The effect of calcium chelators on microsomal pyridine nucleotide-linked dehydrogenases of sugarbeet cells

Cell suspension cultures of Beta vulgaris L., treated with calciumchelators or untreated, were used to characterize pyndine nucleotide-dependentdiaphorases of microsomes. The microsomal activity of NADH-dependentduroquinone reductase from cultures treated with 10 mM Na₂EGTAfor 24 h increased by a factor of 1.8 with respect to controlmicrosomes, and was mainly associated with particles of d=1.17gml^–1. NADPH-duroquinone reductase and NADH-ferricyanidereductase activities showed smaller increases. Bacterial protein-lipopolysaccharidecomplexes (prLPS) also promoted the increase of microsomal diaphorases;CaEGTA was Ineffective. EGTA effects on enzymes of supernatantand mitochondria were negligible, although Na₂EGTA treatmentinduced cell aggregation and strong acidification of the medium. When microsomes from control cultures were solubilized with1% LPC and fractionated in high-efficiency gel permeation columns(FPLC) the diaphorase activities were found associated to threemajor proteins: (i) NADH-specific quinone reductase (NADH-QR)of 340 kDa; (ii) pyndine nucleotide-nonspecific quinone reductase(NAD(P)H-QR) of 85 kDa also having ferricyanide reductase activity;(iii) NADH-specific ferricyanide reductase (NADH-FCR) of 38kDa. The microsomes from EGTA-treated cells also showed a highlyactive NADH-QR having a larger molecular mass (440 kDa) thanin control cells. NAD(P)H-QR also showed increased activity.We conclude that external Ca²⁺ chelation induces changes indehydrogenase components in microsomes. Furthermore, prLPS probablyexert part of their effect on plants through Ca²⁺ chelation. Key words: Beta vulgaris, cell cultures, calcium chelators, diaphorase, NAD(P)H-dehydrogenase, lipopolysaccharide, EGTA, quinone reductase     

On Circuit Functionality in Boolean Networks

It has been proved, for several classes of continuous and discrete dynamical systems, that the presence of a positive (resp. negative) circuit in the interaction graph of a system is a necessary condition for the presence of multiple stable states (resp. a cyclic attractor). A positive (resp. negative) circuit is said to be functional when it “generates” several stable states (resp. a cyclic attractor). However, there are no definite mathematical frameworks translating the underlying meaning of “generates.” Focusing on Boolean networks, we recall and propose some definitions concerning the notion of functionality along with associated mathematical results.     

Antimicrobial activity of various cationic molecules on foodborne pathogens

Antibacterial effects of various arginine- and lysine-rich polycationic proteins and polymers were evaluated by broth and solid dilution assay on a range of foodborne pathogens, Gram-positive and Gram-negative bacteria. The Minimum Inhibitory Concentration (MIC) and the Minimum Bactericidal Concentration (MBC) of α-poly-l-lysine (poly-lys), α-poly-l-arginine (poly-arg) and protamines from herring sperm (clupeine sulphate) and salmon sperm (salmine sulphate) were determined on Bacillus subtilis, Bacillus cereus, Staphylococcus aureus, Listeria monocytogenes, Salmonella typhimurium, Shigella sonnei, Escherichia coli O157:H7 and Pseudomonas aeruginosa. All these molecules showed antibacterial activity on all strains with different MIC and MBC values. The molecular mechanisms underlying the effect of α-poly-l-arginine might be related to the entrance of the molecule into the cell. In fact α-poly-l-arginine labelled with 7-Diethylamino coumarin-3-carboxylic acid, succinimidyl ester (DEAC,SE) showed ability to permeate the cell membrane of B. cereus and E. coli O157:H7.     

The incorporation of glucosamine into enterobacterial core lipopolysaccharide: two enzymatic steps are required

The core lipopolysaccharide (LPS) of Klebsiella pneumoniae is characterized by the presence of disaccharide alphaGlcN-(1,4)-alphaGalA attached by an alpha1,3 linkage to l-glycero-d-manno-heptopyranose II (ld-HeppII). Previously it has been shown that the WabH enzyme catalyzes the incorporation of GlcNAc from UDP-GlcNAc to outer core LPS. The presence of GlcNAc instead of GlcN and the lack of UDP-GlcN in bacteria indicate that an additional enzymatic step is required. In this work we identified a new gene (wabN) in the K. pneumoniae core LPS biosynthetic cluster. Chemical and structural analysis of K. pneumoniae non-polar wabN mutants showed truncated core LPS with GlcNAc instead of GlcN. In vitro assays using LPS truncated at the level of d-galacturonic acid (GalA) and cell-free extract containing WabH and WabN together led to the incorporation of GlcN, whereas none of them alone were able to do it. This result suggests that the later enzyme (WabN) catalyzes the deacetylation of the core LPS containing the GlcNAc residue. Thus, the incorporation of the GlcN residue to core LPS in K. pneumoniae requires two distinct enzymatic steps. WabN homologues are found in Serratia marcescens and some Proteus strains that show the same disaccharide alphaGlcN-(1,4)-alphaGalA attached by an alpha1,3 linkage to ld-HeppII.     

GINsim: a software suite for the qualitative modelling, simulation and analysis of regulatory networks

This paper presents GINsim, a Java software suite devoted to the qualitative modelling, analysis and simulation of genetic regulatory networks. Formally, our approach leans on discrete mathematical and graph-theoretical concepts. GINsim encompasses an intuitive graph editor, enabling the definition and the parameterisation of a regulatory graph, as well as a simulation engine to compute the corresponding qualitative dynamical behaviour. Our computational approach is illustrated by a preliminary model analysis of the inter-cellular regulatory network activating Notch at the dorsal-ventral boundary in the wing imaginal disc of Drosophila. We focus on the cross-regulations between five genes (within and between two cells), which implements the dorsal-ventral border in the developing imaginal disc. Our simulations qualitatively reproduce the wild-type developmental pathway, as well as the outcome of various types of experimental perturbations, such as loss-of-function mutations or ectopically induced gene expression.     

Antarctic scallop (Adamussium colbecki) annual growth rate at Terra Nova Bay

Adamussium colbecki is one of the most well-studied Antarctic molluscs. However, information on its growth rate is currently based on estimates from mark and recapture experiments and from growth-ring analyses. This paper provides the first estimates of the growth pattern of this scallop throughout a year, for individuals maintained under both natural field and laboratory conditions. Results show size-related differences in growth rate, both in the field and in aquaria, with scallops in Terra Nova Bay growing faster than those kept in aquaria. Growth performances were lower than those reported in the literature.     

Modulation of the TGF-β1-induced epithelial to mesenchymal transition (EMT) mediated by P1 and P2 purine receptors in MDCK cells

Epithelial to mesenchymal transition (EMT) occurs during embryogenesis or under pathological conditions such as hypoxia, injury, chronic inflammation, or tissue fibrosis. In renal tubular epithelial cells (MDCK), TGF-β1 induces EMT by reducing or increasing epithelial or mesenchymal marker expression, respectively. In this study, we confirmed that the cAMP analogues, 8-CPT-cAMP or N6-Ph-cAMP, inhibited the TGF-β1-driven overexpression of the mesenchymal markers ZEB-1, Slug, Fibronectin, and α-SMA. Furthermore, we showed that A1, A2A, P2Y1, P2Y11, and P2X7 purine receptor agonists modulated the TGF-β1-induced EMT through the involvement of PKA and/or MAPK/ERK signaling. The stimulation of A2A receptor reduced the overexpression of the EMT-related markers, mainly through the cAMP-dependent PKA pathway, as confirmed by cell pre-treatment with Myr-PKI. Both A1 and P2Y1 receptor stimulation exacerbated the TGF-β1-driven effects, which were reduced by cell pre-treatment with the MAPK inhibitor PD98059, according to the increased ERK1/2 phosphorylation upon receptor activation. The effects induced by P2Y11 receptor activation were oppositely modulated by PKA or MAPK inhibition, in line with the dual nature of the Gs- and Gq-coupled receptor. Differently, P2X7 receptor induced, per se, similar and not additive effects compared to TGF-β1, after prolonged cell exposure to BzATP. These results suggest a putative role of purine receptors as target for anti-fibrotic agents.