Adaptation of an [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay to evaluate the cytotoxicity of the extracellular products of micro-organisms pathogenic to fish

AIMS: Adaptation of a colorimetric assay using [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] (MTT) to evaluate the cytotoxicity of the extracellular products of micro-organisms pathogenic to fish. METHODS AND RESULTS: The optimal conditions for the colorimetric assay were determined and this method was compared with the trypan blue exclusion assay. The protein concentration of extracellular products causing the death of 50% of the cell population (CI50) was determined. CONCLUSIONS: This assay enables quantitative and objective comparison of the cytotoxicity of the extracellular products of micro-organisms pathogenic to fish. It was shown to be more accurate than conventional counting with the trypan blue exclusion assay. SIGNIFICANCE AND IMPACT OF THE STUDY: This method may also be useful for characterizing the cytotoxicity of specific components of extracellular products.     

Genomic Hallmarks of Genes Involved in Chromosomal Translocations in Hematological Cancer

Reciprocal chromosomal translocations (RCTs) leading to the formation of fusion genes are important drivers of hematological cancers. Although the general requirements for breakage and fusion are fairly well understood, quantitative support for a general mechanism of RCT formation is still lacking. The aim of this paper is to analyze available high-throughput datasets with computational and robust statistical methods, in order to identify genomic hallmarks of translocation partner genes (TPGs). Our results show that fusion genes are generally overexpressed due to increased promoter activity of 5′ TPGs and to more stable 3′-UTR regions of 3′ TPGs. Furthermore, expression profiling of 5′ TPGs and of interaction partners of 3′ TPGs indicates that these features can help to explain tissue specificity of hematological translocations. Analysis of protein domains retained in fusion proteins shows that the co-occurrence of specific domain combinations is non-random and that distinct functional classes of fusion proteins tend to be associated with different components of the gene fusion network. This indicates that the configuration of fusion proteins plays an important role in determining which 5′ and 3′ TPGs will combine in specific fusion genes. It is generally accepted that chromosomal proximity in the nucleus can explain the specific pairing of 5′ and 3′ TPGS and the recurrence of hematological translocations. Using recently available data for chromosomal contact probabilities (Hi-C) we show that TPGs are preferentially located in early replicated regions and occupy distinct clusters in the nucleus. However, our data suggest that, in general, nuclear position of TPGs in hematological cancers explains neither TPG pairing nor clinical frequency. Taken together, our results support a model in which genomic features related to regulation of expression and replication timing determine the set of candidate genes more likely to be translocated in hematological tissues, with functional constraints being responsible for specific gene combinations.     

Synthesis and profiling of benzylmorpholine 1,2,4,5-tetraoxane analogue N205: Towards tetraoxane scaffolds with potential for single dose cure of malaria

A series of aryl carboxamide and benzylamino dispiro 1,2,4,5-tetraoxane analogues have been designed and synthesized in a short synthetic sequence from readily available starting materials. From this series of endoperoxides, molecules with in vitro IC50s versus Plasmodium falciparum (3D7) as low as 0.84?nM were identified. Based on an assessment of blood stability and in vitro microsomal stability, N205 (10a) was selected for rodent pharmacokinetic and in vivo antimalarial efficacy studies in the mouse Plasmodium berghei and Plasmodium falciparum Pf3D70087/N9 severe combined immunodeficiency (SCID) mouse models. The results indicate that the 4-benzylamino derivatives have excellent profiles with a representative of this series, N205, an excellent starting point for further lead optimization studies.     

On the path to extinction: Inbreeding and admixture in a declining grey wolf population

Allee effects reduce the viability of small populations in many different ways, which act synergistically to lead populations towards extinction vortexes. The Sierra Morena wolf population, isolated in the south of the Iberian Peninsula and composed of just one or few packs for decades, represents a good example of how diverse threats act additively in very small populations. We sequenced the genome of one of the last wolves identified (and road‐killed) in Sierra Morena and that of another wolf in the Iberian Wolf Captive Breeding Program and compared them with other wolf and dog genomes from around the world (including two previously published genome sequences from northern Iberian wolves). The results showed relatively low overall genetic diversity in Iberian wolves, but diverse population histories including past introgression of dog genes. The Sierra Morena wolf had an extraordinarily high level of inbreeding and long runs of homozygosity, resulting from the long isolation. In addition, about one‐third of the genome was of dog origin. Despite the introgression of dog genes, heterozygosity remained low because of continued inbreeding after several hybridization events. The results thus illustrate the case of a small and isolated wolf population where the low population density may have favoured hybridization and introgression of dog alleles, but continued inbreeding may have resulted in large chromosomal fragments of wolf origin completely disappearing from the population, and being replaced by chromosomal fragments of dog origin. The latest population surveys suggest that this population may have gone extinct.     

Glass‐Type Polyamorphism in Li‐Garnet Thin Film Solid State Battery Conductors

Ceramic Li₇La₃Zr₂O₁₂ garnet materials are promising candidates for the electrolytes in solid state batteries due to their high conductivity and structural stability. In this paper, the existence of “polyamorphism” leading to various glass‐type phases for Li‐garnet structure besides the known crystalline ceramic ones is demonstrated. A maximum in Li‐conductivity exists depending on a frozen thermodynamic glass state, as exemplified for thin film processing, for which the local near range order and bonding unit arrangement differ. Through processing temperature change, the crystallization and evolution through various amorphous and biphasic amorphous/crystalline phase states can be followed for constant Li‐total concentration up to fully crystalline nanostructures. These findings reveal that glass‐type thin film Li‐garnet conductors exist for which polyamorphism can be used to tune the Li‐conductivity being potential new solid state electrolyte phases to avoid Li‐dendrite formation (no grain boundaries) for future microbatteries and large‐scale solid state batteries.     

The <Emphasis Type="Italic">trans</Emphasis> and <Emphasis Type="Italic">cis</Emphasis> zeatin isomers play different roles in regulating growth inhibition induced by high nitrate concentrations in maize

Abscisic acid (ABA), auxins, and cytokinins (CKs) are known to be closely linked to nitrogen signaling. In particular, CKs control the effects of nitrate availability on plant growth. Our group has shown that treatment with high nitrate concentrations limits root growth and leaf development in maize, and conditions the development of younger roots and leaves. CKs also affect source-sink relationships in plants. Based on these results, we hypothesized that CKs regulate the source-sink relationship in maize via a mechanism involving complex crosstalk with the main auxin indole-3-acetic acid (IAA) and ABA. To evaluate this hypothesis, various CK metabolites, IAA, and ABA were quantified in the roots and in source and sink leaves of maize plants treated with high and normal nitrate concentrations. The data obtained suggest that the cis and trans isomers of zeatin play completely distinct roles in maize growth regulation by a complex crosstalk with IAA and ABA. We demonstrate that while trans-zeatin (tZ) and isopentenyladenine (iP) regulate nitrate uptake and thus control final leaf sizes, cis-zeatin (cZ) regulates source and sink strength, and thus controls leaf development. The implications of these findings relating to the roles of ABA and IAA in plants’ responses to varying nitrate concentrations are also discussed.