197 research outputs found
Mechanisms of Eph/ephrin mediated cell-cell communication
Eph receptors and their membrane associated ephrin ligands mediate cell-cell repulsion to guide migrating cells and axons. A peculiarity of this signaling system is that both receptors and ligands can transduce signals into the cell resulting in a bidirectional signaling mode. An important step of ephrinB ligand ‘reverse signaling’ is the regulated tyrosine phosphorylation of the cytoplasmic domain, initiating docking sites for downstream signaling adaptors. Moreover, ephrinB ligands can signal by interactions with various PDZ-domain containing proteins.
Using a broad range of in vitro assays the presented work demonstrates that upon binding to their cognate receptor, ephrinB ligands rapidly activate Src family kinases (SFKs) which subsequently phosphorylate ephrinB. Tyrosine phosphorylation appears to be a transient event that is downregulated by the tyrosine phosphatase PTP-BL, which interacts with ephrinB via one of its PDZ-domains. Studies on PTP-BL and another multiple PDZ domain containing protein GRIP (Glutamate Receptor Interacting Protein) revealed that PDZ interactions with ephrinB are also regulated by EphB receptor binding but, unlike tyrosine phosphorylation, these interactions are long lasting. These findings led to postulate a ‘switch model’ for ephrinB reverse signaling: Tyrosine dependent signaling appears to be a rapid and transient event which is later replaced by stable PDZ-dependent signaling.
EphB ‘forward’ signaling as well as ephrinB ‘reverse’ signaling are important for axonal pathfinding and cell migration during development. Prior to their repellent effect on migrating cells and growth cones, Eph receptors form a high affinity complex with their ligands at sites of cell-cell contact. Therefore, mechanisms have to be in place that allow cells to detach from each other permitting retraction and withdrawal. To overcome this adhesive barrier, the ectodomain of ephrinA ligands is cleaved by metalloproteinases and shed upon receptor binding. Intrigued by the previous findings that activated ephrins cluster in cells, we hypothesized that these Eph/ephrin clusters undergo endocytosis. We developed immunofluorescence internalization and co-culture assays to study clustering and endocytosis at cell-cell contact sites. We established an experimental setup to perform fast time lapse imaging studies of cells expressing different fluorescently tagged proteins. Cell contact-induced ephrinB-EphB complexes are rapidly endocytosed during the retraction of cells and neuronal growth cones. Endocytosis occurs in a bidirectional manner, leading to internalized complexes of full length receptor and ligand, a yet rarely observed phenomenon. Signaling inactive mutants of EphB receptors and ephrinB ligands lead to a strong adhesion between cells. Endocytosis is sufficient to convert this adhesion into the detachment of cells. Bidirectional endocytosis is necessary to efficiently promote axon detachment during growth cone collapse mediated by ephrinB ligands. On the cell biological level, bidirectional endocytosis of two full length transmembrane (TM) proteins is a new phenomenon. Moreover, these studies reveal a novel mechanism of signal termination, de-adhesion and promotion of cell repulsion after intercellular (trans) interaction between two TM proteins
Near-infrared diffuse reflectance spectroscopy for discriminating fruit and vegetable products preserved in glass containers
Near-infrared (NIR) diffuse reflectance spectroscopy was used in combination with multivariate analytical methods to discriminate between different fruit and vegetable products preserved in glass containers, which are commonly used as receptacles for the pasteurization of fruit and vegetable products. To investigate the samples in this way, i.e. inside the sealed glass containers, is important for this specific application in a food processing facility. In order to adapt digitalization technologies to the pasteurization process, it is necessary to investigate usually consumed products with suitable sensors and data analytics. NIR spectroscopy in combination with multivariate data analysis is a mighty tool to unravel various issues in food research and industry. Thus, this combination is in the focus of this investigation. It is shown for the first time that the discrimination between five types of preserved food in glass containers is possible by using NIR diffuse reflectance spectroscopy and multivariate data analysis (including discrimination methods). The performance parameters sensitivity, specificity, and efficiency, are determined for every product group and analyzed in a misclassification table. On average, the results show that
95% of ca. 2100 observations are correctly classified with partial least squares discriminant analysis (PLS-DA)
Comparison of GOME-2/MetOp total ozone data with Brewer spectroradiometer data over the Iberian Peninsula
The main objective of this article is to compare the total ozone data from the new Global Ozone Monitoring Experiment instrument (GOME-2/MetOp) with reliable ground-based measurement recorded by five Brewer spectroradiometers in the Iberian Peninsula. In addition, a similar comparison for the predecessor instrument GOME/ERS-2 is described. The period of study is a whole year from May 2007 to April 2008. The results show that GOME-2/MetOp ozone data already has a very good quality, total ozone columns are on average 3.05% lower than Brewer measurements. This underestimation is higher than that obtained for GOME/ERS-2 (1.46%). However, the relative differences between GOME-2/MetOp and Brewer measurements show significantly lower variability than the differences between GOME/ERS-2 and Brewer data. Dependencies of these relative differences with respect to the satellite solar zenith angle (SZA), the satellite scan angle, the satellite cloud cover fraction (CF), and the ground-based total ozone measurements are analyzed. For both GOME instruments, differences show no significant dependence on SZA. However, GOME-2/MetOp data show a significant dependence on the satellite scan angle (+1.5%). In addition, GOME/ERS-2 differences present a clear dependence with respect to the CF and ground-based total ozone; such differences are minimized for GOME-2/MetOp. The comparison between the daily total ozone values provided by both GOME instruments shows that GOME-2/MetOp ozone data are on average 1.46% lower than GOME/ERS-2 data without any seasonal dependence. Finally, deviations of a priori climatological ozone profile used by the satellite retrieval algorithm from the true ozone profile are analyzed. Although excellent agreement between a priori climatological and measured partial ozone values is found for the middle and high stratosphere, relative differences greater than 15% are common for the troposphere and lower stratosphere.This work was partially supported by Ministerio de Educacion y Ciencia under project CGL2005-05693-C03-03/CLI and by Ministerio de Ciencia e Innovacion under project CGL2008-05939-C03-02/CLI
RECEIVED 1 APRIL; ACCEPTED 9 JULY; PUBLISHED ONLINE
The nematode C. elegans is an excellent model organism for studying behavior at the neuronal level. Because of the organism's small size, it is challenging to deliver stimuli to C. elegans and monitor neuronal activity in a controlled environment. To address this problem, we developed two microfluidic chips, the 'behavior' chip and the 'olfactory' chip for imaging of neuronal and behavioral responses in C. elegans. We used the behavior chip to correlate the activity of AVA command interneurons with the worm locomotion pattern. We used the olfactory chip to record responses from ASH sensory neurons exposed to high-osmotic-strength stimulus. Observation of neuronal responses in these devices revealed previously unknown properties of AVA and ASH neurons. The use of these chips can be extended to correlate the activity of sensory neurons, interneurons and motor neurons with the worm's behavior. How neural circuits process information to generate behavior is a fundamental question in neuroscience. To address this question, one should observe an animal in a well-controlled environment, in which a specific behavior can be generated and corresponding neuronal activity monitored. Ideally such an environment should not disturb normal neuronal function and should be able to reveal the specific neuronal circuit under study. C. elegans, with its optically accessible, stereotyped and compact nervous system, has drawn great scientific attention because of its diverse repertoire of behavioral outputs and its genetic conservation with vertebrates. Initial efforts to measure activity in the C. elegans nervous system relied on electrophysiological recordings from single neurons in dissected worms 1 . The recent development of genetically encoded fluorescent calcium indicators 2 has spawned an increasing interest in optical imaging approaches that permit the tracking of calcium transients in individual neurons in vivo in intact worms 3 . Although transgenic worms that express neuron-specific indicators can now routinely be generated, the present methods for confining and stimulating the worm during imaging are not ideal. The typical experimental setup involves application of glue onto specific segments of the worm to achieve permanent immobilization on a hydrated agar pad. Fluid-filled pipettes, temperature-controlled plates and sharp electrodes have been used in the past to deliver chemical, thermal and mechanical stimuli, respectively 4,5 . Whether the organic glue is toxic to the worm and how it influences neuronal activity are difficult to determine. Moreover, the delivery of chemical stimuli to the glued worm cannot be precisely controlled or separated from mechanical stimuli associated with fluid flow. More concerns arise when the circuit controlling locomotion is under study. The glue immobilizes the worm, not allowing muscles and stretch-receptor neurons, if any, to contract and relax normally. This mechanically restricted microenvironment might affect the function of the proprioceptive sensory neurons as well as motor neurons. Most importantly, the glue setup does not permit most behaviors to be generated, visualized, quantified or correlated to neuronal activity in real time. A system with two objectives 6 has been a welcome step toward simultaneous neuronal-behavior analysis, as has been a new system for tracking thermosensory neurons (albeit at low optical resolution) in freely moving worms 7 . Recent advances in microfabrication technology permit the construction of well-controllable microenvironments with applications ranging from cell analysis to tissue engineering RESULTS The behavior chip The first microfluidic device, the behavior chi
Structural Characterization and Lifetimes of Triple‐Stranded Helical Coinage Metal Complexes: Synthesis, Spectroscopy and Quantum Chemical Calculations
This work reports on a series of polynuclear complexes containing a trinuclear Cu, Ag, or Au core in combination with the fac-isomer of the metalloligand [Ru(pypzH)](PF) (pypzH=3-(pyridin-2-yl)pyrazole). These (in case of the Ag and Au containing species) newly synthesized compounds of the general formula [{Ru(pypz)}M](PF) (2: M=Cu; 3: M=Ag; 4: M=Au) contain triple-stranded helical structures in which two ruthenium moieties are connected by three N-M-N (M=Cu, Ag, Au) bridges. In order to obtain a detailed description of the structure both in the electronic ground and excited states, extensive spectroscopic and quantum chemical calculations are applied. The equilateral coinage metal core triangle in the electronic ground state of 2–4 is distorted in the triplet state. Furthermore, the analyses offer a detailed description of electronic excitations. By using time-resolved IR spectroscopy from the microsecond down to the nanosecond regime, both the vibrational spectra and the lifetime of the lowest lying electronically excited triplet state can be determined. The lifetimes of these almost only non-radiative triplet states of 2–4 show an unusual effect in a way that the Au-containing complex 4 has a lifetime which is by more than a factor of five longer than in case of the Cu complex 2. Thus, the coinage metals have a significant effect on the electronically excited state, which is localized on a pypz ligand coordinated to the Ru atom indicating an unusual cooperative effect between two moieties of the complex
Regulation of two motor patterns enables the gradual adjustment of locomotion strategy in Caenorhabditis elegans
In animal locomotion a tradeoff exists between stereotypy and flexibility: fast long-distance travelling (LDT) requires coherent regular motions, while local sampling and area-restricted search (ARS) rely on flexible movements. We report here on a posture control system in C. elegans that coordinates these needs. Using quantitative posture analysis we explain worm locomotion as a composite of two modes: regular undulations versus flexible turning. Graded reciprocal regulation of both modes allows animals to flexibly adapt their locomotion strategy under sensory stimulation along a spectrum ranging from LDT to ARS. Using genetics and functional imaging of neural activity we characterize the counteracting interneurons AVK and DVA that utilize FLP-1 and NLP-12 neuropeptides to control both motor modes. Gradual regulation of behaviors via this system is required for spatial navigation during chemotaxis. This work shows how a nervous system controls simple elementary features of posture to generate complex movements for goal-directed locomotion strategies
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