Fungicides degradation in an organic biomixture: impact on microbial diversity

Biological systems are being developed all over EU countries to protect water-bodies from pesticide contamination at farm level. A laboratory experiment was carried out to test the efficiency of a mixture of compost and straw in bio-degrading different mixtures of fungicides usually applied in vineyards. At the same time the effects of fungicide applications on microbial community of biomixture were also evaluated. Results showed that the biomixture had a good capability of degrading pesticides. Indeed, at the end of the experiment (112 days), the concentration of most of the pesticides was close to complete degradation. Denaturing gradient gel electrophoresis (DGGE) analysis showed an evident modification of microbial diversity after the addition of fungicides. However, at the end of degradation process, no significant changes in the composition of microbial community were seen. In this specific substrate used in the biomixture, yeast flora and ascomycete filamentous fungi seemed to be involved in the degradation activity.     

Algebraic and geometric tools in phylogenetics.

In this expository work we describe the main aspects of the so-called Phylogenetic Algebraic Geometry which concerns with the study of algebraic varieties representing statistical models of evolution. In particular, we mainly describe how this field of research can be used to infer phylogenies.     

pH dependence of tryptophan synthase catalytic mechanism: I. The first stage, the beta-elimination reaction

The pyridoxal 5'-phosphate-dependent beta-subunit of the tryptophan synthase alpha(2)beta(2) complex catalyzes the condensation of L-serine with indole to form L-tryptophan. The first stage of the reaction is a beta-elimination that involves a very fast interconversion of the internal aldimine in a highly fluorescent L-serine external aldimine that decays, via the alpha-carbon proton removal and beta-hydroxyl group release, to the alpha-aminoacrylate Schiff base. This reaction is influenced by protons, monovalent cations, and alpha-subunit ligands that modulate the distribution between open and closed conformations. In order to identify the ionizable residues that might assist catalysis, we have investigated the pH dependence of the rate of the external aldimine decay by rapid scanning UV-visible absorption and single wavelength fluorescence stopped flow. In the pH range 6-9, the reaction was found to be biphasic with the first phase (rate constants k(1)) accounting for more than 70% of the signal change. In the absence of monovalent cations or in the presence of sodium and potassium ions, the pH dependence of k(1) exhibits a bell shaped profile characterized by a pK(a1) of about 6 and a pK(a2) of about 9, whereas in the presence of cesium ions, the pH dependence exhibits a saturation profile characterized by a single pK(a) of 9. The presence of the allosteric effector indole acetylglycine increases the rate of reaction without altering the pH profile and pK(a) values. By combining structural information for the internal aldimine, the external aldimine, and the alpha-aminoacrylate with kinetic data on the wild type enzyme and beta-active site mutants, we have tentatively assigned pK(a1) to betaAsp-305 and pK(a2) to betaLys-87. The loss of pK(a1) in the presence of cesium ions might be due to a shift to lower values, caused by the selective stabilization of a closed form of the beta-subunit.     

Diffusion NMR studies on neutral and cationic square planar palladium(II) complexes

Diffusion NMR investigations were carried out in CD₂Cl₂ for a series of neutral (1-7) and cationic (8-10) square planar palladium complexes. Diffusion data were elaborated through a modified Stokes-Einstein equation that takes into account the size and shape of molecules. The hydrodynamic volume at infinite dilution of all complexes was found to be similar to the crystallographic volume and always much larger than the van der Waals volume. The self-aggregation tendency of [Pd(N,C⁻)(N,N)][PF₆] ionic complexes [(N,C⁻) = (C₆H₄-(Ph)C(O)-CN-Et); 8, (N,N) = 2,2′-bipirydine; 9, (N,N) = (2,6-(iPr)₂-C₆H₃)NC(Me)-C(Me)N(2,6-(iPr)₂-C₆H₃); 10, (N,N) = (2,6-(iPr)₂-C₆H₃)NC(R′)-C(R′)N(2,6-(iPr)₂-C₆H₃), R′₂ = naphthalene-1,8-diyl] was investigated by performing ¹H and ¹⁹F diffusion experiments as a function of the concentration. Clear evidence for the formation of ion triples containing two cationic units was obtained for 8, most likely due to the establishment of a weak Pd?O interaction. The tendency to form ion triples was much reduced in 9 and 10, having an increased steric hindrance in the apical positions. While 9 showed the usual tendency to afford a mixture of free ions and ion pairs, solvated ions were the predominant species in the case of 10 even at high concentration values (approaching 100 mM).     

The circadian system of ruin lizards: a seasonally changing neuroendocrine loop?

Previous studies have shown that the amplitude of daily melatonin production in cultured ruin lizard pineal organs explanted in the summer is significantly higher than that from organs explanted in the winter. To test whether seasonal photoperiodic changes are decoded autonomously by the pineal gland, pineals explanted in summer were cultured in vitro and exposed to changes between winter and summer photoperiods. The changes in photoperiod duration did not affect the daily profiles of in vitro melatonin production. The discrepancy between the present in vitro results and those from lizards exposed to winter or summer photoperiods before pineal explantation supports the view that circadian information entering the pineal gland via its innervation is involved in determining seasonal changes of melatonin production in ruin lizards. We further examined whether a central component of the circadian system of ruin lizards, specifically the retinae of the lateral eyes, expresses similar seasonal changes in function as does the pineal gland. We did not find any difference between summer and autumn-winter in the effectiveness of either bilateral retinalectomy or optic nerve lesion-at the level of the optic chiasm-in altering circadian locomotor behavior in constant conditions. Both surgical procedures mostly induced a shortening of the free-running period of the locomotor rhythm of similar magnitude in all seasons. Thus, the retinae do not appear to participate in the seasonal reorganization of the circadian system in ruin lizards.     

Highlights from the 5th Annual Meeting of the Italian Society of Virology

The 5th National Congress of the Italian Society of Virology (SIV) was attended by junior- and senior-level virologists to promote interactions and scientific collaborations among the different areas of Virology and allied sciences. The invited and selected lecturers covered the following topics: General Virology and Viral Genetics; Virus-host Interaction and Pathogenesis; Viral Oncogenesis; Viral Immunology and Vaccines; Anti-viral Therapy; Innovative Diagnostics; Viral Biotechnologies and Cell and Gene Therapy. As in the previous editions (Salata and Palù, 2004; Salata et al., 2005), a specific topic was thoroughly covered in a roundtable. This year the elected subject was "HIV: determinants of pathogenicity and clinical implications." The final program and the abstract book can be found at the web site http://www.siv-virologia.it. This report summarizes the lessons learned from the plenary lectures and the selected oral presentations of the 2005 meeting.     

Molecular characterization of a glycerophosphoinositol transporter in mammalian cells

The glycerophosphoinositols are ubiquitous phosphoinositide metabolites involved in the control of several cell functions. They exert their actions both intracellularly and by rapidly equilibrating across the plasma membrane when added to cells, implying the existence of a transporter for their membrane permeation. Such a transporter, GIT1, has been cloned in yeast. By PSI-BLAST analysis, we have identified the Glut2 transporter as a human-genome candidate ortholog of GIT1. This was supported directly through the use of inhibitors, siRNAs and competition studies of specific uptake of GroPIns in HeLa cells over-expressing human Glut2. These data identify Glut2 as a GroPIns transporter in mammals, and define a physiologically relevant cell-permeation mechanism.     

Low resolution structure and stability studies of human GrpE#2, a mitochondrial nucleotide exchange factor

GrpE acts as a nucleotide exchange factor for the Hsp70 chaperone system. Only one GrpE isoform is present in Escherichia coli, but for reasons not yet well understood, two GrpE isoforms have been found in mammalian mitochondria.Therefore, studies aimed at evaluating the physico-chemical characteristics of these proteins are important for the comprehension of the function of the Hsp70 chaperone system in different organisms. Here we report biophysical studies on human mitochondrial GrpE isoform 2. Small angle X-ray scattering measurements of human GrpE isoform 2 showed that this protein has a quaternary structure which is similar to those of human GrpE isoform 1 and E. coli GrpE: a dimer with a cruciform elongated shape. However, mitochondrial isoforms differed from each other regarding chemical and thermal denaturation profiles. This fact, combined with results of distinct expression patterns previously reported, point out that these proteins may have different response to external stimuli. Our results also indicate that human GrpE isoform 2 is more similar to the GrpE from E. coli than to human GrpE isoform 1. These results are relevant because differences in the conformation of Hsp70 co-chaperones are considered to be one of the reasons for functional diversity of this system.     

Hypoxia modulation of peroxisome proliferator-activated receptors (PPARs) in human glioblastoma stem cells. Implications for therapy

Gliobastoma (GB), the most common adult brain tumor, infiltrates normal brain area rendering impossible the complete surgical resection, resulting in a poor median survival (14–15 months), despite the aggressive multimodality treatments post‐surgery, such as radiation and chemo‐therapy. GB is characterized by hypoxic and necrotic regions due to a poorly organized tumor vascularization, leading to inadequate blood supply and consequently to hypoxic and necrotic areas. We have previously shown that, under hypoxia GB primary cells increased the expression of stemness markers as well as the expression of the nuclear receptor peroxisome proliferator‐activated receptor α (PPARα) and also the crucial role played by PPARs in mouse neural stem cells maintenance and differentiation. Due to the importance of lipid signaling in cell proliferation and differentiation, in this work, we analyzed the expression of PPARs in GB neurospheres both in normoxic and hypoxic conditions. The results obtained suggest a differential regulation of the three PPARs by hypoxia, thus indicating a possible therapeutic strategy to counteract GB recurrencies. J. Cell. Biochem. 113: 3342–3352, 2012. © 2012 Wiley Periodicals, Inc.     

Approaches to chemical synthetic biology

Synthetic biology is first represented in terms of two complementary aspects, the bio-engineering one, based on the genetic manipulation of extant microbial forms in order to obtain forms of life which do not exist in nature; and the chemical synthetic biology, an approach mostly based on chemical manipulation for the laboratory synthesis of biological structures that do not exist in nature. The paper is mostly devoted to shortly review chemical synthetic biology projects currently carried out in our laboratory. In particular, we describe: the minimal cell project, then the "Never Born Proteins" and lastly the Never Born RNAs. We describe and critically analyze the main results, emphasizing the possible relevance of chemical synthetic biology for the progress in basic science and biotechnology.