The Binding Pocket at the Interface of Multimeric Telomere G-quadruplexes: Myth or Reality?

Human telomeric DNA with hundreds of repeats of the 5’-TTAGGG-3’ motif plays a crucial role in several biological processes. It folds into G-quadruplex (G4) structures and features a pocket at the interface of two contiguous G4 blocks. Up to now no structural NMR and crystallographic data are available for ligands interacting with contiguous G4s. Naphthalene diimide monomers and dyads were investigated as ligands of a dimeric G4 of human telomeric DNA comparing the results with those of the model monomeric G4. Time-resolved fluorescence, circular dichroism, isothermal titration calorimetry and molecular modeling were used to elucidate binding features. Ligand fluorescence lifetime and induced circular dichroism unveiled occupancy of the binding site at the interface. Thermodynamic parameters confirmed the hypothesis as they remarkably change for the dyad complexes of the monomeric and dimeric telomeric G4. The bi-functional ligand structure of the dyads is a fundamental requisite for binding at the G4 interface as only the dyads engage in complexes with 1 : 1 stoichiometry, lodging in the pocket at the interface and establishing multiple interactions with the DNA skeleton. In the absence of NMR and crystallographic data, our study affords important proofs of binding at the interface pocket and clues on the role played by the ligand structure

A naphthalene diimide dyad for fluorescence switch-on detection of G-quadruplexes

A non-fluorescent dimeric naphthalene diimide dye becomes red emitting upon G-quadruplex binding

Hybrid integration of microfabricated chemοcapacitor arrays with miniaturized read-out electronics towards low-power gas sensing module

AbstractA hybrid gas sensing module consisting of an array of 8 polymer coated capacitive sensors and low power read-out electronics is introduced. The chemocapacitor array is fabricated with standard microelectronics/micromachining processes allowing for the realization of planar InterDigitated Electrodes (IDEs). The read-out electronics sub- module consists of an analog multiplexer for the sequential measurement of the sensor array elements, a capacitance to 24-bit converter and a USB to I2C interface. The compact hybrid module has been successfully applied in the detection of sub-100ppm concentrations of p-xylene and toluene. The responses to various humidity levels have been also evaluated

New applications for known drugs: Human glycogen synthase kinase 3 inhibitors as modulators of Aspergillus fumigatus growth [Postprint]

Invasive aspergillosis (IA) is one of the most severe forms of fungi infection. IA disease is mainly due to Aspergillus fumigatus, an air-borne opportunistic pathogen. Mortality rate caused by IA is still very high (50-95%), because of difficulty in early diagnostics and reduced antifungal treatment options, thus new and efficient drugs are necessary. The aim of this work is, using Aspergillus nidulans as non-pathogen model, to develop efficient drugs to treat IA. The recent discovered role of glycogen synthase kinase-3 homologue, GskA, in A. fumigatus human infection and our previous experience on human GSK-3 inhibitors focus our attention on this kinase as a target for the development of antifungal drugs. With the aim to identify effective inhibitors of colonial growth of A. fumigatus we use A. nidulans as an accurate model for in vivo and in silico studies. Several well-known human GSK-3β inhibitors were tested for inhibition of A. nidulans colony growth. Computational tools as docking studies and binding site prediction was used to explain the different biological profile of the tested inhibitors. Three of the five tested hGSK3β inhibitors are able to reduce completely the colonial growth by covalent bind to the enzyme. Therefore these compounds may be useful in different applications to eradicate IA.SAF2012-37979-C03-01 to A.M; BFU2012-33142 to E.A.EN

Dyads of G-Quadruplex Ligands Triggering DNA Damage Response and Tumour Cell Growth Inhibition at Subnanomolar Concentration

Naphthalene diimide (NDI) dyads exhibiting a different substitution pattern and linker length have been synthesised and evaluated as G-quadruplex (G4) ligands, by investigating their cytotoxicity in selected cell lines. The dyads with the long C7 linker exhibit extremely low IC50 values, below 10\u2005nm, on different cancer cell lines. Contrary, the dyads with the shorter C4 linker were much less effective, with IC values increasing up to 1\u2005\u3bcm. Among the three dyads with the longest linker, small differences in the IC50 values emerge, suggesting that the linker length plays a more important role than the substitution pattern. We have further shown that the dyads are able to induce cellular DNA damage response, which is not limited to the telomeric regions and is likely the origin of their cytotoxicity. Both absorption titration and dynamic light scattering of the most cytotoxic dyads in the presence of hTel22 highlight their ability to induce effective G4 aggregation, acting as non-covalent cross-linking agents

The structure of the Shiga toxin 2a A-subunit dictates the interactions of the toxin with blood components

Hemolytic uremic syndrome (eHUS) is a severe complication of human infections with Shiga toxins (Stxs)-producing Escherichia coli. A key step in the pathogenesis of eHUS is the interaction of Stxs with blood components before the targeting of renal endothelial cells. Here, we show that a single proteolytic cleavage in the Stx2a A-subunit, resulting into two fragments (A1 and A2) linked by a disulfide bridge (cleaved Stx2a), dictates different binding abilities. Uncleaved Stx2a was confirmed to bind to human neutrophils and to trigger leukocyte/platelet aggregate formation, whereas cleaved Stx2a was ineffective. Conversely, binding of complement factor H was confirmed for cleaved Stx2a and not for uncleaved Stx2a. It is worth noting that uncleaved and cleaved Stx2a showed no differences in cytotoxicity for Vero cells or Raji cells, structural conformation, and contaminating endotoxin. These results have been obtained by comparing two Stx2a batches, purified in different laboratories by using different protocols, termed Stx2a(cl; cleaved toxin, Innsbruck) and Stx2a(uncl; uncleaved toxin, Bologna). Stx2a(uncl) behaved as Stx2a(cl) after mild trypsin treatment. In this light, previous controversial results obtained with purified Stx2a has to be critically re-evaluated; furthermore, characterisation of the structure of circulating Stx2a is mandatory to understand eHUS-pathogenesis and to develop therapeutic approaches

Correlation-maximizing surrogate gene space for visual mining of gene expression patterns in developing barley endosperm tissue

<p>Abstract</p> <p>Background</p> <p>Micro- and macroarray technologies help acquire thousands of gene expression patterns covering important biological processes during plant ontogeny. Particularly, faithful visualization methods are beneficial for revealing interesting gene expression patterns and functional relationships of coexpressed genes. Such screening helps to gain deeper insights into regulatory behavior and cellular responses, as will be discussed for expression data of developing barley endosperm tissue. For that purpose, high-throughput multidimensional scaling (HiT-MDS), a recent method for similarity-preserving data embedding, is substantially refined and used for (a) assessing the quality and reliability of centroid gene expression patterns, and for (b) derivation of functional relationships of coexpressed genes of endosperm tissue during barley grain development (0–26 days after flowering).</p> <p>Results</p> <p>Temporal expression profiles of 4824 genes at 14 time points are faithfully embedded into two-dimensional displays. Thereby, similar shapes of coexpressed genes get closely grouped by a correlation-based similarity measure. As a main result, by using power transformation of correlation terms, a characteristic cloud of points with bipolar sandglass shape is obtained that is inherently connected to expression patterns of pre-storage, intermediate and storage phase of endosperm development.</p> <p>Conclusion</p> <p>The new HiT-MDS-2 method helps to create global views of expression patterns and to validate centroids obtained from clustering programs. Furthermore, functional gene annotation for developing endosperm barley tissue is successfully mapped to the visualization, making easy localization of major centroids of enriched functional categories possible.</p

Robust Hydrophobic and Hydrophilic Polymer Fibers Sensitized by Inorganic and Hybrid Lead Halide Perovskite Nanocrystal Emitters

Advances in the technology and processing of flexible optical materials have paved the way toward the integration of semiconductor emitters and polymers into functional light emitting fabrics. Lead halide perovskite nanocrystals appear as highly suitable optical sensitizers for such polymer fiber emitters due to their ease of fabrication, versatile solution-processing and highly efficient, tunable, and narrow emission across the visible spectrum. A beneficial byproduct of the nanocrystal incorporation into the polymer matrix is that it provides a facile and low-cost method to chemically and structurally stabilize the perovskite nanocrystals under ambient conditions. Herein, we demonstrate two types of robust fiber composites based on electrospun hydrophobic poly(methyl methacrylate) (PMMA) or hydrophilic polyvinylpyrrolidone (PVP) fibrous membranes sensitized by green-emitting all-inorganic CsPbBr3 or hybrid organic-inorganic FAPbBr3 nanocrystals. We perform a systematic investigation on the influence of the nanocrystal-polymer relative content on the structural and optical properties of the fiber nanocomposites and we find that within a wide content range, the nanocrystals retain their narrow and high quantum yield emission upon incorporation into the polymer fibers. Quenching of the radiative recombination at the higher/lower bound of the nanocrystal:polymer mass ratio probed is discussed in terms of nanocrystal clustering/ligand desorption due to dilution effects, respectively. The nanocomposite's optical stability over an extended exposure in air and upon immersion in water is also discussed. The studies confirm the demonstration of robust and bright polymer-fiber emitters with promising applications in backlighting for LCD displays and textile-based light emitting devices

Catecholaminergic signalling through thymic nerve fibres, thymocytes and stromal cells is dependent on both circulating and locally synthesized glucocorticoids

Glucocorticoids have been shown to modulate the expression of noradrenaline metabolizing enzymes and beta(2)- and alpha(1B)-adrenoceptors in a tissue- and cell- specific manner. In the thymus, apart from extensive sympathetic innervation, a regulatory network has been identified that encompasses catecholamine-containing non-lymphoid and lymphoid cells. We examined a putative role of adrenal- and thymus-derived glucocorticoids in modulation of rat thymic noradrenaline levels and adrenoceptor expression. Seven days postadrenalectomy, the thymic levels of mRNAs encoding tyrosine hydroxylase, dopamine beta-hydroxylase, monoamine oxidase-A and, consequently, noradrenaline were decreased. Catecholamine content was diminished in autofluorescent nerve fibres (judging by the intensity of fluorescence) and thymocytes (considering HPLC measurements of noradrenaline and the frequency of tyrosine hydroxylase-positive cells), while it remained unaltered in non-lymphoid autofluorescent cells. In addition, adrenalectomy diminished the thymocyte expression of beta(2)- and alpha(1B)-adrenoceptors at both mRNA and protein levels. Administration of ketoconazole (an inhibitor of glucocorticoid synthesis/action; 25 mg kg(-1) day(-1), s.c.) to glucocorticoid-deprived rats increased the thymic levels of tyrosine hydroxylase, dopamine beta-hydroxylase and, consequently, noradrenaline. The increased intensity of the autofluorescent cell fluorescence in ketoconazole-treated rats indicated an increase in their catecholamine content, and suggested differential glucocorticoid-mediated regulation of catecholamines in thymic lymphoid and non-lymphoid cells. In addition, ketoconazole increased the thymocyte expression of alpha(1B)-adrenoceptors. Thus, this study indicates that in the thymus, as in some other tissues, glucocorticoids not only act in concert with cateholamines, but they may modulate catecholamine action by tuning thymic catecholamine metabolism and adrenoceptor expression in a cell-specific manner. Additionally, the study indicates a role of thymus-derived glucocorticoids in this modulation