scholarly journals Mutational analysis of the alpha subunit of eIF2B provides insights into the role of eIF2B bodies in translational control and VWM disease

2020 ◽  
pp. jbc.RA120.014956
Author(s):  
Karl Norris ◽  
Rachel E Hodgson ◽  
Tawni Dornelles ◽  
K Elizabeth Allen ◽  
Ben M Abell ◽  
...  
Keyword(s):  
Translational Control ◽  
Translational Regulation ◽  
Mutational Analysis ◽  
Control Point ◽  
The Body ◽  
Initiation Factor ◽  
Regulatory Subunit ◽  
Missense Mutations ◽  
Eif2α Phosphorylation

Eukaryotic initiation factor 2B (eIF2B) serves as a vital control point within protein synthesis and regulates translation initiation in response to cellular stress. Mutations within eIF2B result in the fatal disease, leukoencephalopathy with vanishing white matter (VWM). Previous biochemical studies on VWM mutations have illustrated that changes in the activity of eIF2B poorly correlates with disease severity. This suggests that there may be additional characteristics of eIF2B contributing to VWM pathogenesis. Here, we investigated whether the localisation of eIF2B to eIF2B bodies was integral for function and whether this localisation could provide insight into the pathogenesis of VWM. We demonstrate that the regulatory subunit, eIF2Bα, is required for the assembly of eIF2B bodies in yeast and that loss of eIF2B bodies correlates with an inability of cells to regulate eIF2B activity.  Mutational analysis of eIF2Bα showed that missense mutations which disrupt the regulation of eIF2B similarly disrupt the assembly of eIF2B bodies. In contrast, when eIF2Bα mutations which impact the catalytic activity of eIF2B were analysed, eIF2B bodies were absent and instead eIF2B localised to small foci, termed microfoci. FRAP analysis highlighted that within these microfoci, eIF2 shuttles more slowly indicating that formation of eIF2B bodies correlates with full eIF2B activity. When eIF2Bα VWM mutations were analysed a diverse impact on localisation was observed, which did not seem to correlate with eIF2B activity.  These findings provide key insights into how the eIF2B body assembles and suggest that the body is a fundamental part of the translational regulation via eIF2α phosphorylation.

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 Pharmacological dimerization and activation of the exchange factor eIF2B antagonizes the integrated stress response

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Carmela Sidrauski ◽  
Jordan C Tsai ◽  
Martin Kampmann ◽  
Brian R Hearn ◽  
Punitha Vedantham ◽  
...  
Keyword(s):  
Stress Response ◽  
Translational Control ◽  
Control Point ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Integrated Stress Response ◽  
Nucleotide Exchange ◽  
Eif2α Phosphorylation ◽  
Symmetric Molecule ◽  
Invaluable Tool

The general translation initiation factor eIF2 is a major translational control point. Multiple signaling pathways in the integrated stress response phosphorylate eIF2 serine-51, inhibiting nucleotide exchange by eIF2B. ISRIB, a potent drug-like small molecule, renders cells insensitive to eIF2α phosphorylation and enhances cognitive function in rodents by blocking long-term depression. ISRIB was identified in a phenotypic cell-based screen, and its mechanism of action remained unknown. We now report that ISRIB is an activator of eIF2B. Our reporter-based shRNA screen revealed an eIF2B requirement for ISRIB activity. Our results define ISRIB as a symmetric molecule, show ISRIB-mediated stabilization of activated eIF2B dimers, and suggest that eIF2B4 (δ-subunit) contributes to the ISRIB binding site. We also developed new ISRIB analogs, improving its EC50 to 600 pM in cell culture. By modulating eIF2B function, ISRIB promises to be an invaluable tool in proof-of-principle studies aiming to ameliorate cognitive defects resulting from neurodegenerative diseases.

scholarly journals The Regulation of Hepatic Protein Synthesis during Fasting in the Rat

2005 ◽  
Vol 280 (16) ◽  
pp. 16427-16436 ◽  
Author(s):  
Padmanabhan Anand ◽  
Philip A. Gruppuso
Keyword(s):  
Protein Synthesis ◽  
Translational Control ◽  
Translational Regulation ◽  
Initiation Factor ◽  
Pcr Analysis ◽  
Mrna Levels ◽  
Eif2α Phosphorylation ◽  
Cap Binding Complex ◽  
Branched Chain ◽  
Rps6 Phosphorylation

We have studied translational control in the model of 48 h of fasting in the rat. Our initial observations showed a paradoxical increase in ribosomal protein S6 (rpS6) phosphorylation and a decrease in eukaryotic initiation factor 2α (eIF2α) phosphorylation. These effects, which would favor an increase in protein synthesis, could be attributed to increased circulating concentrations of branched-chain amino acids in fasting. To determine what mechanisms might account for decreased hepatic translation in fasting, we examined the cap binding complex. eIF4E-bound 4E-BP1 did not increase. However, eIF4E-bound eIF4G and total cellular eIF4G were profoundly decreased in fasted liver. eIF4G mRNA levels were not lower after fasting. Based on the hypothesis that decreased eIF4G translation might account for the reduced eIF4G content, we fractionated ribosomes by sucrose density centrifugation. Immunoblotting for rpS6 showed modest polysomal disaggregation upon fasting. PCR analysis of polysome profiles revealed that a spectrum of mRNAs undergo different translational regulation in the fasted state. In particular, eIF4G was minimally affected by fasting. This indicated that reduced eIF4G abundance in fasting may be a function of its stability, whereas its recovery upon refeeding is necessarily independent of its own involvement in the cap binding complex. Western immunoblotting of polysome fractions showed that phosphorylated rpS6 was disproportionately present in translating polysomes in fed and fasted animals, consistent with a role in translational control. However, the translation of rpS8, an mRNA with a 5′-oligopyrimidine tract, did not coincide with rpS6 phosphorylation, thus dissociating rpS6 phosphorylation from the translational control of this subset of mRNAs.

scholarly journals A Retrospective on eIF2A—and Not the Alpha Subunit of eIF2

2020 ◽  
Vol 21 (6) ◽  
pp. 2054
Author(s):  
Anton A. Komar ◽  
William C. Merrick
Keyword(s):  
Translational Control ◽  
Ribosomal Subunit ◽  
Initiation Factor ◽  
Single Chain ◽  
Initiation Factors ◽  
Eif2α Phosphorylation ◽  
Specific Mrnas ◽  
Internal Initiation ◽  
And Function ◽  
Polypeptide Chains

Initiation of protein synthesis in eukaryotes is a complex process requiring more than 12 different initiation factors, comprising over 30 polypeptide chains. The functions of many of these factors have been established in great detail; however, the precise role of some of them and their mechanism of action is still not well understood. Eukaryotic initiation factor 2A (eIF2A) is a single chain 65 kDa protein that was initially believed to serve as the functional homologue of prokaryotic IF2, since eIF2A and IF2 catalyze biochemically similar reactions, i.e., they stimulate initiator Met-tRNAi binding to the small ribosomal subunit. However, subsequent identification of a heterotrimeric 126 kDa factor, eIF2 (α,β,γ) showed that this factor, and not eIF2A, was primarily responsible for the binding of Met-tRNAi to 40S subunit in eukaryotes. It was found however, that eIF2A can promote recruitment of Met-tRNAi to 40S/mRNA complexes under conditions of inhibition of eIF2 activity (eIF2α-phosphorylation), or its absence. eIF2A does not function in major steps in the initiation process, but is suggested to act at some minor/alternative initiation events such as re-initiation, internal initiation, or non-AUG initiation, important for translational control of specific mRNAs. This review summarizes our current understanding of the eIF2A structure and function.

scholarly journals Neuronal BC RNAs cooperate with eIF4B to mediate activity-dependent translational control

2014 ◽  
Vol 207 (2) ◽  
pp. 237-252 ◽  
Author(s):  
Taesun Eom ◽  
Ilham A. Muslimov ◽  
Panayiotis Tsokas ◽  
Valerio Berardi ◽  
Jun Zhong ◽  
...  
Keyword(s):  
Neuronal Activity ◽  
Translational Control ◽  
Translational Regulation ◽  
Regulation Of Gene Expression ◽  
Ribosomal Subunit ◽  
Initiation Factor ◽  
Neuronal Stimulation ◽  
Mechanistic Basis ◽  
Protein Kinase Mζ ◽  
Activity Dependent

In neurons, translational regulation of gene expression has been implicated in the activity-dependent management of synapto-dendritic protein repertoires. However, the fundamentals of stimulus-modulated translational control in neurons remain poorly understood. Here we describe a mechanism in which regulatory brain cytoplasmic (BC) RNAs cooperate with eukaryotic initiation factor 4B (eIF4B) to control translation in a manner that is responsive to neuronal activity. eIF4B is required for the translation of mRNAs with structured 5′ untranslated regions (UTRs), exemplified here by neuronal protein kinase Mζ (PKMζ) mRNA. Upon neuronal stimulation, synapto-dendritic eIF4B is dephosphorylated at serine 406 in a rapid process that is mediated by protein phosphatase 2A. Such dephosphorylation causes a significant decrease in the binding affinity between eIF4B and BC RNA translational repressors, enabling the factor to engage the 40S small ribosomal subunit for translation initiation. BC RNA translational control, mediated via eIF4B phosphorylation status, couples neuronal activity to translational output, and thus provides a mechanistic basis for long-term plastic changes in nerve cells.

scholarly journals A Blood Pact: the Significance and Implications of eIF4E on Lymphocytic Leukemia

2018 ◽  
pp. 363-382
Author(s):  
V. VENTURI ◽  
T. MASEK ◽  
M. POSPISEK
Keyword(s):  
Translational Control ◽  
Lymphoblastic Leukemia ◽  
Lymphocytic Leukemia ◽  
Initiation Factor ◽  
Oncogenic Signaling ◽  
Translation Factor ◽  
Eukaryotic Initiation Factor 4E ◽  
Biological Circuits ◽  
Oncogenic Signaling Pathways

Elevated levels of eukaryotic initiation factor 4E (eIF4E) are implicated in neoplasia, with cumulative evidence pointing to its role in the etiopathogenesis of hematological diseases. As a node of convergence for several oncogenic signaling pathways, eIF4E has attracted a great deal of interest from biologists and clinicians whose efforts have been targeting this translation factor and its biological circuits in the battle against leukemia. The role of eIF4E in myeloid leukemia has been ascertained and drugs targeting its functions have found their place in clinical trials. Little is known, however, about the pertinence of eIF4E to the biology of lymphocytic leukemia and a paucity of literature is available in this regard that prospectively evaluates the topic to guide practice in hematological cancer. A comprehensive analysis on the significance of eIF4E translation factor in the clinical picture of leukemia arises, therefore, as a compelling need. This review presents aspects of eIF4E involvement in the realm of the lymphoblastic leukemia status; translational control of immunological function via eIF4E and the state-of-the-art in drugs will also be outlined.

scholarly journals Inhibition of a constitutive translation initiation factor 2α phosphatase, CReP, promotes survival of stressed cells

2003 ◽  
Vol 163 (4) ◽  
pp. 767-775 ◽  
Author(s):  
Céline Jousse ◽  
Seiichi Oyadomari ◽  
Isabel Novoa ◽  
Phoebe Lu ◽  
Yuhong Zhang ◽  
...  
Keyword(s):  
Phosphatase Activity ◽  
Translation Initiation ◽  
Mammalian Cells ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Regulatory Subunit ◽  
Gene Encoding ◽  
Eukaryotic Translation ◽  
Eif2α Phosphorylation ◽  
Stressed Cells

Phosphorylation of eukaryotic translation initiation factor 2α (eIF2α) on serine 51 is effected by specific stress-activated protein kinases. eIF2α phosphorylation inhibits translation initiation promoting a cytoprotective gene expression program known as the integrated stress response (ISR). Stress-induced activation of GADD34 feeds back negatively on this pathway by promoting eIF2α dephosphorylation, however, GADD34 mutant cells retain significant eIF2α-directed phosphatase activity. We used a somatic cell genetic approach to identify a gene encoding a novel regulatory subunit of a constitutively active holophosphatase complex that dephosphorylates eIF2α. RNAi of this gene, which we named constitutive repressor of eIF2α phosphorylation (CReP, or PPP1R15B), repressed the constitutive eIF2α-directed phosphatase activity and activated the ISR. CReP RNAi strongly protected mammalian cells against oxidative stress, peroxynitrite stress, and more modestly against accumulation of malfolded proteins in the endoplasmic reticulum. These findings suggest that therapeutic inhibition of eIF2α dephosphorylation by targeting the CReP-protein–phosphatase-1 complex may be used to access the salubrious qualities of the ISR.

scholarly journals Established and Emerging Regulatory Roles of Eukaryotic Translation Initiation Factor 5B (eIF5B)

2021 ◽  
Vol 12 ◽  
Author(s):  
Prakash Amruth Raj Chukka ◽  
Stacey D. Wetmore ◽  
Nehal Thakor
Keyword(s):  
Translation Initiation ◽  
Translational Control ◽  
Translational Regulation ◽  
Eukaryotic Cell ◽  
Translation Initiation Factor ◽  
Initiation Factor ◽  
Eukaryotic Translation ◽  
Cellular Mrnas ◽  
Eukaryotic Translation Initiation ◽  
Initiation Stage

Translational control (TC) is one the crucial steps that dictate gene expression and alter the outcome of physiological process like programmed cell death, metabolism, and proliferation in a eukaryotic cell. TC occurs mainly at the translation initiation stage. The initiation factor eIF5B tightly regulates global translation initiation and facilitates the expression of a subset of proteins involved in proliferation, inhibition of apoptosis, and immunosuppression under stress conditions. eIF5B enhances the expression of these survival proteins to allow cancer cells to metastasize and resist chemotherapy. Using eIF5B as a biomarker or drug target could help with diagnosis and improved prognosis, respectively. To achieve these goals, it is crucial to understand the role of eIF5B in translational regulation. This review recapitulates eIF5B’s regulatory roles in the translation initiation of viral mRNA as well as the cellular mRNAs in cancer and stressed eukaryotic cells.

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