The Experts below are selected from a list of 120 Experts worldwide ranked by ideXlab platform
Jilong Chen - One of the best experts on this subject based on the ideXlab platform.
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influenza a virus induced degradation of eukaryotic translation Initiation Factor 4B contributes to viral replication by suppressing ifitm3 protein expression
Journal of Virology, 2014Co-Authors: Song Wang, Xiaojuan Chi, Haitao Wei, Yuhai Chen, Zhilong Chen, Shile Huang, Jilong ChenAbstract:Although alteration in host cellular translation machinery occurs in virus-infected cells, the role of such alteration and the precise pathogenic processes are not well understood. Influenza A virus (IAV) infection shuts off host cell gene expression at transcriptional and translational levels. Here, we found that the protein level of eukaryotic translation Initiation Factor 4B (eIF4B), an integral component of the translation Initiation apparatus, was dramatically reduced in A549 cells as well as in the lung, spleen, and thymus of mice infected with IAV. The decrease in eIF4B level was attributed to lysosomal degradation of eIF4B, which was induced by viral NS1 protein. Silencing eIF4B expression in A549 cells significantly promoted IAV replication, and conversely, overexpression of eIF4B markedly inhibited the viral replication. Importantly, we observed that eIF4B knockdown transgenic mice were more susceptible to IAV infection, exhibiting faster weight loss, shorter survival time, and more-severe organ damage. Furthermore, we demonstrated that eIF4B regulated the expression of interferon-induced transmembrane protein 3 (IFITM3), a critical protein involved in immune defense against a variety of RNA viruses, including influenza virus. Taken together, our findings reveal that eIF4B plays an important role in host defense against IAV infection at least by regulating the expression of IFITM3, which restricts viral entry and thereby blocks early stages of viral production. These data also indicate that influenza virus has evolved a strategy to overcome host innate immunity by downregulating eIF4B protein. IMPORTANCE Influenza A virus (IAV) infection stimulates the host innate immune system, in part, by inducing interferons (IFNs). Secreted IFNs activate the Janus kinase/signal transducers and activators of transcription (JAK/STAT) pathway, leading to elevated transcription of a large group of IFN-stimulated genes that have antiviral function. To circumvent the host innate immune response, influenza virus has evolved multiple strategies for suppressing the production of IFNs. Here, we show that IAV infection induces lysosomal degradation of eIF4B protein; and eIF4B inhibits IAV replication by upregulating expression of interferon-induced transmembrane protein 3 (IFITM3), a key protein that protects the host from virus infection. Our finding illustrates a critical role of eIF4B in the host innate immune response and provides novel insights into the complex mechanisms by which influenza virus interacts with its host.
Sarah E Walker - One of the best experts on this subject based on the ideXlab platform.
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the n terminal domain of eukaryotic Initiation Factor 4B drives yeast translational control in response to urea
bioRxiv, 2021Co-Authors: Xiaozhuo Liu, Houtan Moshiri, Ansuman Sahoo, Sarah E WalkerAbstract:The yeast eukaryotic Initiation Factor 4B binds the 40S subunit in translation preInitiation complexes (PICs), promoting mRNA binding. Recent evidence suggests mRNAs have variable dependence on eIF4B, suggesting this Factor could promote changes in mRNA selection for translation, in order to adapt to stressors. However, the importance of eIF4B and its constituent domains for mRNA selection under diverse cellular and environmental conditions remain undefined. Here we compared the effects of disrupting eIF4B RNA- and ribosome-binding motifs under ~1400 growth conditions. The RNA-Recognition Motif (RRM) was dispensable for stress responses, but the 40S-binding N-terminal Domain (NTD) promoted growth in response to various stressors. In particular, the NTD conferred a strong growth advantage in the presence of urea. Ribosome profiling revealed that the NTD promoted translation of mRNAs with long and highly structured 5-prime untranslated regions, both with and without urea exposure. Our results suggest eIF4B controls mRNA loading and scanning as a part of the PIC, rather than by activating mRNPs prior to ribosome binding. Furthermore, our data indicate the yeast response to urea includes a translational component, driven by production of proteins associated with the cellular periphery. Together our analyses suggest general eIFs can promote diverse cellular responses.
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Binding to the Ribosome by Eukaryotic Initiation Factor 4B Drives Yeast Translational Control in Response to Urea
2019Co-Authors: Xiaozhuo Liu, Houtan Moshiri, Sarah E WalkerAbstract:ABSTRACT The yeast eukaryotic Initiation Factor 4B binds the 40S subunit in translation preInitiation complexes (PICs), promoting mRNA binding. Recent evidence suggests mRNAs have variable dependence on eIF4B, suggesting this Factor could promote changes in mRNA selection to adapt to stressors. However, the importance of eIF4B and its constituent domains for mRNA selection under diverse cellular and environmental conditions remain undefined. Here we compared the effects of disrupting eIF4B RNA- and ribosome-binding under ∼1400 growth conditions. The RNA-Recognition Motif (RRM) was dispensable for stress responses, but ribosome binding by the N-terminal Domain (NTD) promoted growth in response to various stressors. In particular, the NTD conferred a strong growth advantage in the presence of urea. Ribosome profiling revealed that the NTD promoted translation of mRNAs with long and highly structured 5-prime untranslated regions, both with and without urea exposure. Because these changes required 40S binding, our results suggest eIF4B regulates mRNA loading and scanning as a part of the PIC, rather than by activating mRNPs prior to ribosome binding. Furthermore, our data indicate the yeast response to urea includes a translational component, driven by translation of mRNAs encoding proteins associated with the cellular periphery, suggesting general eIFs can promote diverse cellular responses.
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identification and characterization of functionally critical conserved motifs in the internal repeats and n terminal domain of yeast translation Initiation Factor 4B yeif4B
Journal of Biological Chemistry, 2014Co-Authors: Fujun Zhou, Sarah E Walker, Jon R Lorsch, Sarah F Mitchell, Alan G HinnebuschAbstract:Abstract eIF4B has been implicated in attachment of the 43S preInitiation complex (PIC) to mRNAs and scanning to the start codon. We recently determined that the internal 7 repeats (of ~26 amino acids each) of S. cerevisiae eIF4B (yeIF4B) comprise the region most critically required to enhance mRNA recruitment by 43S PICs in vitro and stimulate general translation Initiation in yeast. Moreover, whereas the N-terminal domain (NTD) of yeIF4B contributes to these activities, the RNA recognition motif (RRM) is dispensable. We have now determined that only 2 of the 7 internal repeats are sufficient for wild-type (WT) yeIF4B function in vivo when all other domains are intact. However, three or more repeats are needed in the absence of the NTD or when the functions of eIF4F components are compromised. We corroborated these observations in the reconstituted system by demonstrating that yeIF4B variants with only one or two repeats display substantial activity in promoting mRNA recruitment by the PIC, whereas additional repeats are required at lower levels of eIF4A or when the NTD is missing. These findings indicate functional overlap among the 7-repeat and NTD domains of yeIF4B and eIF4A in mRNA recruitment. Interestingly, only three highly conserved positions in the 26-aa repeat are essential for function in vitro and in vivo. Finally, we identified conserved motifs in the NTD and demonstrate functional overlap of two such motifs. These results provide a comprehensive description of the critical sequence elements in yeIF4B that support eIF4F function in mRNA recruitment by the PIC.
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identification and characterization of functionally critical conserved motifs in the internal repeats and n terminal domain of yeast translation Initiation Factor 4B yeif4B
Journal of Biological Chemistry, 2014Co-Authors: Fujun Zhou, Sarah E Walker, Jon R Lorsch, Sarah F Mitchell, Alan G HinnebuschAbstract:eIF4B has been implicated in attachment of the 43 S preInitiation complex (PIC) to mRNAs and scanning to the start codon. We recently determined that the internal seven repeats (of ∼26 amino acids each) of Saccharomyces cerevisiae eIF4B (yeIF4B) compose the region most critically required to enhance mRNA recruitment by 43 S PICs in vitro and stimulate general translation Initiation in yeast. Moreover, although the N-terminal domain (NTD) of yeIF4B contributes to these activities, the RNA recognition motif is dispensable. We have now determined that only two of the seven internal repeats are sufficient for wild-type (WT) yeIF4B function in vivo when all other domains are intact. However, three or more repeats are needed in the absence of the NTD or when the functions of eIF4F components are compromised. We corroborated these observations in the reconstituted system by demonstrating that yeIF4B variants with only one or two repeats display substantial activity in promoting mRNA recruitment by the PIC, whereas additional repeats are required at lower levels of eIF4A or when the NTD is missing. These findings indicate functional overlap among the 7-repeats and NTD domains of yeIF4B and eIF4A in mRNA recruitment. Interestingly, only three highly conserved positions in the 26-amino acid repeat are essential for function in vitro and in vivo. Finally, we identified conserved motifs in the NTD and demonstrate functional overlap of two such motifs. These results provide a comprehensive description of the critical sequence elements in yeIF4B that support eIF4F function in mRNA recruitment by the PIC.
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yeast eukaryotic Initiation Factor 4B eif4B enhances complex assembly between eif4a and eif4g in vivo
Journal of Biological Chemistry, 2013Co-Authors: Eun Hee Park, Sarah E Walker, Fujun Zhou, Joseph M Lee, Vaishnavi Rajagopal, Jon R Lorsch, Alan G HinnebuschAbstract:Translation Initiation Factor eIF4F (eukaryotic Initiation Factor 4F), composed of eIF4E, eIF4G, and eIF4A, binds to the m7G cap structure of mRNA and stimulates recruitment of the 43S preInitiation complex and subsequent scanning to the Initiation codon. The HEAT domain of eIF4G stabilizes the active conformation of eIF4A required for its RNA helicase activity. Mammalian eIF4B also stimulates eIF4A activity, but this function appears to be lacking in yeast, making it unclear how yeast eIF4B (yeIF4B/Tif3) stimulates translation. We identified Ts− mutations in the HEAT domains of yeast eIF4G1 and eIF4G2 that are suppressed by overexpressing either yeIF4B or eIF4A, whereas others are suppressed only by eIF4A overexpression. Importantly, suppression of HEAT domain substitutions by yeIF4B overexpression was correlated with the restoration of native eIF4A·eIF4G complexes in vivo, and the rescue of specific mutant eIF4A·eIF4G complexes by yeIF4B was reconstituted in vitro. Association of eIF4A with WT eIF4G in vivo also was enhanced by yeIF4B overexpression and was impaired in cells lacking yeIF4B. Furthermore, we detected native complexes containing eIF4G and yeIF4B but lacking eIF4A. These and other findings lead us to propose that yeIF4B acts in vivo to promote eIF4F assembly by enhancing a conformation of the HEAT domain of yeast eIF4G conducive for stable binding to eIF4A.
Alan G Hinnebusch - One of the best experts on this subject based on the ideXlab platform.
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identification and characterization of functionally critical conserved motifs in the internal repeats and n terminal domain of yeast translation Initiation Factor 4B yeif4B
Journal of Biological Chemistry, 2014Co-Authors: Fujun Zhou, Sarah E Walker, Jon R Lorsch, Sarah F Mitchell, Alan G HinnebuschAbstract:eIF4B has been implicated in attachment of the 43 S preInitiation complex (PIC) to mRNAs and scanning to the start codon. We recently determined that the internal seven repeats (of ∼26 amino acids each) of Saccharomyces cerevisiae eIF4B (yeIF4B) compose the region most critically required to enhance mRNA recruitment by 43 S PICs in vitro and stimulate general translation Initiation in yeast. Moreover, although the N-terminal domain (NTD) of yeIF4B contributes to these activities, the RNA recognition motif is dispensable. We have now determined that only two of the seven internal repeats are sufficient for wild-type (WT) yeIF4B function in vivo when all other domains are intact. However, three or more repeats are needed in the absence of the NTD or when the functions of eIF4F components are compromised. We corroborated these observations in the reconstituted system by demonstrating that yeIF4B variants with only one or two repeats display substantial activity in promoting mRNA recruitment by the PIC, whereas additional repeats are required at lower levels of eIF4A or when the NTD is missing. These findings indicate functional overlap among the 7-repeats and NTD domains of yeIF4B and eIF4A in mRNA recruitment. Interestingly, only three highly conserved positions in the 26-amino acid repeat are essential for function in vitro and in vivo. Finally, we identified conserved motifs in the NTD and demonstrate functional overlap of two such motifs. These results provide a comprehensive description of the critical sequence elements in yeIF4B that support eIF4F function in mRNA recruitment by the PIC.
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identification and characterization of functionally critical conserved motifs in the internal repeats and n terminal domain of yeast translation Initiation Factor 4B yeif4B
Journal of Biological Chemistry, 2014Co-Authors: Fujun Zhou, Sarah E Walker, Jon R Lorsch, Sarah F Mitchell, Alan G HinnebuschAbstract:Abstract eIF4B has been implicated in attachment of the 43S preInitiation complex (PIC) to mRNAs and scanning to the start codon. We recently determined that the internal 7 repeats (of ~26 amino acids each) of S. cerevisiae eIF4B (yeIF4B) comprise the region most critically required to enhance mRNA recruitment by 43S PICs in vitro and stimulate general translation Initiation in yeast. Moreover, whereas the N-terminal domain (NTD) of yeIF4B contributes to these activities, the RNA recognition motif (RRM) is dispensable. We have now determined that only 2 of the 7 internal repeats are sufficient for wild-type (WT) yeIF4B function in vivo when all other domains are intact. However, three or more repeats are needed in the absence of the NTD or when the functions of eIF4F components are compromised. We corroborated these observations in the reconstituted system by demonstrating that yeIF4B variants with only one or two repeats display substantial activity in promoting mRNA recruitment by the PIC, whereas additional repeats are required at lower levels of eIF4A or when the NTD is missing. These findings indicate functional overlap among the 7-repeat and NTD domains of yeIF4B and eIF4A in mRNA recruitment. Interestingly, only three highly conserved positions in the 26-aa repeat are essential for function in vitro and in vivo. Finally, we identified conserved motifs in the NTD and demonstrate functional overlap of two such motifs. These results provide a comprehensive description of the critical sequence elements in yeIF4B that support eIF4F function in mRNA recruitment by the PIC.
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yeast eukaryotic Initiation Factor 4B eif4B enhances complex assembly between eif4a and eif4g in vivo
Journal of Biological Chemistry, 2013Co-Authors: Eun Hee Park, Sarah E Walker, Fujun Zhou, Joseph M Lee, Vaishnavi Rajagopal, Jon R Lorsch, Alan G HinnebuschAbstract:Translation Initiation Factor eIF4F (eukaryotic Initiation Factor 4F), composed of eIF4E, eIF4G, and eIF4A, binds to the m7G cap structure of mRNA and stimulates recruitment of the 43S preInitiation complex and subsequent scanning to the Initiation codon. The HEAT domain of eIF4G stabilizes the active conformation of eIF4A required for its RNA helicase activity. Mammalian eIF4B also stimulates eIF4A activity, but this function appears to be lacking in yeast, making it unclear how yeast eIF4B (yeIF4B/Tif3) stimulates translation. We identified Ts− mutations in the HEAT domains of yeast eIF4G1 and eIF4G2 that are suppressed by overexpressing either yeIF4B or eIF4A, whereas others are suppressed only by eIF4A overexpression. Importantly, suppression of HEAT domain substitutions by yeIF4B overexpression was correlated with the restoration of native eIF4A·eIF4G complexes in vivo, and the rescue of specific mutant eIF4A·eIF4G complexes by yeIF4B was reconstituted in vitro. Association of eIF4A with WT eIF4G in vivo also was enhanced by yeIF4B overexpression and was impaired in cells lacking yeIF4B. Furthermore, we detected native complexes containing eIF4G and yeIF4B but lacking eIF4A. These and other findings lead us to propose that yeIF4B acts in vivo to promote eIF4F assembly by enhancing a conformation of the HEAT domain of yeast eIF4G conducive for stable binding to eIF4A.
Nahum Sonenberg - One of the best experts on this subject based on the ideXlab platform.
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Control of cell survival and proliferation by mammalian eukaryotic Initiation Factor 4B.
Molecular and cellular biology, 2010Co-Authors: David Shahbazian, Armen Parsyan, Emmanuel Petroulakis, Ivan Topisirovic, Yvan Martineau, Bernard F. Gibbs, Yuri V. Svitkin, Nahum SonenbergAbstract:Translation Initiation plays an important role in cell growth, proliferation, and survival. The translation Initiation Factor eIF4B (eukaryotic Initiation Factor 4B) stimulates the RNA helicase activity of eIF4A in unwinding secondary structures in the 5′ untranslated region (5′UTR) of the mRNA in vitro. Here, we studied the effects of eIF4B depletion in cells using RNA interference (RNAi). In agreement with the role of eIF4B in translation Initiation, its depletion resulted in inhibition of this step. Selective reduction of translation was observed for mRNAs harboring strong to moderate secondary structures in their 5′UTRs. These mRNAs encode proteins, which function in cell proliferation (Cdc25C, c-myc, and ODC [ornithine decarboxylase]) and survival (Bcl-2 and XIAP [X-linked inhibitor of apoptosis]). Furthermore, eIF4B silencing led to decreased proliferation rates, promoted caspase-dependent apoptosis, and further sensitized cells to camptothecin-induced cell death. These results demonstrate that eIF4B is required for cell proliferation and survival by regulating the translation of proliferative and prosurvival mRNAs.
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14 3 3σ controls mitotic translation to facilitate cytokinesis
Nature, 2007Co-Authors: Erik Wilker, Nahum Sonenberg, Yun Feng, Marcel A. T. M. Van Vugt, Stephen C. Artim, Paul H. Huang, Christian P. Petersen, Phillip A. Sharp, Christian H Reinhardt, Forest M. WhiteAbstract:The protein known as 14-3-3σ is an important p53-regulated human tumour suppressor that is lost early in cancer development. A possible molecular basis for its function has now been found. While mRNA translation normally switches from a 'cap-dependent' to 'cap-independent' mechanism during mitosis, cells lacking 14-3-3σ don't make this switch, and consequently have cell division abnormalities. A function for 14-3-3σ in regulating the translation of proteins in mitosis is defined. During mitosis, translation normally switches from a 'cap-dependent' to 'cap-independent' mechanism, but cells lacking 14-3-3σ don't make this switch and consequently have difficulties in cell division. 14-3-3 proteins are crucial in a wide variety of cellular responses including cell cycle progression, DNA damage checkpoints and apoptosis. One particular 14-3-3 isoform, σ, is a p53-responsive gene, the function of which is frequently lost in human tumours, including breast and prostate cancers as a result of either hypermethylation of the 14-3-3σ promoter or induction of an oestrogen-responsive ubiquitin ligase that specifically targets 14-3-3σ for proteasomal degradation1,2,3,4,5,6,7,8,9. Loss of 14-3-3σ protein occurs not only within the tumours themselves but also in the surrounding pre-dysplastic tissue (so-called field cancerization), indicating that 14-3-3σ might have an important tumour suppressor function that becomes lost early in the process of tumour evolution3,9. The molecular basis for the tumour suppressor function of 14-3-3σ is unknown. Here we report a previously unknown function for 14-3-3σ as a regulator of mitotic translation through its direct mitosis-specific binding to a variety of translation/Initiation Factors, including eukaryotic Initiation Factor 4B in a stoichiometric manner. Cells lacking 14-3-3σ, in marked contrast to normal cells, cannot suppress cap-dependent translation and do not stimulate cap-independent translation during and immediately after mitosis. This defective switch in the mechanism of translation results in reduced mitotic-specific expression of the endogenous internal ribosomal entry site (IRES)-dependent form of the cyclin-dependent kinase Cdk11 (p58 PITSLRE), leading to impaired cytokinesis, loss of Polo-like kinase-1 at the midbody, and the accumulation of binucleate cells. The aberrant mitotic phenotype of 14-3-3σ-depleted cells can be rescued by forced expression of p58 PITSLRE or by extinguishing cap-dependent translation and increasing cap-independent translation during mitosis by using rapamycin. Our findings show how aberrant mitotic translation in the absence of 14-3-3σ impairs mitotic exit to generate binucleate cells and provides a potential explanation of how 14-3-3σ-deficient cells may progress on the path to aneuploidy and tumorigenesis.
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Phosphorylation of eucaryotic translation Initiation Factor 4B Ser422 is modulated by S6 kinases.
The EMBO journal, 2004Co-Authors: Brian Raught, David Shahbazian, Nahum Sonenberg, Franck Peiretti, Anne-claude Gingras, Mark Livingstone, Greg L. Mayeur, Roberto D. Polakiewicz, John W.b. HersheyAbstract:The eucaryotic translation Initiation Factor 4B (eIF4B) stimulates the helicase activity of the DEAD box protein eIF4A to unwind inhibitory secondary structure in the 5′ untranslated region of eucaryotic mRNAs. Here, using phosphopeptide mapping and a phosphospecific antiserum, we identify a serum-responsive eIF4B phosphorylation site, Ser422, located in an RNA-binding region required for eIF4A helicase-promoting activity. Ser422 phosphorylation appears to be regulated by the S6Ks: (a) Ser422 phosphorylation is sensitive to pharmacological inhibitors of phosphoinositide-3 kinase and the mammalian target of rapamycin; (b) S6K1/S6K2 specifically phosphorylate Ser422 in vitro; and (c) rapamycin-resistant S6Ks confer rapamycin resistance upon Ser422 phosphorylation in vivo. Substitution of Ser422 with Ala results in a loss of activity in an in vivo translation assay, indicating that phosphorylation of this site plays an important role in eIF4B function. We therefore propose that eIF4B may mediate some of the effects of the S6Ks on translation.
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In vitro RNA selection identifies RNA ligands that specifically bind to eukaryotic translation Initiation Factor 4B: the role of the RNA remotif.
RNA (New York N.Y.), 1996Co-Authors: Nathalie Méthot, G Pickett, Jack D. Keene, Nahum SonenbergAbstract:Translation Initiation Factor elF-4B is an RNA-binding protein that promotes the association of the mRNA to the 40S ribosomal subunit. One of its better characterized features is the ability to stimulate the activity of the DEAD box RNA hilicase elF-4A. In addition to an RNA recognition motif (RRM) located near its amino-terimus, elF-4B contains an RNA-binding region in its carboxy-terminal half. The elF-4A helicase stimulatory activity resides in the carboxy-terminal half of elF-4B, and the RRM has little impact on this function.To better understand the role of the elF-4B RRM, it was of interest to identify its specific RNA target sequence. To this end, it vitro RNA selection/amplifications were performed using various portions of elF-4B. These experiments were designed to test the RNA recognition specificity of the two elF-4B regions implicated in RNA binding and to assess the influence of elF-4A on the RNA-binding specificity. The RRM was shown to bind with high affinity to an RNA stem-loop structure with conserved primary sequence elements. Discrete point mutations in an in vitro-selected RNA identified residues critical for RNA binding. Neither the carboxy-terminal RNA-interaction region, nor elF-4A, influenced the structure of the high-affinity RNA ligands selected by elF-4B, and elF-4A by itself did not select any specific RNA target. Previous studies have demonstrated an interaction of elF-4B with ribosomes, and it was suggested that this association is mediated through binding to ribosomal RNA. We show that the RRM of elF-4B interacts directly with 18S rRNA and this interaction is inhibited by an excess of the elF-4B in vitro-selected RNA. ElF-4B could bind simultaneously to two different RNA molecules, supporting a model whereby elF-4B promotes ribosome binding to the 5 untranslated region of a mRNA by bridging it to 18S rRNA.
Daniel R. Gallie - One of the best experts on this subject based on the ideXlab platform.
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Eukaryotic Initiation Factor 4B and the poly(A)-binding protein bind eIF4G competitively
Translation (Austin Tex.), 2013Co-Authors: Shijun Cheng, Daniel R. GallieAbstract:The eukaryotic translation Initiation Factor (eIF) 4G functions as a scaffold protein that assembles components of the translation Initiation complex required to recruit the 40S ribosomal subunit to an mRNA. Although many eukaryotes express two highly similar eIF4G isoforms, those in plants are highly divergent in size and sequence from one another and are referred to as eIF4G and eIFiso4G. Although the domain organization of eIFiso4G differs substantially from eIF4G orthologs in other species, the domain organization of plant eIF4G is largely unknown despite the fact that it is more similar in size and sequence to eIF4G of other eukaryotes. In this study, we show that eIF4G differs from eIFiso4G in that it contains two distinct interaction domains for the poly(A) binding protein (PABP) and eIF4B but is similar to eIFiso4G in having two eIF4A interaction domains. PABP and eIF4B bind the same N-terminal region of eIF4G as they do to a region C-proximal to the HEAT-1 domain in the middle domain of eIF4G, resulting in competitive binding between eIF4B and PABP to each site. eIF4G also differs from eIFiso4G in that no competitive binding was observed between PABP and eIF4A or between eIF4B and eIF4A to its HEAT-1-containing region. These results demonstrate that despite substantial differences in size, sequence, and domain organization, PABP and eIF4B bind to eIF4G and eIFiso4G competitively.
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Translation Initiation Factor 4B Homodimerization, RNA Binding, and Interaction with Poly(A)-binding Protein Are Enhanced by Zinc
The Journal of biological chemistry, 2008Co-Authors: Shijun Cheng, Dixie J. Goss, Shemaila Sultana, Daniel R. GallieAbstract:Abstract The eukaryotic translation Initiation Factor (eIF) 4B promotes the RNA-dependent ATP hydrolysis activity and ATP-dependent RNA helicase activity of eIF4A and eIF4F during translation Initiation. eIF4B also helps to organize the assembly of the translational machinery through its interactions with eIF4A, eIF4G, eIF3, the poly(A)-binding protein (PABP), and RNA. Although the function of eIF4B is conserved among plants, animals, and yeast, eIF4B is one of the least conserved of Initiation Factors at the sequence level. Mammalian eIF4B is a constitutive dimer; however, conflicting reports have suggested that plant eIF4B may exist as a monomer or a dimer. In this study, we show that eIF4B from wheat can form a dimer and we identify the region responsible for its dimerization. Zinc stimulated homodimerization of eIF4B and bound eIF4B with a Kd of 19.7 nm. Zinc increased the activity of the eIF4B C-terminal RNA-binding domain specifically. Zinc promoted the interaction between eIF4B and PABP but not the interaction between eIF4B and eIF4A or eIFiso4G, demonstrating that the effect of zinc was highly specific. The interaction between PABP and eIFiso4G was also stimulated by zinc but required significantly higher levels of zinc. Interestingly zinc abolished the ability of eIFiso4G to compete with eIF4B in binding to their overlapping binding sites in PABP by preferentially promoting the interaction between eIF4B and PABP. Our observations suggest that wheat eIF4B can dimerize but requires zinc. Moreover zinc controls the partner protein selection of PABP such that the interaction with eIF4B is preferred over eIFiso4G.
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Wheat Eukaryotic Initiation Factor 4B Organizes Assembly of RNA and eIFiso4G, eIF4A, and Poly(A)-binding Protein
The Journal of biological chemistry, 2006Co-Authors: Shijun Cheng, Daniel R. GallieAbstract:The eukaryotic translation Initiation Factor (eIF) 4B promotes the RNA-dependent ATP hydrolysis activity and ATP-dependent RNA helicase activity of eIF4A and eIF4F during translation Initiation. Although this function is conserved among plants, animals, and yeast, eIF4B is one of the least conserved of Initiation Factors at the sequence level. To gain insight into its functional conservation, the organization of the functional domains of eIF4B from wheat has been investigated. Plant eIF4B contains three RNA binding domains, one more than reported for mammalian or yeast eIF4B, and each domain exhibits a preference for purine-rich RNA. In addition to a conserved RNA recognition motif and a C-terminal RNA binding domain, wheat eIF4B contains a novel N-terminal RNA binding domain that requires a short, lysine-rich containing sequence. Both the lysine-rich motif and an adjacent, C-proximal motif are conserved with an N-proximal sequence in human and yeast eIF4B. The C-proximal motif within the N-terminal RNA binding domain in wheat eIF4B is required for interaction with eIFiso4G, an interaction not reported for other eIF4B proteins. Moreover, each RNA binding domain requires dimerization for binding activity. Two binding sites for the poly(A)-binding protein were mapped to a region within each of two conserved 41-amino acid repeat domains on either side of the C-terminal RNA binding domain. eIF4A bound to an adjacent region within each repeat, supporting a central role for these conserved eIF4B domains in facilitating interaction with other components of the translational machinery. These results support the notion that eIF4B functions by organizing multiple components of the translation Initiation machinery and RNA.