A mouse model for Li-Fraumeni-Like Syndrome with cardiac angiosarcomas associated to POT1 mutations

The shelterin protein POT1 has been found mutated in many different familial and sporadic cancers, however, no mouse models to understand the pathobiology of these mutations have been developed so far. To address the molecular mechanisms underlying the tumorigenic effects of POT1 mutant proteins in humans, we have generated a mouse model for the human POT1^R117C mutation found in Li-Fraumeni-Like families with cases of cardiac angiosarcoma by introducing this mutation in the Pot1a endogenous locus, knock-in for Pot1a^R117C. We find here that both mouse embryonic fibroblasts (MEFs) and tissues from Pot1a^+/ki mice show longer telomeres than wild-type controls. Longer telomeres in Pot1a^+/ki MEFs are dependent on telomerase activity as they are not found in double mutant Pot1a^+/ki Tert^-/- telomerase-deficient MEFs. By using complementation assays we further show that POT1a pR117C exerts dominant-negative effects at telomeres. As in human Li-Fraumeni patients, heterozygous Pot1a^+/ki mice spontaneously develop a high incidence of angiosarcomas, including cardiac angiosarcomas, and this is associated to the presence of abnormally long telomeres in endothelial cells as well as in the tumors. The Pot1a^+/R117C mouse model constitutes a useful tool to understand human cancers initiated by POT1 mutations.     

Genotoxicity of Nicotiana tabacum leaves on Helix aspersa

Tobacco farmers are routinely exposed to complex mixtures of inorganic and organic chemicals present in tobacco leaves. In this study, we examined the genotoxicity of tobacco leaves in the snail Helix aspersa as a measure of the risk to human health. DNA damage was evaluated using the micronucleus test and the Comet assay and the concentration of cytochrome P450 enzymes was estimated. Two groups of snails were studied: one fed on tobacco leaves and one fed on lettuce (Lactuca sativa L) leaves (control group). All of the snails received leaves (tobacco and lettuce leaves were the only food provided) and water ad libitum. Hemolymph cells were collected after 0, 24, 48 and 72 h. The Comet assay and micronucleus test showed that exposure to tobacco leaves for different periods of time caused significant DNA damage. Inhibition of cytochrome P450 enzymes occurred only in the tobacco group. Chemical analysis indicated the presence of the alkaloid nicotine, coumarins, saponins, flavonoids and various metals. These results show that tobacco leaves are genotoxic in H. aspersa and inhibit cytochrome P450 activity, probably through the action of the complex chemical mixture present in the plant.     

Analysis of the marine ornamental fish trade at Ceará State, northeast Brazil

Brazil is one of the leading exporters of ornamental fishes, mostly freshwater; however, monitoring of the trade is nearly non-existent in the country. This paper provides an initial assessment of a new venture, the marine aquarium fish trade at Ceará State, northeast Brazil, aiming to document the species traded, to provide preliminary estimates of numbers of specimens traded, and to identify priorities in data collection and monitoring. A total of 143 species and 199 304 fishes were traded. From the total, 109 species were native and represented 84% of the fishes traded. Thirty-four exotic species figured on the permits and amounted to nearly 16% of the exports; however, most of them consist of misidentified native species. Nearly 90% of the fish trade was directed to the international market. Official figures represent an underestimation of the total number of captured specimens.     

Alexandrium catenella and Alexandrium minutum blooms in the Mediterranean Sea: Toward the identification of ecological niches

Annual recurrent blooms of the toxic dinoflagellates Alexandrium catenella and Alexandrium minutum were detected from 2000 to 2003 in harbours along the Catalan coast. The interrelation study between the occurrence of the blooms and specific external conditions at the study sites demonstrated that different factors are required for the bloom of each Alexandrium species. Concentrations higher than 10⁵ cells l⁻¹ of A. catenella were only detected in Tarragona harbour. These blooms were associated with water surface temperature between 21 and 25 °C and salinities of around 34 psu or higher than 37 psu. A. minutum appeared widely spread along the Catalan coast, though the most intensive and recurrent blooms of this species were observed in Arenys de Mar harbour. Concentrations of millions of cells per litre of A. minutum were associated with water temperatures below 14 °C and salinities of around 34–36 psu. A. minutum cell densities showed a positive significant correlation with NO₃ but a negative correlation with NH₄. On the other hand, A. catenella blooms dominated when both NO₃ and NH₄ levels were high. The prevailing inorganic nitrogen form (NO₃ vs. NH₄) could explain why these two species rarely coincide in the same harbours. Accumulation of cysts in the sediment was found to be an important potential factor for the recurrence of these species. The 4.3 × 10³ A. catenella cysts cm⁻³ of wet sediment in Tarragona harbour and the 3.02 × 10³ A. minutum cysts cm⁻³ of wet sediment in Vilanova harbour were the highest concentrations observed from the cyst study. Confined waters such as harbours play an important role as reservoirs for the accumulation of cysts and vegetative cells, which contributes to the expansion of these dinoflagellates in the region. However, the particular environmental conditions are also decisive factors of bloom intensity.     

Stereoselective convergent synthesis of 24-substituted metabolites and analogues of vitamin D

The synthesis of vitamin D(3) active metabolites [24R,25-(OH)(2)-D(3), 24S,25-(OH)(2)-D(3) and 1alpha,24R,25-(OH)(3)-D(3)] and the first 24-aminovitamin D(3) derivatives [24S-benzoylamino-25-OH-D(3) and 24S-benzoylamino-1alpha,25-(OH)(2)-D(3)] are reported. The stereogenic center at C-24 was generated through ultrasonically induced aqueous conjugate addition of iodide 8 to dioxolanone 6 or oxazolidinone 7. The vitamin D triene system was constructed using the Lythgoe approach. The synthetic route, which is both short (6 or 7 steps from iodide 8) and efficient (32-45% overall yield), constitutes a practical method for the preparation of 24-functionalized metabolites and analogues of vitamin D(3). The ultrasonically induced conjugate addition in the key step provides a novel example of a highly stereoselective reaction promoted by the zinc-copper couple in aqueous media.     

Strategy for fluorescent labeling of human acidic fibroblast growth factor without impairment of mitogenic activity: a bona fide tracer

A deuterium reagent, 1-(d5) phenyl-3-methyl-5-pyrazolone (d5-PMP), has been synthesized and used for relative quantitative analysis of oligosaccharides by mass spectrometry (MS) using d0/d5-PMP stable isotopic labeling. Previously reported permethylation-based isotopic labels generate variable mass differences, and reductive amination-based isotopic labels cause a loss of some acid-labile groups in carbohydrates. In contrast, d0/d5-PMP stable isotopic labeling is performed at the reducing end of glycans under basic conditions without desialylation, and the mass difference (Δm = 10 Da) between the heavy form (d5-PMP derivative) and light form (d0-PMP derivative) of each glycan is invariable. When the two derivative forms of a glycan are mixed in equimolar amounts, a pair of peaks with a 10-Da mass differences is observed in the MS profile. The difference at relative intensity between the d0- and d5-PMP derivatives reflects the difference in quantity of glycans in two samples, making it possible to carry out both qualitative and relative quantitative analyses of glycans in glycomic studies. Application of this method on DP₂ to DP₆ maltodextrin oligosaccharides and N-linked glycans released from ribonuclease B and bovine fetuin demonstrates a 10-fold relative quantitative dynamic range, a satisfying reproducibility (coefficient of variation [CV] ? 8.34%), and good accuracy (relative error [RE] ? 5.1%) of the method. The suggested technique has been successfully applied for comparative quantitative analysis of free oligosaccharides in human and bovine milk.     

Diverse mechanisms of metaeffector activity in an intracellular bacterial pathogen,Legionella pneumophila

Pathogens deliver complex arsenals of translocated effector proteins to host cells during infection, but the extent to which these proteins are regulated once inside the eukaryotic cell remains poorly defined. Among all bacterial pathogens, Legionella pneumophila maintains the largest known set of translocated substrates, delivering over 300 proteins to the host cell via its Type IVB, Icm/Dot translocation system. Backed by a few notable examples of effector–effector regulation in L. pneumophila, we sought to define the extent of this phenomenon through a systematic analysis of effector–effector functional interaction. We used Saccharomyces cerevisiae, an established proxy for the eukaryotic host, to query > 108,000 pairwise genetic interactions between two compatible expression libraries of ~330 L. pneumophila‐translocated substrates. While capturing all known examples of effector–effector suppression, we identify fourteen novel translocated substrates that suppress the activity of other bacterial effectors and one pair with synergistic activities. In at least nine instances, this regulation is direct—a hallmark of an emerging class of proteins called metaeffectors, or “effectors of effectors”. Through detailed structural and functional analysis, we show that metaeffector activity derives from a diverse range of mechanisms, shapes evolution, and can be used to reveal important aspects of each cognate effector's function. Metaeffectors, along with other, indirect, forms of effector–effector modulation, may be a common feature of many intracellular pathogens—with unrealized potential to inform our understanding of how pathogens regulate their interactions with the host cell.