Translation control of gene expression.
ABSTRACT The bacteriophage lambda cIII gene product is an early regulator of the lysogenic pathway. The availability of a set of cIII expression mutants allowed us to establish the structure-function relationship of the cIII mRNA. We demonstrated, using defined in vitro systems, that the cIII mRNA is present in two conformations at equilibrium. Mutations that have been shown to lead to cIII overexpression were found to freeze the RNA in one conformation (structure B), and permit efficient binding to the 30S ribosomal subunit. Mutations that have been shown to prevent cIII translation cause the mRNA to assume the alternative conformation (structure A). In this structure, the translation initiation region is occluded, thereby preventing 30S ribosomal subunit binding. Translation of the cIII gene is regulated by RNaseIII. We have localized the RNaseIII responsive element (RRE) to the cIII coding region. We suggest that the regulation of the equilibrium between the two mRNA conformations provides a mechanism for the control of cIII gene expression. The way in which RNaseIII participates in this regulation is as yet unknown.
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ABSTRACT: Memory consolidation is defined temporally based on pharmacological interventions such as inhibitors of mRNA translation (molecular consolidation) or post-acquisition deactivation of specific brain regions (systems level consolidation). However, the relationship between molecular and systems consolidation are poorly understood. Molecular consolidation mechanisms involved in translation initiation and elongation have previously been studied in the cortex using taste-learning paradigms. For example, the levels of phosphorylation of eukaryotic elongation factor 2 (eEF2) were found to be correlated with taste learning in the gustatory cortex (GC), minutes following learning. In order to isolate the role of the eEF2 phosphorylation state at Thr-56 in both molecular and system consolidation, we analyzed cortical-dependent taste learning in eEF2K (the only known kinase for eEF2) ki mice, which exhibit reduced levels of eEF2 phosphorylation but normal levels of eEF2 and eEF2K. These mice exhibit clear attenuation of cortical-dependent associative, but not of incidental, taste learning. In order to gain a better understanding of the underlying mechanisms, we compared brain activity as measured by MEMRI (manganese-enhanced magnetic resonance imaging) between eEF2K ki mice and WT mice during conditioned taste aversion (CTA) learning and observed clear differences between the two but saw no differences under basal conditions. Our results demonstrate that adequate levels of phosphorylation of eEF2 are essential for cortical-dependent associative learning and suggest that malfunction of memory processing at the systems level underlies this associative memory impairment.Learning & memory (Cold Spring Harbor, N.Y.) 02/2012; 19(3):116-25. DOI:10.1101/lm.023937.111 · 4.38 Impact Factor
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ABSTRACT: Translation is the vital process by which the information contained in messenger RNAs (mRNA) is used to synthesise proteins. This is a highly regulated process that involves a complex machinery and tight control at every step. In eukaryotes, translation initiation is the rate-limiting step and the most tightly controlled. The translation of an mRNA is preceded by multiple post-transcription steps including, splicing, export and chemical modification including the addition of a 5'-end cap structure. The initiation factor eIF4E binds to this cap structure while in the nucleus, and initiates the translation by recruiting the ribosome after export to the cytoplasm. Controlling the availability of eIF4E within the various compartments of the cell has a direct effect on the efficiency of translation initiation and indirectly on the rate of proliferation of the cell. On the other hand, disturbing the level of eIF4E synthesis could lead to various pathologies especially tumour development. eIF4E requires multiple binding partners to fulfill its mission. The factor 4E-T is involved in eIF4E transport, while eIF4G enhances the binding of eIF4E to the cap structure. eIF4E can be inactivated upon sequestration by the 4E Binding Protein (4E-BP) who competes with eIF4G for binding of eIF4E. The localization of eIF4E inside the cell is clearly critical for its normal function. It is known that many external stresses can influence cellular translation and this prompted us to investigate the effect of such stresses on the cellular localization of eIF4E and to study the role of eIF4E-binding partners under these conditions. The present work demonstrates that, during heat shock and oxidative stress, 4E-BP plays an essential role in controlling the localization of eIF4E to the stress response cytoplasmic foci, known as the stress granules (SGs). In addition, eIF4E is partially retained in the nucleus during heat shock but not in oxidative stress. These observations suggest that upon stress the cellular translation mechanism is delayed or even stopped by reducing the availability of eIF4E to the translation complex. On the other hand, we show that eIF4E nuclear translocation upon poliovirus infection is correlated with eIF4G cleavage. This translocation could favour the shutdown of host cell protein synthesis by reducing the cap dependent translation and preventing mRNA circularization. In our study, we focused on the role of eIF4E as a key player for cellular survival under stressful conditions. Therefore, identifying reagents that induce the relocalization of eIF4E to the nucleus or to SGs could help in the development of anti-proliferative drugs. La traduction est un processus vital par lequel l'information contenue dans l'ARN messager est utilisée pour la synthèse protéique. Chaque étape de ce procédé est strictement régulé grâce à l'implication d'une machinerie complexe hautement contrôlée. Chez les eucaryotes, l'initiation est l'étape limitante et la mieux contrôlée du processus de traduction. Surviennent ensuite, les étapes dites post-traductionnelles qui incluent, l'épissage, l'exportation nucléaire et les modifications biochimiques. Le facteur de transcription eIF4E lie la coiffe du messager en 5' au niveau du noyau, permettant son exportation au cytoplasme et l'initiation de la traduction grâce au recrutement des sous-unites ribosomales. Le contrôle d'eIF4E dans les différentes parties cellulaires est crucial pour l'efficacité traductionnelle, mais génère également un effet indirect sur la prolifération et division cellulaire. Aussi, un défaut de synthèse ou de niveau d'expression d'eIF4E amènent de nombreuses anomalies, spécifiquement lors du développement tumoral. eIF4E requiert de nombreux partenaires pour agir efficacement. Le facteur 4E-T est impliqué dans le transport d'eIF4E tandis que eIF4G favorise sa liaison a la coiffe. eIF4E est connu pour être inactive par séquestration de 4E-BP, principal compétiteur pour la liaison a la coiffe. La localisation d'eIF4E au sein de la cellule est critique pour son bon fonctionnement. De nombreux facteurs de stress sont connus pour influencer la traduction cellulaire. Je me suis donc concentre à étudier les effets des facteurs de stress dans la localisation cellulaire d'eIF4E et le rôle respectif de chacun de ses partenaires dans ces conditions. J'ai ainsi démontré que durant un choc de chaleur et un stress oxydatif, 4E-BP joue un rôle essentiel dans le contrôle de la localisation d'eIF4E au niveau des granules de stress. De plus, eIF4E est partiellement retenu au noyau lors d'un stress de chaleur mais pas au cours d'un stress oxydatif. Ces observations suggèrent que lors d'un stress, la machinerie traductionnelle est retardée ou arrêtée par une baisse de la disponibilité d'eIF4E a ce complexe protéique. D'un autre coté, j'ai montré que la translocation nucléaire d'eIF4E au cours d'une infection au Poliovirus est corrélée avec le clivage d'eIF4G. Cette translocation pourrait favoriser l'arrêt de la synthèse protéique de la cellule hote en réduisant la traduction dépendant de la coiffe et en prévenant la circularisation du messager. En conclusion, eIF4E est un élément clé dans la survie cellulaire associée aux conditions de stress. De plus, l'identification de réactifs pouvant induire la relocalisation d'eIF4E au noyau ou aux granules de stress aiderait au développement de composes anti-prolifératifs.
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ABSTRACT: A number of bacterial and viral genes take part in the decision between lysis and lysogenization in temperate bacteriophages. In the lambda case, at least five viral genes (cI, cro, cII, N and cIII) and several bacterial genes are involved. Several attempts have been made to model this complex regulatory network. Our approach is based on a logical method described in the first paper of the series which formalizes the interactions between the elements of a regulatory network in terms of discrete variables, functions and parameters. In this paper two models are described and discussed, the first (two-variable model) focused on cI and cro interactions, the second (four-variable model) considering, in addition, genes cII and N. The treatment presented emphasizes the roles of positive and negative feedback loops and their interactions in the development of the phage. The role of the loops between cI and cro, and of cI on itself (which both have to be positive loops) was discovered earlier; this group's contribution to this aspect mainly deals with the possibility of treating these loops as parts of a more extended network. In contrast, the role of the negative loop of cro on itself had apparently remained unexplained. We realized that this loop buffers the expression of genes cro itself. cII, O and P against the inflation due to the rapid replication of the phage. More generally, negative auto-control of a gene appears an efficient way to render its expression insensitive (or less sensitive) to gene dosage, whereas a simple negative control would not provide this result.Bulletin of Mathematical Biology 04/1995; 57(2):277-97. DOI:10.1016/0092-8240(94)00037-D · 1.29 Impact Factor