Annexin-actin interactions

The actin cytoskeleton is a malleable framework of polymerised actin monomers that may be rapidly restructured to enable diverse cellular activities such as motility, endocytosis and cytokinesis. The regulation of actin dynamics involves the coordinated activity of numerous proteins, among which members of the annexin family of Ca2+- and phospholipid-binding proteins play an important role. Although the roles of annexins in actin dynamics are not understood at a mechanistic level, annexins have the requisite properties to integrate Ca2+-signaling with actin dynamics at membrane contact sites. In this review we discuss the current state of knowledge on this topic, and consider how and where annexins may fit into the complex molecular machinery that regulates the actin cytoskeleton.     

<Emphasis Type="Italic">Erysiphe catalpae</Emphasis> and <Emphasis Type="Italic">Erysiphe elevata</Emphasis> in Europe

The recent epidemic spread of the North American powdery mildew Erysiphe elevata in Europe is described and discussed. Since 2002, this plant pathogenic fungus has been collected on Catalpa bignonioides, C. erubescens and C. speciosa in the Czech Republic, Germany, Hungary, Slovakia and Switzerland. The diagnostically important anamorph of E. elevata, so far unknown, is described and illustrated in detail. Type material of Erysiphe catalpae and two specimens of E. catalpae recently collected in Poland have been examined and compared with E. elevata. The anamorph as well as the teleomorph of E. catalpae proved to be easily distinguishable from E. elevata. The supposition that E. catalpae, introduced in Armenia, was based on immature ascomata of E. elevata proved to be wrong. The origin and distribution of E. catalpae are discussed, and a key to powdery mildew fungi on Catalpa spp. in Europe is provided.     

Emotional and cognitive reinforcement of rat hippocampal long-term potentiation by different learning paradigms

In earlier studies we have shown that a protein-synthesis-independent, early, long-term potentiaton (early-LTP) that lasts up to 4-5 hours can be transformed (reinforced) into a protein-synthesis-dependent late-LTP that lasts > or = 8 hours by either an emotional challenge (e.g. swim stress) or mastering a cognitive task (e.g. spatial learning). In the present study we show that LTP-reinforcement by spatial training depends on the specific constraints of the learning paradigm. In a holeboard paradigm,LTP-reinforcement is related to the formation of a lasting reference memory whereas water-maze training gives more heterogenous results. Thus, cognitive aspects interfere with emotionally challenging components of the latter paradigm. These data indicate that different spatial-learning tasks are weighted distinctly by the animal. Thus, we show that aspects of specific spatial learning paradigms such as shifts of attention and emotional content directly influence functional plasticity and memory formation.     

A catalog of human cDNA expression clones and its application to structural genomics

We describe here a systematic approach to the identification of human proteins and protein fragments that can be expressed as soluble proteins in Escherichia coli. A cDNA expression library of 10,825 clones was screened by small-scale expression and purification and 2,746 clones were identified. Sequence and protein-expression data were entered into a public database. A set of 163 clones was selected for structural analysis and 17 proteins were prepared for crystallization, leading to three new structures.     

Behaviour-based modelling of hexapod locomotion: linking biology and technical application

Walking in insects and most six-legged robots requires simultaneous control of up to 18 joints. Moreover, the number of joints that are mechanically coupled via body and ground varies from one moment to the next, and external conditions such as friction, compliance and slope of the substrate are often unpredictable. Thus, walking behaviour requires adaptive, context-dependent control of many degrees of freedom. As a consequence, modelling legged locomotion addresses many aspects of any motor behaviour in general. Based on results from behavioural experiments on arthropods, we describe a kinematic model of hexapod walking: the distributed artificial neural network controller walknet. Conceptually, the model addresses three basic problems in legged locomotion. (I) First, coordination of several legs requires coupling between the step cycles of adjacent legs, optimising synergistic propulsion, but ensuring stability through flexible adjustment to external disturbances. A set of behaviourally derived leg coordination rules can account for decentralised generation of different gaits, and allows stable walking of the insect model as well as of a number of legged robots. (II) Second, a wide range of different leg movements must be possible, e.g. to search for foothold, grasp for objects or groom the body surface. We present a simple neural network controller that can simulate targeted swing trajectories, obstacle avoidance reflexes and cyclic searching-movements. (III) Third, control of mechanically coupled joints of the legs in stance is achieved by exploiting the physical interactions between body, legs and substrate. A local positive displacement feedback, acting on individual leg joints, transforms passive displacement of a joint into active movement, generating synergistic assistance reflexes in all mechanically coupled joints.     

Fast identification of folded human protein domains expressed in <Emphasis Type="Italic">E. coli</Emphasis> suitable for structural analysis

Background  

High-throughput protein structure analysis of individual protein domains requires analysis of large numbers of expression clones to identify suitable constructs for structure determination. For this purpose, methods need to be implemented for fast and reliable screening of the expressed proteins as early as possible in the overall process from cloning to structure determination.     

Hypoxia-induced inhibition of whole cell membrane currents and ion transport of A549 cells

In excitable cells, hypoxia inhibits K channels, causes membrane depolarization, and initiates complex adaptive mechanisms. It is unclear whether K channels of alveolar epithelial cells reveal a similar response to hypoxia. A549 cells were exposed to hypoxia during whole cell patch-clamp measurements. Hypoxia reversibly inhibited a voltage-dependent outward current, consistent with a K current, because tetraethylamonium (TEA; 10 mM) abolished this effect; however, iberiotoxin (0.1 microM) does not. In normoxia, TEA and iberiotoxin inhibited whole cell current (-35%), whereas the K-channel inhibitors glibenclamide (1 microM), barium (1 mM), chromanol B293 (10 microM), and 4-aminopyridine (1 mM) were ineffective. (86)Rb uptake was measured to see whether K-channel modulation also affected transport activity. TEA, iberiotoxin, and 4-h hypoxia (1.5% O(2)) inhibited total (86)Rb uptake by 40, 20, and 35%, respectively. Increased extracellular K also inhibited (86)Rb uptake in a dose-dependent way. The K-channel opener 1-ethyl-2-benzimidazolinone (1 mM) increased (86)Rb uptake by 120% in normoxic and hypoxic cells by activation of Na-K pumps (+60%) and Na-K-2Cl cotransport (+170%). However, hypoxic transport inhibition was also seen in the presence of 1-ethyl-2-benzimidazolinone, TEA, and iberiotoxin. These results indicate that hypoxia, membrane depolarization, and K-channel inhibition decrease whole cell membrane currents and transport activity. It appears, therefore, that a hypoxia-induced change in membrane conductance and membrane potential might be a link between hypoxia and alveolar ion transport inhibition.     

Activated protein C inhibits the release of macrophage inflammatory protein-1-alpha from THP-1 cells and from human monocytes

Several lines of evidence have implicated activated protein C (APC) to be an endogenous inhibitor of the inflammatory septic cascade. APC may exhibit direct anti-inflammatory properties, independent of its antithrombotic effects. Chemokines influence the interaction of monocytes at the endothelium during infection and sepsis and are involved in the molecular events leading to an adverse and lethal outcome of sepsis. Defining regulatory mechanisms on the monocytic release profile of the proinflammatory C-C chemokines macrophage inflammatory protein-1-alpha (MIP-1-alpha) and monocyte chemoattractant protein-1 (MCP-1) might have therapeutic implications for the treatment of sepsis. We established a monocytic cell model of inflammation by the addition of lipopolysaccharide (LPS) and examined the effect of human APC on LPS-stimulated chemokine release from the monocytic cell line THP-1. We found that human APC in supra-physiological concentrations of 2.5-10 microg/ml inhibited the LPS-induced release of the chemokines MIP-1-alpha and MCP-1, as measured by enzyme-linked immunosorbent assays (ELISA) at 6 up to 24 h. In addition to experiments on THP-1 cells, recombinant human APC in concentrations of 50 ng/ml was found to have an inhibiting effect on the release of MIP-1-alpha from freshly isolated mononuclear cells of septic patients. The ability of APC to decrease the release of the C-C chemokine MIP-1-alpha from the monocytic cell line THP-1 and from human monocytes may identify a novel immunomodulatory pathway by which APC exerts its anti-inflammatory action and may contribute to control the inflammatory response in sepsis.