"Shaping" of cell signaling via AKAP-tethered PDE4D: Probing with AKAR2-AKAP5 biosensor

ABSTRACT: BACKGROUND: PKA, a key regulator of cell signaling, phosphorylates a diverse and important array of target molecules and is spatially docked to members of the A-kinase Anchoring Protein (AKAP) family. AKAR2 is a biosensor which yields a FRET signal in vivo, when phosphorylated by PKA. AKAP5, a prominent member of the AKAP family, docks several signaling molecules including PKA, PDE4D, as well as GPCRs, and is obligate for the propagation of the activation of the mitogen-activated protein kinase cascade from GPCRs to ERK1,2. RESULTS: Using an AKAR2-AKAP5 fusion "biosensor", we investigated the spatial-temporal activation of AKAP5 undergoing phosphorylation by PKA in response to beta-adrenergic stimulation. The pattern of PKA activation reported by AKAR2-AKAP5 is a more rapid and spatially distinct from those "sensed" by AKAR2-AKAP12. Spatial-temporal restriction of activated PKA by AKAP5 was found to "shape" the signaling response. Phosphatase PDE4D tethered to AKAP5 also later reverses within 60 s elevated intracellular cyclic AMP levels stimulated by beta-adrenergic agonist. AKAP12, however, fails to attenuate the rise in cyclic AMP over this time. Fusion of the AKAP5 PDE4D-binding-domain to AKAP12 was found to accelerate a reversal of accumulation of intracellular cyclic AMP. CONCLUSION: AKAPs, which are scaffolds with tethered enzymes, can "shape" the temporal and spatial aspects of cell signaling.     

Interaction of monocytes with NK cells upon Toll-like receptor-induced expression of the NKG2D ligand MICA

Reciprocal interactions between NK cells and dendritic cells have been shown to influence activation of NK cells, maturation, or lysis of dendritic cells and subsequent adaptive immune responses. However, little is known about the crosstalk between monocytes and NK cells and the receptors involved in this interaction. We report in this study that human monocytes, upon TLR triggering, up-regulate MHC class I-Related Chain (MIC) A, but not other ligands for the activating immunoreceptor NKG2D like MICB or UL-16 binding proteins 1-3. MICA expression was associated with CD80, MHC class I and MHC class II up-regulation, secretion of proinflammatory cytokines, and apoptosis inhibition, but was not accompanied by release of MIC molecules in soluble form. TLR-induced MICA on the monocyte cell surface was detected by autologous NK cells as revealed by NKG2D down-regulation. Although MICA expression did not render monocytes susceptible for NK cell cytotoxicity, LPS-treated monocytes stimulated IFN-gamma production of activated NK cells which was substantially dependent on MICA-NKG2D interaction. No enhanced NK cell proliferation or cytotoxicity against third-party target cells was observed after stimulation of NK cells with LPS-activated monocytes. Our data indicate that MICA-NKG2D interaction constitutes a mechanism by which monocytes and NK cells as an early source of IFN-gamma may communicate directly during an innate immune response to infections in humans.     

Silicon improves salinity tolerance in wheat plants

Durum wheat (Triticum durum cv. Gediz-75) and bread wheat (Triticum aestivum cv. Izmir-85) were grown in a complete nutrient solution in a growth room to investigate effect of silicone supplied to the nutrient solution on plants grown at salt stress. The experiment was a 2 × 2 factorial arrangement with two levels of NaCl in nutrient solution, 0 and 100 mM, and two levels of silicone (Si) in nutrient solution, 0.25 and 0.50 mM, as Na₂SiO₃. The plants grown at 100 mM NaCl produced less dry matter and chlorophyll content than those without NaCl. Supplementary Si at both 0.25 and 0.5 mM ameliorated the negative effects of salinity on plant dry matter and chlorophyll content. Membrane permeability and proline content in leaves increased with addition of 100 mM NaCl and these increases were decreased with Si treatments. Sodium (Na) concentration in plant tissues increased in both leaves and roots of plants in the high NaCl treatment and Si treatments lowered significantly the concentrations of Na in both leaves and roots. Bread wheat was more tolerant to salinity than durum wheat. The accumulation of Na in roots indicates a possible mechanism whereby bread wheat copes with salinity in the rooting medium and/or may indicate the existence of an inhibition mechanism of Na transport to leaves. Concentrations of both Ca and K were lower in the plants grown at high NaCl than in those in the control treatment and these two element concentrations were increased by Si treatments in both shoots and roots but remained lower than control values in most cases.     

Modeling Response of N Addition on C and N Allocation in Scandinavian Norway Spruce Stands

Because nitrogen is considered to be the major growth-limiting element in boreal forests, the increasing nitrogen availability from deposition should lead to increasing growth. We have tested this assumption by simulating, with a simple model, carbon and nitrogen development in seven long-term fertilization experiments in three Nordic countries. The only differences between sites in the model are climate, the ambient nitrogen deposition, nitrogen fertilization regimes, and the initial conditions at the start of the experiment. The model simulates the observed stand development well as long as nitrogen remains the limiting factor. The simulated retention of deposited nitrogen is in general low (less than 50%), whereas retention of fertilizer nitrogen is higher. This seems to imply that the higher production in the fertilized stands will not be maintained once the fertilization is stopped. The model results also indicate that the major effect of climate on site productivity is through soil processes, not tree physiology.     

Retinoic Acid and Vitamin E Modulate Expression and Release of CD178 in Carcinoma Cells: Consequences for Induction of Apoptosis in CD95-Sensitive Cells

CD178 (CD95-ligand) is expressed on several tumor cells and likely influences the interaction of the tumor with the host immune system. However, little is known about the mechanisms that regulate its expression on the cell surface. We have evaluated the ability of various compounds and cytokines to regulate cell surface expression and release of soluble CD178 in various carcinoma cell lines. Vitamin E succinate (VES) and retinoic acid (RA) were found to reduce CD178 surface expression, whereas interferon-gamma stimulated a slight upregulation. At 48 h, the regulation of surface CD178 by VES and RA arose from a small decrease in CD178 mRNA and to a greater extent due to an increase in the release of soluble CD178; the latter was blocked by addition of a metalloproteinase inhibitor. Accordingly, VES and RA treatment diminished the ability of tumor cells to kill CD95-sensitive cells and this effect was markedly reduced by the presence of a metalloproteinase inhibitor. Our results indicate that, in vitro, CD178 expression on the cell surface of tumor cells can be regulated by agents that alter both expression and release of the ligand. In vivo, such treatments may play an important role in the outcome of tumor sensitivity or resistance to host immune mechanisms.     

Automated identification of diseases related to lymph system from lymphography data using artificial immune recognition system with fuzzy resource allocation mechanism (fuzzy-AIRS)

Artificial immune recognition system (AIRS) classification algorithm, which has an important place among classification algorithms in the field of artificial immune systems, has showed an effective and intriguing performance on the problems it was applied. AIRS was previously applied to some medical classification problems including breast cancer, Cleveland heart disease, diabetes and it obtained very satisfactory results. So, AIRS proved to be an efficient artificial intelligence technique in medical field. In this study, the resource allocation mechanism of AIRS was changed with a new one determined by fuzzy-logic. This system, named as fuzzy-AIRS was used as a classifier in the diagnosis of lymph diseases, which is of great importance in medicine. The classifications of lymph diseases dataset taken from University of California at Irvine (UCI) Machine Learning Repository were done using 10-fold cross-validation method. Reached classification accuracies were evaluated by comparing them with reported classifiers in UCI web site in addition to other systems that are applied to the related problems. Also, the obtained classification performances were compared with AIRS with regard to the classification accuracy, number of resources and classification time. While only AIRS algorithm obtained 83.138% classification accuracy, fuzzy-AIRS classified the lymph diseases dataset with 90.00% accuracy. For lymph diseases dataset, fuzzy-AIRS obtained the highest classification accuracy according to the UCI web site. Beside of this success, fuzzy-AIRS gained an important advantage over the AIRS by means of classification time. By reducing classification time as well as obtaining high classification accuracies in the applied datasets, fuzzy-AIRS classifier proved that it could be used as an effective classifier for medical problems.     

Turnover of Phospholipids in an Unsaturated Fatty Acid Auxotroph of Escherichia coli

The membrane phospholipids of an unsaturated fatty acid auxotroph of Escherichia coli were found to undergo turnover. These phospholipids were excreted into the culture medium, and were replaced in the cell with newly synthesized phospholipids. Phospholipids of growing cells supplemented with elaidic acid underwent rapid turnover, while those of cells supplemented with oleate, or cis-vaccenate plus palmitoleate, underwent slow turnover. Starvation for required amino acids stimulated this turnover in the latter two cases. Protein was also lost from growing cells. However, after amino acid starvation this loss ceased while phospholipid turnover continued. Electron micrographs of growing cells indicated that large pieces of membrane-like material were separating from the cell surface.     

Purification and characterization of a bacteriocin from an oenological strain of Leuconostoc mesenteroides subsp. cremoris

Malolactic fermentation (MLF), which improves organoleptic properties and biologic stability of some wines, may cause wine spoilage if uncontrolled. Bacteriocins were reported as efficient preservatives to control MLF through their bactericidal effect on malolactic bacteria. Leuconostoc mesenteroides subsp. cremoris W3 isolated from wine produces an inhibitory substance that is bactericidal against malolactic bacteria in model wine medium. Treatment of the culture supernatant of strain W3 with proteases eliminated the inhibitory activity, which proved that it is a true bacteriocin and we tentatively termed it mesentericin W3. The bacteriocin inhibited the growth of food-borne pathogenic bacteria such as Enterococcus faecalis, Listeria monocytogenes, and malolactic bacteria. It was active over a wide pH range and stable to organic solvents and heat. Mesentericin W3 was purified to homogeneity by a pH-mediated cell adsorption–desorption method, cation exchange, hydrophobic interaction, and reverse-phase chromatography. Matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectroscopy (MS) and partial amino acid sequence analysis revealed that mesentericin W3 was identical to mesentericin Y105.