Synthetic lethality between eIF5A and Ypt1 reveals a connection between translation and the secretory pathway in yeast

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

eIF5A has a function in the elongation step of translation in yeast

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Structural modeling and mutational analysis of yeast eukaryotic translation initiation factor 5A reveal new critical residues and reinforce its involvement in protein synthesis

Eukaryotic translation initiation factor 5A (eIF5A) is a protein that is highly conserved and essential for cell viability. This factor is the only protein known to contain the unique and essential amino acid residue hypusine. This work focused on the structural and functional characterization of Saccharomyces cerevisiae eIF5A. The tertiary structure of yeast eIF5A was modeled based on the structure of its Leishmania mexicana homologue and this model was used to predict the structural localization of new site-directed and randomly generated mutations. Most of the 40 new mutants exhibited phenotypes that resulted from eIF-5A protein-folding defects. Our data provided evidence that the C-terminal alpha-helix present in yeast eIF5A is an essential structural element, whereas the eIF5A N-terminal 10 amino acid extension not present in archaeal eIF5A homologs, is not. Moreover, the mutants containing substitutions at or in the vicinity of the hypusine modification site displayed nonviable or temperature-sensitive phenotypes and were defective in hypusine modification. Interestingly, two of the temperature-sensitive strains produced stable mutant eIF5A proteins - eIF5A(K56A) and eIF5A(Q22H,L93F)- and showed defects in protein synthesis at the restrictive temperature. Our data revealed important structural features of eIF5A that are required for its vital role in cell viability and underscored an essential function of eIF5A in the translation step of gene expression.

Is there a role for eIF5A in translation?

The putative translation factor eIF5A is essential for cell viability and is highly conserved from archaebacteria to mammals. This factor is the only cellular protein that undergoes an essential posttranslational modification dependent on the polyamine spermidine, called hypusination. This review focuses on the functional characterization of eIF5A. Although this protein was originally identified as a translation initiation factor, subsequent studies did not support a role for eIF5A in general translation initiation. eIF5A has also been implicated in nuclear export of HIV-1 Rev and mRNA decay, but these findings are controversial in the literature and may reflect secondary effects of eIF-5A function. Next, the involvement of eIF5A and hypusination in the control of the cell cycle and proliferation in various organisms is reviewed. Finally, recent evidence in favor of reconsidering the role of eIF5A as a translation factor is discussed. Future studies may reveal the specific mechanism by which eIF5A affects protein synthesis.

eIF5A binds to translational machinery components and affects translation in yeast

The putative translation factor eIF5A is essential for cell viability and is highly conserved from archebacteria to mammals. Although this protein was originally identified as a translation initiation factor, subsequent experiments did not support a role for eIF5A in general translation. In this work, we demonstrate that eIF-5A interacts with structural components of the 80S ribosome, as well as with the translation elongation factor 2 (eEF2). Moreover, eIF5A is further shown to cofractionate with monosomes in a translation-dependent manner. Finally, eIF5A mutants show altered polysome profiles and are sensitive to translation inhibitors. Our results re-establish a function for eIF5A in translation and suggest a role for this factor in translation elongation instead of translation initiation. (c) 2006 Elsevier B.V. All rights reserved.

Case study: a 3D resistivity and induced polarization imaging from downstream a waste disposal site in Brazil

A contaminated site from a downstream municipal solid waste disposal site in Brazil was investigated by using a 3D resistivity and induced polarization (IP) imaging technique. This investigation purpose was to detect and delineate contamination plume produced by wastes. The area was selected based on previous geophysical investigations, and chemical analyses carried out in the site, indicating the presence of a contamination plume in the area. Resistivity model has successfully imaged waste presence (rho < 20 Omega m), water table depth, and groundwater flow direction. A conductive anomaly (rho < 20 Omega m) outside wastes placement was interpreted as a contamination plume. Chargeability model was also able to imaging waste presence (m > 31 mV/V), water table depth, and groundwater flow direction. A higher chargeability zone (m > 31 mV/V) outside wastes placement and following conductive anomaly was interpreted as a contamination plume. Normalized chargeability (MN = m/rho) confirmed polarizable zone, which could be an effect of a salinity increase (contamination plume), and the clay presence in the environment.

The fate of copepod populations in the Paranapanema River (São Paulo, Brazil), downstream from the Jurumirim dam

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Phraseological translation

Almost all texts contain some complex lexical units, belonging to the phraseology of the language of a specialized field or of the general language. The translator must first identify this phraseologism, and then understand its meaning. However, it is not enough to propose an explanation in the target language: the translator has to establish its phraseologically equivalent lexical unit in meaning and in phraseological formulations.

Annexin-A1 gene expression during liver development and post-translation modification after experimental endotoxemia

Objective and design: We have previously reported a role for annexin-A1 in liver proliferation and tumorogenicity as well as its action as an acute phase protein in a model of endotoxemia in interleukin-6 null mice.Material and methods: In this study, we have investigated the analysis of the gene and protein expression in annexin-A1 null mice and the wild type livers during foetal and adult life, and in the presence of a proinflammatory stimulus.Results: The data indicate a link between the expression of the annexin-A1 as serine-phosphorylated-protein during early events of the inflammatory response and as tyrosine-phosphorylated-form at later time-points, during the resolution of inflammation.Conclusions: The study of annexin-A1 post-translation modification may promote a new annexin-A1 peptide discovery programme to treat specific pathologies.

Effect of atypical rainfall on lotic zooplankton: comparing downstream of a reservoir and tributaries with free stretches

To contribute to the knowledge of lotic zooplankton from dammed and free stretches, we tested the hypothesis that the ecological attributes are different between these environments, which are subjected to several effects caused by different hydrological conditions. The study was carried out in the low Iguacu river, a large hydrographic basin in the south of Brasil. Two samplings were performed, one in the dry period (April/04) and the other in the atypical rainy period (July/04), in five stations downstream a large reservoir, and other 12 stations in four tributaries representing the upper, intermediate and low regions of each river. The observations suggest a clear spatial distribution of zooplankton in lotic stretches subjected or not to damming, mainly due to the effects of physical, chemical and biological variables. Furthermore, the atypical rainfall promoted alterations in community structure when compared to the dry period.

Conserved sequences in the β subunit of archaeal and eukaryal translation initiation factor 2 (elF2), absent from elF5, mediate interaction with elF2γ

The eukaryotic translation initiation factor 2 (eIF2) binds the methionyl-initiator tRNA in a GTP-dependent mode. This complex associates with the 40 S ribosomal particle, which then, with the aid of other factors, binds to the 5' end of the mRNA and migrates to the first AUG codon, where eIF5 promotes GTP hydrolysis, followed by the formation of the 80 S ribosome. Here we provide a comparative sequence analysis of the β subunit of eIF2 and its archaeal counterpart (aIF2β). aIF2β differs from eIF2β in not possessing an N-terminal extension implicated in binding RNA, eIF5 and eIF2B. The remaining sequences are highly conserved, and are shared with eIF5. Previously isolated mutations in the yeast eIF2β, which allow initiation of translation at UUG codons due to the uncovering of an intrinsic GTPase activity in eIF2, involve residues that are conserved in aIF2β, but not in eIF5. We show that the sequence of eIF2B homologous to aIF2β is sufficient for binding eIF2γ, the only subunit with which it interacts, and comprises, at the most, 78 residues, eIF5 does not interact with eIF2γ, despite its similarity with eIF2β, probably because of a gap in homology in this region. These observations have implications for the evolution of the mechanism of translation initiation.

Genetic interactions of yeast eukaryotic translation initiation factor 5a (eIF5A) reveal connections to poly(A)-binding protein and protein kinase C signaling

The highly conserved eukaryotic translation initiation factor eIF5A has been proposed to have various roles in the cell, from translation to mRNA decay to nuclear protein export. To further our understanding of this essential protein, three temperature-sensitive alleles of the yeast TIF51A gene have been characterized. Two mutant eIF5A proteins contain mutations in a proline residue at the junction between the two eIFSA domains and the third, strongest allele encodes a protein with a single mutation in each domain, both of which are required for the growth defect. The stronger tif51A alleles cause defects in degradation of short-lived mRNAs, supporting a role for this protein in mRNA decay. A multicopy suppressor screen revealed six genes, the overexpression of which allows growth of a tif51A-1 strain at high temperature; these genes include PAB1, PKC1, and PKC1 regulators WSC1, WSC2, and WSC3. Further results suggest that eIFSA may also be involved in ribosomal synthesis and the WSC/PKC1 signaling pathway for cell wall integrity or related processes.

Inhibition of eukaryotic translation initiation factor 5A (eIF5A) hypusination impairs melanoma growth

The eukaryotic translation initiation factor 5A (eIF5A) undergoes a specific post-translational modification called hypusination. This modification is required for the functionality of this protein. The compound N1-guanyl-1,7-diaminoheptane (GC7) is a potent and selective inhibitor of deoxyhypusine synthase, which catalyses the first step of eIF5A hypusination process. In the present study, the effects of GC7 on cell death were investigated using two cell lines: melan-a murine melanocytes and Tm5 marine melanoma. In vitro treatment with GC7 increased by 3-fold the number of cells presenting DNA fragmentation in Tm5 cells. Exposure to GC7 also decreased viability to both cell lines. This study also describes, for the first time, the in vivo antitumour effect of GC7, as indicated by impaired melanoma growth in C57BL/6 mice. Copyright © 2006 John Wiley & Sons, Ltd.

Leucine-rich diet alters the eukaryotic translation initiation factors expression in skeletal muscle of tumour-bearing rats

Background: Cancer-cachexia induces a variety of metabolic disorders on protein turnorver, decreasing protein synthesis and increasing protein degradation. Controversly, insulin, other hormones, and branched-chain amino acids, especially leucine, stimulate protein synthesis and modulate the activity of translation initiation factors involved in protein synthesis. Since the tumour effects are more pronounced when associated with pregnancy, ehancing muscle-wasting proteolysis, in this study, the influence of a leucine-rich diet on the protein synthesis caused by cancer were investigated. Methods: Pregnant rats with or without Walker 256 tumour were distributed into six groups. During 20 days of experiment, three groups were fed with a control diet: C - pregnant control, W - tumour-bearing, and P - pair-fed, which received the same amount of food as ingested by the W group; three other groups of pregnant rats were fed a leucine-rich diet: L - pregnant leucine, WL - tumour-bearing, and PL - pair-fed, which received the same amount of food as ingested by the WL group. Results: The gastrocnemius muscle of WL rats showed increased incorporation of leucine in protein compared to W rats; the leucine-rich diet also prevented the decrease in plasma insulin normally seen in W. The expression of translation initiation factors increased when tumour-bearing rats fed leucine-rich diet, with increase of ∼35% for eIF2α and eIF5, ∼17% for eIF4E and 20% for eIF4G; the expression of protein kinase S6K1 and protein kinase C was also highly enhanced. Conclusion: The results suggest that a leucine-rich diet increased the protein synthesis in skeletal muscle in tumour-bearing rats possibly through the activation of eIF factors and/or the S6kinase pathway. © 2007 Ventrucci et al; licensee BioMed Central Ltd.

Discharge pulses of hydroelectric dams and their effects in the downstream limnological conditions: A case study in a large tropical river (SE Brazil)

To address daily fluctuations in electricity demands, the quantities of water passing through the turbines of hydropower plants can vary significantly (up to fourfold) during a 24-h cycle. This study evaluates the effects of hourly variations in water discharges on the limnological conditions observed in two below-dam river stretches. The study reservoirs, Capivara and Taquaruçu, are the 9th and 10th reservoirs in a cascade of dams in the Paranapanema River in south-east Brazil. The reservoirs exhibit different trophic conditions, water retention times, thermal regimes and spillway positions. Capivara Reservoir is deeper, meso-eutrophic, with a high water retention time and hypolimnetic discharges (32 m) varying between 500 and 1400 m3 s-1. In contrast, Taquaruçu Reservoir is relatively shallow, oligo-mesotrophic, and has a low retention time, with water discharges varying between 500 and 2000 m3 s-1. Its turbine water intake zone also is more superficial (7 m). For two periods of the year, winter and summer, profiles of limnological measurements were developed in the lacustrine (above-dam site) zones of the reservoirs, as well as in the downstream river stretches (below-dam site). In both cases, the sampling was carried out at 4-h intervals over a complete nictemeral cycle. The results demonstrated that the reservoir operating regime (water discharge variations) promoted significant differences in the conditions of the river below the dams, especially for water velocity, turbidity, and nutrient and suspended solids concentrations. The reservoir physical characteristics, including depth, thermal stratification and outlet structure, are also key factors influencing the limnology and water quality at the below-dam sampling sites. In the case of Capivara Reservoir, for example, the low dissolved oxygen concentration (<5.0 mg L-1) in its bottom water layer was transferred to the downstream river stretches during the summer. These study results demonstrated that it is important to continue such investigations as a means of verifying whether or not these high-amplitude/low-frequency variations could negatively affect the downstream river biota. © 2009 The Authors. Journal compilation © 2009 Blackwell Publishing Asia Pty Ltd.