Genetic modification of potato against microbial diseases: in vitro and in planta activity of a dermaseptin B1 derivative, MsrA2

Dermaseptin B1 is a potent cationic antimicrobial peptide found in skin secretions of the arboreal frog Phyllomedusa bicolor. A synthetic derivative of dermaseptin B1, MsrA2 (N-Met-dermaseptin B1), elicited strong antimicrobial activities against various phytopathogenic fungi and bacteria in vitro. To assess its potential for plant protection, MsrA2 was expressed at low levels (1–5 g/g of fresh tissue) in the transgenic potato (Solanum tuberosum L.) cv. Desiree. Stringent challenges of these transgenic potato plants with a variety of highly virulent fungal phytopathogens—Alternaria, Cercospora, Fusarium, Phytophthora, Pythium, Rhizoctonia and Verticillium species—and with the bacterial pathogen Erwinia carotovora demonstrated that the plants had an unusually broad-spectrum and powerful resistance to infection. MsrA2 profoundly protected both plants and tubers from diseases such as late blight, dry rot and pink rot and markedly extended the storage life of tubers. Due to these properties in planta, MsrA2 is proposed as an ideal antimicrobial peptide candidate to significantly increase resistance to phytopathogens and improve quality in a variety of crops worldwide with the potential to obviate fungicides and facilitate storage under difficult conditions.     

X-ray structure of two crystalline forms of a streptomycete ribonuclease with cytotoxic activity

Ribonuclease (RNase) Sa3 is secreted by the Gram-positive bacterium Streptomyces aureofaciens. The enzyme catalyzes the cleavage of RNA on the 3' side of guanosine residues. Here, x-ray diffraction analysis was used to determine the three-dimensional structure of two distinct crystalline forms of RNase Sa3 to a resolution of 2.0 and 1.7 A. These two structures are similar to each other as well as to that of a homolog, RNase Sa. All of the key active-site residues of RNase Sa (Asn(42), Glu(44), Glu(57), Arg(72), and His(88)) are located in the putative active site of RNase Sa3. Also herein, RNase Sa3 is shown to be toxic to human erythroleukemia cells in culture. Like onconase, which is an amphibian ribonuclease in Phase III clinical trials as a cancer chemotherapeutic, RNase Sa3 is not inhibited by the cytosolic ribonuclease inhibitor protein. Thus, a prokaryotic ribonuclease can be toxic to mammalian cells.     

Complement mediates the binding of HIV to erythrocytes

A fraction of HIV is associated with erythrocytes even when the virus becomes undetectable in plasma under antiretroviral therapy. The aim of the present work was to further characterize this association in vitro. We developed an in vitro model to study the factors involved in the adherence of HIV-1 to erythrocytes. Radiolabeled HIV-1 (HIV) and preformed HIV-1/anti-HIV immune complexes (HIV-IC) were opsonized in various human sera, purified using sucrose density gradient ultracentrifugation, and incubated with human erythrocytes. We observed that, when opsonized in normal human serum, not only HIV-IC, but also HIV, bound to erythrocytes, although the adherence of HIV was lower than that of HIV-IC. The adherence was abolished when the complement system was blocked, but was maintained in hypogammaglobulinemic sera. Complement-deficient sera indicated that both pathways of complement were important for optimal adherence. No adherence was seen in C1q-deficient serum, and the adherence of HIV was reduced when the alternative pathway was blocked using anti-factor D Abs. The adherence could be inhibited by an mAb against complement receptor 1. At supraphysiological concentrations, purified C1q mediated the binding of a small fraction of HIV and HIV-IC to erythrocytes. In conclusion, HIV-IC bound to erythrocytes as other types of IC do when exposed to complement. Of particular interest was that HIV alone bound also to erythrocytes in a complement/complement receptor 1-dependent manner. Thus, erythrocytes may not only deliver HIV-IC to organs susceptible to infection, but free HIV as well. This may play a crucial role in the progression of the primary infection.     

Preservation of Metabolic Flexibility in Skeletal Muscle by a Combined Use of n-3 PUFA and Rosiglitazone in Dietary Obese Mice

Insulin resistance, the key defect in type 2 diabetes (T2D), is associated with a low capacity to adapt fuel oxidation to fuel availability, i.e., metabolic inflexibility. This, in turn, contributes to a further damage of insulin signaling. Effectiveness of T2D treatment depends in large part on the improvement of insulin sensitivity and metabolic adaptability of the muscle, the main site of whole-body glucose utilization. We have shown previously in mice fed an obesogenic high-fat diet that a combined use of n-3 long-chain polyunsaturated fatty acids (n-3 LC-PUFA) and thiazolidinediones (TZDs), anti-diabetic drugs, preserved metabolic health and synergistically improved muscle insulin sensitivity. We investigated here whether n-3 LC-PUFA could elicit additive beneficial effects on metabolic flexibility when combined with a TZD drug rosiglitazone. Adult male C57BL/6N mice were fed an obesogenic corn oil-based high-fat diet (cHF) for 8 weeks, or randomly assigned to various interventions: cHF with n-3 LC-PUFA concentrate replacing 15% of dietary lipids (cHF+F), cHF with 10 mg rosiglitazone/kg diet (cHF+ROSI), cHF+F+ROSI, or chow-fed. Indirect calorimetry demonstrated superior preservation of metabolic flexibility to carbohydrates in response to the combined intervention. Metabolomic and gene expression analyses in the muscle suggested distinct and complementary effects of the interventions, with n-3 LC-PUFA supporting complete oxidation of fatty acids in mitochondria and the combination with n-3 LC-PUFA and rosiglitazone augmenting insulin sensitivity by the modulation of branched-chain amino acid metabolism. These beneficial metabolic effects were associated with the activation of the switch between glycolytic and oxidative muscle fibers, especially in the cHF+F+ROSI mice. Our results further support the idea that the combined use of n-3 LC-PUFA and TZDs could improve the efficacy of the therapy of obese and diabetic patients.     

Impact of the auxin signaling inhibitor p-chlorophenoxyisobutyric acid on short-term Cd-induced hydrogen peroxide production and growth response in barley root tip

Short-term treatment (30min) of barley roots with a low 10μM Cd concentration induced significant H(2)O(2) production in the elongation and differentiation zone of the root tip 3h after treatment. This elevated H(2)O(2) production was accompanied by root growth inhibition and probably invoked root swelling in the elongation zone of the root tip. By contrast, a high 60μM Cd concentration induced robust H(2)O(2) production in the elongation zone of the root tip already 1h after short-term treatment. This robust H(2)O(2) generation caused extensive cell death 6h after short-term treatment. Similarly to low Cd concentration, exogenously applied H(2)O(2) caused marked root growth inhibition, which at lower H(2)O(2) concentration was accompanied by root swelling. The auxin signaling inhibitor p-chlorophenoxyisobutyric acid effectively inhibited 10μM Cd-induced root growth inhibition, H(2)O(2) production and root swelling, but was ineffective in the alleviation of 60μM Cd-induced root growth inhibition and H(2)O(2) production. Our results demonstrated that Cd-induced mild oxidative stress caused root growth inhibition, likely trough the rapid reorientation of cell growth in which a crucial role was played by IAA signaling in the root tip. Strong oxidative stress induced by high Cd concentration caused extensive cell death in the elongation zone of the root tip, resulting in the cessation of root growth or even in root death.     

STDP rule endowed with the BCM sliding threshold accounts for hippocampal heterosynaptic plasticity

We have combined the nearest neighbour additive spike-timing-dependent plasticity (STDP) rule with the Bienenstock, Cooper and Munro (BCM) sliding modification threshold in a computational model of heterosynaptic plasticity in the hippocampal dentate gyrus. As a result we can reproduce (1) homosynaptic long-term potentiation of the tetanized input, and (2) heterosynaptic long-term depression of the untetanized input, as observed in real experiments. Action Editor: Nicolas Brunel     

The influence of simvastatin, atorvastatin and high-cholesterol diet on acetylcholinesterase activity, amyloid beta and cholesterol synthesis in rat brain

OBJECTIVE: There is evidence to suppose that cholesterol-lowering medicine might confer protection against dementia, probably via modulation of cholesterol synthesis in the brain. The aim of the present study was to investigate the potential influence of statins and cholesterol diet on selected parameters relevant to Alzheimer's disease pathophysiology. METHODS: For 15 days, rats were orally administered simvastatin (10 or 20mg/kg b.wt.), atorvastatin (10 or 20mg/kg b.wt.), or aqua (control group); and one group was fed high-cholesterol (2%) diet. At the end of experiments brain (and plasma) cholesterol, lathosterol, hydroxymethylglutaryl-coenzyme A reductase protein, acetylcholinesterase activity, amyloid beta (40 and 42) and cholesterol synthesis rate (using the incorporation of deuterium from deuterated water) were determined and statistically compared to those of aqua. RESULTS: Both statins were able to lower cholesterol in the plasma, but none elicited an effect on total brain cholesterol. Significant reductions of brain lathosterol and cholesterol synthesis rate were observed after simvastatin and atorvastatin treatment. Acetylcholinesterase activity, amyloid beta and hydroxymethylglutaryl-coenzyme A reductase levels remained unaffected by the two drugs. CONCLUSIONS: This study brings additional evidence of a role for statins in cholesterol synthesis in the brain. Our data question the relationship between amyloid beta, acetylcholinesterase activity and cholesterol synthesis in the rat brain as well as the assumption about no exchange between peripheral and brain cholesterol pools.