ◽  
Er Stress ◽  
Translational Control ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Regulatory Subunit ◽  
Dependent Manner ◽  
Eukaryotic Translation ◽  
Eif2α Phosphorylation ◽  
The Unfolded Protein Response

The unfolded protein response (UPR) regulates endoplasmic reticulum (ER) homeostasis and protects cells from ER stress. IRE1α is a central regulator of the UPR that activates the transcription factor XBP1s through an unconventional splicing mechanism using its endoribonuclease activity. IRE1α also cleaves certain mRNAs containing XBP1-like secondary structures to promote the degradation of these mRNAs, a process known as regulated IRE1α-dependent decay (RIDD). We show here that the mRNA of CReP/Ppp1r15b, a regulatory subunit of eukaryotic translation initiation factor 2α (eIF2α) phosphatase, is a RIDD substrate. eIF2α plays a central role in the integrated stress response by mediating the translational attenuation to decrease the stress level in the cell. CReP expression was markedly suppressed in XBP1-deficient mice livers due to hyperactivated IRE1α. Decreased CReP expression caused the induction of eIF2α phosphorylation and the attenuation of protein synthesis in XBP1-deficient livers. ER stress also suppressed CReP expression in an IRE1α-dependent manner, which increased eIF2α phosphorylation and consequently attenuated protein synthesis. Taken together, the results of our study reveal a novel function of IRE1α in the regulation of eIF2α phosphorylation and the translational control.

scholarly journals mTORC1 signalling and eIF4E/4E-BP1 translation initiation factor stoichiometry influence recombinant protein productivity from GS-CHOK1 cells

2016 ◽  
Vol 473 (24) ◽  
pp. 4651-4664 ◽  
Author(s):  
Lyne Jossé ◽  
Jianling Xie ◽  
Christopher G. Proud ◽  
C. Mark Smales
Keyword(s):  
Protein Synthesis ◽  
Cell Growth ◽  
Recombinant Protein ◽  
Cell Lines ◽  
Translation Initiation ◽  
Mammalian Cells ◽  
Mrna Translation ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Cho Cell

Many protein-based biotherapeutics are produced in cultured Chinese hamster ovary (CHO) cell lines. Recent reports have demonstrated that translation of recombinant mRNAs and global control of the translation machinery via mammalian target of rapamycin (mTOR) signalling are important determinants of the amount and quality of recombinant protein such cells can produce. mTOR complex 1 (mTORC1) is a master regulator of cell growth/division, ribosome biogenesis and protein synthesis, but the relationship between mTORC1 signalling, cell growth and proliferation and recombinant protein yields from mammalian cells, and whether this master regulating signalling pathway can be manipulated to enhance cell biomass and recombinant protein production (rPP) are not well explored. We have investigated mTORC1 signalling and activity throughout batch culture of a panel of sister recombinant glutamine synthetase-CHO cell lines expressing different amounts of a model monoclonal IgG4, to evaluate the links between mTORC1 signalling and cell proliferation, autophagy, recombinant protein expression, global protein synthesis and mRNA translation initiation. We find that the expression of the mTORC1 substrate 4E-binding protein 1 (4E-BP1) fluctuates throughout the course of cell culture and, as expected, that the 4E-BP1 phosphorylation profiles change across the culture. Importantly, we find that the eIF4E/4E-BP1 stoichiometry positively correlates with cell productivity. Furthermore, eIF4E amounts appear to be co-regulated with 4E-BP1 amounts. This may reflect a sensing of either change at the mRNA level as opposed to the protein level or the fact that the phosphorylation status, as well as the amount of 4E-BP1 present, is important in the co-regulation of eIF4E and 4E-BP1.

scholarly journals Complementation of Cold Shock Proteins by Translation Initiation Factor IF1 In Vivo

2001 ◽  
Vol 183 (24) ◽  
pp. 7381-7386 ◽  
Author(s):  
Michael H. W. Weber ◽  
Carsten L. Beckering ◽  
Mohamed A. Marahiel
Keyword(s):  
Translation Initiation ◽  
Cold Shock ◽  
Optimal Growth ◽  
Stationary Growth Phase ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Cold Shock Proteins ◽  
Double Deletion ◽  
Shock Proteins

ABSTRACT The cold shock response in both Escherichia coli andBacillus subtilis is induced by an abrupt downshift in growth temperature and leads to a dramatic increase in the production of a homologous class of small, often highly acidic cold shock proteins. This protein family is the prototype of the cold shock domain (CSD) that is conserved from bacteria to humans. For B. subtilis it has been shown that at least one of the three resident cold shock proteins (CspB to D) is essential under optimal growth conditions as well as during cold shock. Analysis of theB. subtilis cspB cspC double deletion mutant revealed that removal of these csp genes results in pleiotropic alteration of protein synthesis, cell lysis during the entry of stationary growth phase, and the inability to differentiate into endospores. We show here that heterologous expression of the translation initiation factor IF1 from E. coli in aB. subtilis cspB cspC double deletion strain is able to cure both the growth and the sporulation defects observed for this mutant, suggesting that IF1 and cold shock proteins have at least in part overlapping cellular function(s). Two of the possible explanation models are discussed.

scholarly journals Inhibition of the Ubiquitin-Proteasome System Induces Stress Granule Formation

2007 ◽  
Vol 18 (7) ◽  
pp. 2603-2618 ◽  
Author(s):  
Rachid Mazroui ◽  
Sergio Di Marco ◽  
Randal J. Kaufman ◽  
Imed-Eddine Gallouzi
Keyword(s):  
Mrna Decay ◽  
Proteasome Inhibitors ◽  
Ubiquitin Proteasome System ◽  
Early Response ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Dependent Manner ◽  
Cellular Mechanisms ◽  
Granule Formation ◽  
Eif2α Phosphorylation

The inhibition of the ubiquitin-dependent proteasome system (UPS) via specific drugs is one type of approach used to combat cancer. Although it has been suggested that UPS inhibition prevents the rapid decay of AU-rich element (ARE)-containing messages, very little is known about the cellular mechanisms leading to this effect. Here we establish a link between the inhibition of UPS activity, the formation of cytoplasmic stress granules (SGs), and mRNA metabolism. The assembly of the SGs requires the phosphorylation of the translation initiation factor eIF2α by a mechanism involving the stress kinase GCN2. On prolonged UPS inhibition and despite the maintenance of eIF2α phosphorylation, SGs disassemble and translation recovers in an Hsp72 protein-dependent manner. The formation of these SGs coincides with the disassembly of processing bodies (PBs), known as mRNA decay entities. As soon as the SGs assemble, they recruit ARE-containing messages such as p21cip1 mRNA, which are stabilized under these conditions. Hence, our findings suggest that SGs could be considered as one of the players that mediate the early response of the cell to proteasome inhibitors by interfering temporarily with mRNA decay pathways.

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 The Leader Protein of Theiler's Virus Prevents the Activation of PKR

2019 ◽  
Vol 93 (19) ◽  
Author(s):  
Fabian Borghese ◽  
Frédéric Sorgeloos ◽  
Teresa Cesaro ◽  
Thomas Michiels
Keyword(s):  
Stress Granule ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Wild Type Virus ◽  
Granule Formation ◽  
Double Stranded Rna ◽  
Eif2α Phosphorylation ◽  
Leader Protein ◽  
Infected Cells ◽  
L Proteins

ABSTRACT Leader (L) proteins encoded by cardioviruses are multifunctional proteins that contribute to innate immunity evasion. L proteins of Theiler’s murine encephalomyelitis virus (TMEV), Saffold virus (SAFV), and encephalomyocarditis virus (EMCV) were reported to inhibit stress granule assembly in infected cells. Here, we show that TMEV L can act at two levels in the stress granule formation pathway: on the one hand, it can inhibit sodium arsenite-induced stress granule assembly without preventing eIF2α phosphorylation and, thus, acts downstream of eIF2α; on the other hand, it can inhibit eucaryotic translation initiation factor 2 alpha kinase 2 (PKR) activation and the consequent PKR-mediated eIF2α phosphorylation. Interestingly, coimmunostaining experiments revealed that PKR colocalizes with viral double-stranded RNA (dsRNA) in cells infected with L-mutant viruses but not in cells infected with the wild-type virus. Furthermore, PKR coprecipitated with dsRNA from cells infected with L-mutant viruses significantly more than from cells infected with the wild-type virus. These data strongly suggest that L blocks PKR activation by preventing the interaction between PKR and viral dsRNA. In infected cells, L also rendered PKR refractory to subsequent activation by poly(I·C). However, no interaction was observed between L and either dsRNA or PKR. Taken together, our results suggest that, unlike other viral proteins, L indirectly acts on PKR to negatively regulate its responsiveness to dsRNA. IMPORTANCE The leader (L) protein encoded by cardioviruses is a very short multifunctional protein that contributes to evasion of the host innate immune response. This protein notably prevents the formation of stress granules in infected cells. Using Theiler’s virus as a model, we show that L proteins can act at two levels in the stress response pathway leading to stress granule formation, the most striking one being the inhibition of eucaryotic translation initiation factor 2 alpha kinase 2 (PKR) activation. Interestingly, the leader protein appears to inhibit PKR via a novel mechanism by rendering this kinase unable to detect double-stranded RNA, its typical activator. Unlike other viral proteins, such as influenza virus NS1, the leader protein appears to interact with neither PKR nor double-stranded RNA, suggesting that it acts indirectly to trigger the inhibition of the kinase.

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