Genes driving potato tuber initiation and growth: identification based on transcriptional changes using the POCI array

The increasing amount of available expressed gene sequence data makes whole-transcriptome analysis of certain crop species possible. Potato currently has the second largest number of publicly available expressed sequence tag (EST) sequences among the Solanaceae. Most of these ESTs, plus other proprietary sequences, were combined and used to generate a unigene assembly. The set of 246,182 sequences produced 46,345 unigenes, which were used to design a 44K 60-mer oligo array (Potato Oligo Chip Initiative: POCI). In this study, we attempt to identify genes controlling and driving the process of tuber initiation and growth by implementing large-scale transcriptional changes using the newly developed POCI array. Major gene expression profiles could be identified exhibiting differential expression at key developmental stages. These profiles were associated with functional roles in cell division and growth. A subset of genes involved in the regulation of the cell cycle, based on their Gene Ontology classification, exhibit a clear transient upregulation at tuber onset indicating increased cell division during these stages. The POCI array allows the study of potato gene expression on a much broader level than previously possible and will greatly enhance analysis of transcriptional control mechanisms in a wide range of potato research areas. POCI sequence and annotation data are publicly available through the POCI database (http://pgrc.ipk-gatersleben.de/poci).

Ice-stream initiation, duration and thinning on James Ross Island, northern Antarctic Peninsula

Predicting the future response of the Antarctic Ice Sheet to climate change requires an understanding of the ice streams that dominate its dynamics. Here we use cosmogenic isotope exposure-age dating (26Al, 10Be and 36Cl) of erratic boulders on ice-free land on James Ross Island, north-eastern Antarctic Peninsula, to define the evolution of Last Glacial Maximum (LGM) ice in the adjacent Prince Gustav Channel. These data include ice-sheet extent, thickness and dynamical behaviour. Prior to ∼18 ka, the LGM Antarctic Peninsula Ice Sheet extended to the continental shelf-edge and transported erratic boulders onto high-elevation mesas on James Ross Island. After ∼18 ka there was a period of rapid ice-sheet surface-lowering, coincident with the initiation of the Prince Gustav Ice Stream. This timing coincided with rapid increases in atmospheric temperature and eustatic sea-level rise around the Antarctic Peninsula. Collectively, these data provide evidence for a transition from a thick, cold-based LGM Antarctic Peninsula Ice Sheet to a thinner, partially warm-based ice sheet during deglaciation.

Stabilization of the i-motif structure by the intra-strand cross-link formation

The i-motif structures are formed by oligonucleotides containing cytosine tracts under acidic conditions. The folding of the i-motif under physiological conditions is of great interest because of its biological role. In this study, we investigated the effect of the intra-strand cross-link on the stability of the i-motif structure. The 4-vinyl-substituted analog of thymidine (T-vinyl) was incorporated into the 5′-end of the human telomere complementary strand, which formed the intra-strand cross-link with the internal adenine. The intra-strand cross-linked i-motif displayed CD spectra similar to that of the natural i-motif at acidic pH, which was transformed into a random coil with the increasing pH. The pH midpoint for the transition from the i-motif to random coil increased from pH 6.1 for the natural one to pH 6.8 for the cross-linked one. The thermodynamic parameters were obtained by measuring the thermal melting behaviors by CD and UV, and it was determined that the intra-strand cross-linked i-motif is stabilized due to a favorable entropy effect. Thus, this study has clearly indicated the validity of the intra-strand cross-linking for stabilization of the i-motif structure.

The novel Syk inhibitor R406 reveals mechanistic differences in the initiation of GPVI and CLEC-2 signaling in platelets.

The inhibitory effect of R406 provides direct evidence of a role for Syk in GPVI, CLEC-2 and integrin alphaIIbbeta3 signaling in human platelets. Further, the results demonstrate a critical role for Syk in mediating tyrosine phosphorylation of CLEC-2, suggesting a novel model in which both Src and Syk kinases regulate tyrosine phosphorylation of the C-type lectin receptor leading to platelet activation.

Sea-breeze dynamics and convection initiation: the influence of convective parameterization in weather and climate model biases

There are some long-established biases in atmospheric models that originate from the representation of tropical convection. Previously, it has been difficult to separate cause and effect because errors are often the result of a number of interacting biases. Recently, researchers have gained the ability to run multiyear global climate model simulations with grid spacings small enough to switch the convective parameterization off, which permits the convection to develop explicitly. There are clear improvements to the initiation of convective storms and the diurnal cycle of rainfall in the convection-permitting simulations, which enables a new process-study approach to model bias identification. In this study, multiyear global atmosphere-only climate simulations with and without convective parameterization are undertaken with the Met Office Unified Model and are analyzed over the Maritime Continent region, where convergence from sea-breeze circulations is key for convection initiation. The analysis shows that, although the simulation with parameterized convection is able to reproduce the key rain-forming sea-breeze circulation, the parameterization is not able to respond realistically to the circulation. A feedback of errors also occurs: the convective parameterization causes rain to fall in the early morning, which cools and wets the boundary layer, reducing the land–sea temperature contrast and weakening the sea breeze. This is, however, an effect of the convective bias, rather than a cause of it. Improvements to how and when convection schemes trigger convection will improve both the timing and location of tropical rainfall and representation of sea-breeze circulations.

Insights on eukaryotic translation initiation factor 5A (eIF5A) in the brain and aging

Long-term memory, a persistent form of synaptic plasticity, requires translation of a subset of mRNA present in neuronal dendrites during a short and critical period through a mechanism not yet fully elucidated. Western blotting analysis revealed a high content of eukaryotic translation initiation factor 5A (eIF5A) in the brain of neonatal rats, a period of intense neurogenesis rate, differentiation and synaptic establishment, when compared to adult rats. Immunohistochemistry analysis revealed that eIF5A is present in the whole brain of adult rats showing a variable content among the cells from different areas (e.g. cortex, hippocampus and cerebellum). A high content of eIF5A in the soma and dendrites of Purkinje cells, key neurons in the control of motor long-term memory in the cerebellum, was observed. Detection of high eIF5A content was revealed in dendritic varicosities of Purkinje cells. Evidence is presented herein that a reduction of eIF5A content is associated to brain aging. (C) 2008 Elsevier B.V. All rights reserved.

HOST-MEDIATED INDUCTION OF alpha-AMYLASES BY LARVAE OF THE MEXICAN BEAN WEEVIL Zabrotes subfasciatus (COLEOPTERA: CHRYSOMELIDAE: BRUCHINAE) IS IRREVERSIBLE AND OBSERVED FROM THE INITIATION OF THE FEEDING PERIOD

Larvae of Zabrotes subfasciatus secrete alpha-amylases that are insensitive to the alpha-amylase inhibitor found in seeds of Phaseolus vulgaris. By analyzing amylase activities during larval development on P. vulgaris, we detected activity of the constitutive amylase and the two inducible amylase isoforms at all stages. When larvae were transferred from the non alpha-amylase inhibitor containing seeds of Vigna unguiculata to P. vulgaris, the inducible alpha-amylases were expressed at the same level as in control larvae fed on P. vulgaris. Interestingly, when larvae were transferred from seeds of P. vulgaris to those of V. unguiculata, inducible alpha-amylases continued to be expressed at a level similar to that found in control larvae fed P. vulgaris continuously. When 10-day-old larvae were removed from seeds of V. unguiculata and transferred into capsules containing flour of P. vulgaris cotyledons, and thus maintained until completing 17 days ( age when the larvae stopped feeding), we could detect higher activity of the inducible alpha-amylases. However, when larvae of the same age were transferred from P. vulgaris into capsules containing flour of V. unguiculata, the inducible alpha-amylases remained up-regulated. These results suggest that the larvae of Z. subfasciatus have the ability to induce insensitive amylases early in their development. A short period of feeding on P. vulgaris cotyledon flour was sufficient to irreversibly induce the inducible alpha-amylase isoforms. Incubations of brush border membrane vesicles with the alpha-amylase inhibitor 1 from P. vulgaris suggest that the inhibitor is recognized by putative receptors found in the midgut microvillar membranes. (C) 2010 Wiley Periodicals, Inc.

Involvement of Eukaryotic Translation Initiation Factor 5A (eIF5A) in Skeletal Muscle Stem Cell Differentiation

The eukaryotic translation initiation factor 5A (eIF5A) contains a special amino acid residue named hypusine that is required for its activity, being produced by a post-translational modification using spermidine as substrate. Stem cells from rat skeletal muscles (satellite cells) were submitted to differentiation and an increase of eIF5A gene expression was observed. Higher content of eIF5A protein was found in satellite cells on differentiation in comparison to non-differentiated satellite cells and skeletal muscle. The treatment with NI-guanyl- 1,7-diaminoheptane (GC7), a hypusination inhibitor, reversibly abolished the differentiation process. In association with the differentiation blockage, an increase of glucose consumption and lactate production and a decrease of glucose and palmitic acid oxidation were observed. A reduction in cell proliferation and protein synthesis was also observed. L-Arginine, a spermidine precursor and partial suppressor of muscle dystrophic phenotype, partially abolished the GC7 inhibitory effect on satellite cell differentiation. These results reveal a new physiological role for eIF5A and contribute to elucidate the molecular mechanisms involved in muscle regeneration.

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.

DNA strand breaks and base modifications induced by cholesterol hydroperoxides

Cholesterol (Ch) can be oxidized by reactive oxygen species, forming oxidized products such as Ch hydroperoxides (ChOOH). These hydroperoxides can disseminate the peroxidative stress to other cell compartments. In this work, the ability of ChOOH to induce strand breaks and/or base modifications in a plasmid DNA model was evaluated. In addition, HPLC/MS/MS analyses were performed to investigate the formation of 8-oxo-7,8-dihydro-2`-deoxyguanosine (8-oxodGuo) after the incubation of 2`-deoxyguanosine (dGuo) with ChOOH and Cu(2+). In the presence of copper ions, ChOOH induced DNA strand breaks in time and concentration-dependent manners. Purine and pyrimidine base modifications were also observed, as assessed respectively by the treatment with Fpg and Endo III repair enzymes. The detection of 8-oxodGuo by HPLC/MS/MS is in agreement with the dGuo oxidation in plasmid DNA. ChOOH-derived DNA damage adds further support to the role of lipid peroxidation in inducing DNA modifications and mutation.

Double-strand DNA cleavage induced by oxindole-Schiff base copper(II) complexes with potential antitumor activity

Some oxindole-Schiff base copper(II) complexes have already shown potential antitumor activity towards different cells, inducing apoptosis in a process modulated by the ligand, and having nuclei and mitochondria as main targets. Here, three novel copper(II) complexes with analogous ligands were isolated and characterized by spectroscopic techniques, having their reactivity compared to the so far most active complex in this class. Cytotoxicity experiments carried out toward human neuroblastoma SH-SY5Y cells confirmed its proapoptosis property. DNA cleavage studies were then performed in the presence of these complexes, in order to verify the influence of ligand structural features in its nuclease activity. All of them were able to cause double-strand DNA scissions, giving rise to nicked circular Form II and linear Form III species, in the presence of hydrogen peroxide. Additionally, DNA Form II was also detected in the absence of peroxide when the most active complex, [Cu(isaepy)(2)](2+) 1, was used. In an effort to better elucidate their interactions with DNA, solutions of the different complexes titrated with DNA had their absorption spectra monitored. An absorbance hyperchromism observed at 260 nm pointed to the intercalation of these complexes into the DNA structure. Further, investigations of 2-deoxy-D-ribose (DR) oxidation catalyzed by each of those complexes, using 2-thiobarbituric acid reactive species (TBARS) method, and detection of reactive oxygen species (ROS) formation by spin-trapping EPR, suggested that their mechanism of action in performing efficiently DNA cleavage occurs preferentially, but not only by oxidative pathways. (C) 2007 Elsevier Inc. All rights reserved.