Termination of second messenger signaling in olfaction.

PubMed Central

Boekhoff, I; Breer, H

1992-01-01

By using isolated rat olfactory cilia and a fast kinetics methodology, it has been demonstrated that odorant-induced second messenger signaling in the millisecond time range is terminated via phosphorylation reactions catalyzed by specific protein kinases. The cyclic adenosine nucleotide pathway is turned off by kinase A activity, whereas the inositol trisphosphate cascade is terminated by kinase C. The data support the concept that desensitization of odorant responses involves phosphorylation of key elements in the transduction cascade. PMID:1370581

Signal transduction in a covalent post-assembly modification cascade

NASA Astrophysics Data System (ADS)

Pilgrim, Ben S.; Roberts, Derrick A.; Lohr, Thorsten G.; Ronson, Tanya K.; Nitschke, Jonathan R.

2017-12-01

Natural reaction cascades control the movement of biomolecules between cellular compartments. Inspired by these systems, we report a synthetic reaction cascade employing post-assembly modification reactions to direct the partitioning of supramolecular complexes between phases. The system is composed of a self-assembled tetrazine-edged FeII8L12 cube and a maleimide-functionalized FeII4L6 tetrahedron. Norbornadiene (NBD) functions as the stimulus that triggers the cascade, beginning with the inverse-electron-demand Diels-Alder reaction of NBD with the tetrazine moieties of the cube. This reaction generates cyclopentadiene as a transient by-product, acting as a relay signal that subsequently undergoes a Diels-Alder reaction with the maleimide-functionalized tetrahedron. Cyclooctyne can selectively inhibit the cascade by outcompeting NBD as the initial trigger. Initiating the cascade with 2-octadecyl NBD leads to selective alkylation of the tetrahedron upon cascade completion. The increased lipophilicity of the C18-tagged tetrahedron drives this complex into a non-polar phase, allowing its isolation from the initially inseparable mixture of complexes.

Single-Molecule Imaging of Cellular Signaling

NASA Astrophysics Data System (ADS)

De Keijzer, Sandra; Snaar-Jagalska, B. Ewa; Spaink, Herman P.; Schmidt, Thomas

Single-molecule microscopy is an emerging technique to understand the function of a protein in the context of its natural environment. In our laboratory this technique has been used to study the dynamics of signal transduction in vivo. A multitude of signal transduction cascades are initiated by interactions between proteins in the plasma membrane. These cascades start by binding a ligand to its receptor, thereby activating downstream signaling pathways which finally result in complex cellular responses. To fully understand these processes it is important to study the initial steps of the signaling cascades. Standard biological assays mostly call for overexpression of the proteins and high concentrations of ligand. This sets severe limits to the interpretation of, for instance, the time-course of the observations, given the large temporal spread caused by the diffusion-limited binding processes. Methods and limitations of single-molecule microscopy for the study of cell signaling are discussed on the example of the chemotactic signaling of the slime-mold Dictyostelium discoideum. Single-molecule studies, as reviewed in this chapter, appear to be one of the essential methodologies for the full spatiotemporal clarification of cellular signaling, one of the ultimate goals in cell biology.

Presence of Ca2+-dependent K+ channels in chemosensory cilia support a role in odor transduction.

PubMed

Delgado, Ricardo; Saavedra, M Veronica; Schmachtenberg, Oliver; Sierralta, Jimena; Bacigalupo, Juan

2003-09-01

Olfactory receptor neurons (ORNs) respond to odorants with changes in the action potential firing rate. Excitatory responses, consisting of firing increases, are mediated by a cyclic AMP cascade that leads to the activation of cationic nonselective cyclic nucleotide-gated (CNG) channels and Ca2+-dependent Cl- (ClCa) channels. This process takes place in the olfactory cilia, where all protein components of this cascade are confined. ORNs from various vertebrate species have also been shown to generate inhibitory odor responses, expressed as decreases in action potential discharges. Odor inhibition appears to rely on Ca2+-dependent K+ (KCa) channels, but the underlying transduction mechanism remains unknown. If these channels are involved in odor transduction, they are expected to be present in the olfactory cilia. We found that a specific antibody against a large conductance KCa recognized a protein of approximately 116 kDa in Western blots of purified rat olfactory ciliary membranes. Moreover, the antibody labeled ORN cilia in isolated ORNs from rat and toad (Caudiverbera caudiverbera). In addition, single-channel recordings from inside-out membrane patches excised from toad chemosensory cilia showed the presence of 4 different types of KCa channels, with unitary conductances of 210, 60, 12, and 29 and 60 pS, high K+-selectivity, and Ca2+ sensitivities in the low micromolar range. Our work demonstrates the presence of K+ channels in the ORN cilia and supports their participation in odor transduction.

Ultrasensitivity in signaling cascades revisited: Linking local and global ultrasensitivity estimations.

PubMed

Altszyler, Edgar; Ventura, Alejandra C; Colman-Lerner, Alejandro; Chernomoretz, Ariel

2017-01-01

Ultrasensitive response motifs, capable of converting graded stimuli into binary responses, are well-conserved in signal transduction networks. Although it has been shown that a cascade arrangement of multiple ultrasensitive modules can enhance the system's ultrasensitivity, how a given combination of layers affects a cascade's ultrasensitivity remains an open question for the general case. Here, we introduce a methodology that allows us to determine the presence of sequestration effects and to quantify the relative contribution of each module to the overall cascade's ultrasensitivity. The proposed analysis framework provides a natural link between global and local ultrasensitivity descriptors and it is particularly well-suited to characterize and understand mathematical models used to study real biological systems. As a case study, we have considered three mathematical models introduced by O'Shaughnessy et al. to study a tunable synthetic MAPK cascade, and we show how our methodology can help modelers better understand alternative models.

A Prize-Collecting Steiner Tree Approach for Transduction Network Inference

NASA Astrophysics Data System (ADS)

Bailly-Bechet, Marc; Braunstein, Alfredo; Zecchina, Riccardo

Into the cell, information from the environment is mainly propagated via signaling pathways which form a transduction network. Here we propose a new algorithm to infer transduction networks from heterogeneous data, using both the protein interaction network and expression datasets. We formulate the inference problem as an optimization task, and develop a message-passing, probabilistic and distributed formalism to solve it. We apply our algorithm to the pheromone response in the baker’s yeast S. cerevisiae. We are able to find the backbone of the known structure of the MAPK cascade of pheromone response, validating our algorithm. More importantly, we make biological predictions about some proteins whose role could be at the interface between pheromone response and other cellular functions.

The ethylene signal transduction pathway in Arabidopsis

NASA Technical Reports Server (NTRS)

Kieber, J. J.; Evans, M. L. (Principal Investigator)

1997-01-01

The gaseous hormone ethylene is an important regulator of plant growth and development. Using a simple response of etiolated seedlings to ethylene as a genetic screen, genes involved in ethylene signal transduction have been identified in Arabidopsis. Analysis of two of these genes that have been cloned reveals that ethylene signalling involves a combination of a protein (ETR1) with similarity to bacterial histidine kinases and a protein (CTR1) with similarity to Raf-1, a protein kinase involved in multiple signalling cascades in eukaryotic cells. Several lines of investigation provide compelling evidence that ETR1 encodes an ethylene receptor. For the first time there is a glimpse of the molecular circuitry underlying the signal transduction pathway for a plant hormone.

Signal transduction and amplification through enzyme-triggered ligand release and accelerated catalysis.

PubMed

Goggins, Sean; Marsh, Barrie J; Lubben, Anneke T; Frost, Christopher G

2015-08-01

Signal transduction and signal amplification are both important mechanisms used within biological signalling pathways. Inspired by this process, we have developed a signal amplification methodology that utilises the selectivity and high activity of enzymes in combination with the robustness and generality of an organometallic catalyst, achieving a hybrid biological and synthetic catalyst cascade. A proligand enzyme substrate was designed to selectively self-immolate in the presence of the enzyme to release a ligand that can bind to a metal pre-catalyst and accelerate the rate of a transfer hydrogenation reaction. Enzyme-triggered catalytic signal amplification was then applied to a range of catalyst substrates demonstrating that signal amplification and signal transduction can both be achieved through this methodology.

Targeted Ablation of the Pde6h Gene in Mice Reveals Cross-species Differences in Cone and Rod Phototransduction Protein Isoform Inventory*

PubMed Central

Brennenstuhl, Christina; Tanimoto, Naoyuki; Burkard, Markus; Wagner, Rebecca; Bolz, Sylvia; Trifunovic, Dragana; Kabagema-Bilan, Clement; Paquet-Durand, Francois; Beck, Susanne C.; Huber, Gesine; Seeliger, Mathias W.; Ruth, Peter; Wissinger, Bernd; Lukowski, Robert

2015-01-01

Phosphodiesterase-6 (PDE6) is a multisubunit enzyme that plays a key role in the visual transduction cascade in rod and cone photoreceptors. Each type of photoreceptor utilizes discrete catalytic and inhibitory PDE6 subunits to fulfill its physiological tasks, i.e. the degradation of cyclic guanosine-3′,5′-monophosphate at specifically tuned rates and kinetics. Recently, the human PDE6H gene was identified as a novel locus for autosomal recessive (incomplete) color blindness. However, the three different classes of cones were not affected to the same extent. Short wave cone function was more preserved than middle and long wave cone function indicating that some basic regulation of the PDE6 multisubunit enzyme was maintained albeit by a unknown mechanism. To study normal and disease-related functions of cone Pde6h in vivo, we generated Pde6h knock-out (Pde6h−/−) mice. Expression of PDE6H in murine eyes was restricted to both outer segments and synaptic terminals of short and long/middle cone photoreceptors, whereas Pde6h−/− retinae remained PDE6H-negative. Combined in vivo assessment of retinal morphology with histomorphological analyses revealed a normal overall integrity of the retinal organization and an unaltered distribution of the different cone photoreceptor subtypes upon Pde6h ablation. In contrast to human patients, our electroretinographic examinations of Pde6h−/− mice suggest no defects in cone/rod-driven retinal signaling and therefore preserved visual functions. To this end, we were able to demonstrate the presence of rod PDE6G in cones indicating functional substitution of PDE6. The disparities between human and murine phenotypes caused by mutant Pde6h/PDE6H suggest species-to-species differences in the vulnerability of biochemical and neurosensory pathways of the visual signal transduction system. PMID:25739440

MicroRNA-133 modulates the β1-adrenergic receptor transduction cascade.

PubMed

Castaldi, Alessandra; Zaglia, Tania; Di Mauro, Vittoria; Carullo, Pierluigi; Viggiani, Giacomo; Borile, Giulia; Di Stefano, Barbara; Schiattarella, Gabriele Giacomo; Gualazzi, Maria Giovanna; Elia, Leonardo; Stirparo, Giuliano Giuseppe; Colorito, Maria Luisa; Pironti, Gianluigi; Kunderfranco, Paolo; Esposito, Giovanni; Bang, Marie-Louise; Mongillo, Marco; Condorelli, Gianluigi; Catalucci, Daniele

2014-07-07

The sympathetic nervous system plays a fundamental role in the regulation of myocardial function. During chronic pressure overload, overactivation of the sympathetic nervous system induces the release of catecholamines, which activate β-adrenergic receptors in cardiomyocytes and lead to increased heart rate and cardiac contractility. However, chronic stimulation of β-adrenergic receptors leads to impaired cardiac function, and β-blockers are widely used as therapeutic agents for the treatment of cardiac disease. MicroRNA-133 (miR-133) is highly expressed in the myocardium and is involved in controlling cardiac function through regulation of messenger RNA translation/stability. To determine whether miR-133 affects β-adrenergic receptor signaling during progression to heart failure. Based on bioinformatic analysis, β1-adrenergic receptor (β1AR) and other components of the β1AR signal transduction cascade, including adenylate cyclase VI and the catalytic subunit of the cAMP-dependent protein kinase A, were predicted as direct targets of miR-133 and subsequently validated by experimental studies. Consistently, cAMP accumulation and activation of downstream targets were repressed by miR-133 overexpression in both neonatal and adult cardiomyocytes following selective β1AR stimulation. Furthermore, gain-of-function and loss-of-function studies of miR-133 revealed its role in counteracting the deleterious apoptotic effects caused by chronic β1AR stimulation. This was confirmed in vivo using a novel cardiac-specific TetON-miR-133 inducible transgenic mouse model. When subjected to transaortic constriction, TetON-miR-133 inducible transgenic mice maintained cardiac performance and showed attenuated apoptosis and reduced fibrosis compared with control mice. miR-133 controls multiple components of the β1AR transduction cascade and is cardioprotective during heart failure. © 2014 American Heart Association, Inc.

Transmembrane signaling in Saccharomyces cerevisiae as a model for signaling in metazoans: state of the art after 25 years.

PubMed

Engelberg, David; Perlman, Riki; Levitzki, Alexander

2014-12-01

In the very first article that appeared in Cellular Signalling, published in its inaugural issue in October 1989, we reviewed signal transduction pathways in Saccharomyces cerevisiae. Although this yeast was already a powerful model organism for the study of cellular processes, it was not yet a valuable instrument for the investigation of signaling cascades. In 1989, therefore, we discussed only two pathways, the Ras/cAMP and the mating (Fus3) signaling cascades. The pivotal findings concerning those pathways undoubtedly contributed to the realization that yeast is a relevant model for understanding signal transduction in higher eukaryotes. Consequently, the last 25 years have witnessed the discovery of many signal transduction pathways in S. cerevisiae, including the high osmotic glycerol (Hog1), Stl2/Mpk1 and Smk1 mitogen-activated protein (MAP) kinase pathways, the TOR, AMPK/Snf1, SPS, PLC1 and Pkr/Gcn2 cascades, and systems that sense and respond to various types of stress. For many cascades, orthologous pathways were identified in mammals following their discovery in yeast. Here we review advances in the understanding of signaling in S. cerevisiae over the last 25 years. When all pathways are analyzed together, some prominent themes emerge. First, wiring of signaling cascades may not be identical in all S. cerevisiae strains, but is probably specific to each genetic background. This situation complicates attempts to decipher and generalize these webs of reactions. Secondly, the Ras/cAMP and the TOR cascades are pivotal pathways that affect all processes of the life of the yeast cell, whereas the yeast MAP kinase pathways are not essential. Yeast cells deficient in all MAP kinases proliferate normally. Another theme is the existence of central molecular hubs, either as single proteins (e.g., Msn2/4, Flo11) or as multisubunit complexes (e.g., TORC1/2), which are controlled by numerous pathways and in turn determine the fate of the cell. It is also apparent that lipid signaling is less developed in yeast than in higher eukaryotes. Finally, feedback regulatory mechanisms seem to be at least as important and powerful as the pathways themselves. In the final chapter of this essay we dare to imagine the essence of our next review on signaling in yeast, to be published on the 50th anniversary of Cellular Signalling in 2039. Copyright © 2014 Elsevier Inc. All rights reserved.

Therapeutic peptides for cancer therapy. Part I - peptide inhibitors of signal transduction cascades.

PubMed

Bidwell, Gene L; Raucher, Drazen

2009-10-01

Therapeutic peptides have great potential as anticancer agents owing to their ease of rational design and target specificity. However, their utility in vivo is limited by low stability and poor tumor penetration. The authors review the development of peptide inhibitors with potential for cancer therapy. Peptides that inhibit signal transduction cascades are discussed. The authors searched Medline for articles concerning the development of therapeutic peptides and their delivery. Given our current knowledge of protein sequences, structures and interaction interfaces, therapeutic peptides that inhibit interactions of interest are easily designed. These peptides are advantageous because they are highly specific for the interaction of interest, and they are much more easily developed than small molecule inhibitors of the same interactions. The main hurdle to application of peptides for cancer therapy is their poor pharmacokinetic and biodistribution parameters. Therefore, successful development of peptide delivery vectors could potentially make possible the use of this new and very promising class of anticancer agents.

Signalling crosstalk in plants: emerging issues.

PubMed

Taylor, Jane E; McAinsh, Martin R

2004-01-01

The Oxford English Dictionary defines crosstalk as 'unwanted transfer of signals between communication channels'. How does this definition relate to the way in which we view the organization and function of signalling pathways? Recent advances in the field of plant signalling have challenged the traditional view of a signalling transduction cascade as isolated linear pathways. Instead the picture emerging of the mechanisms by which plants transduce environmental signals is of the interaction between transduction chains. The manner in which these interactions occur (and indeed whether the transfer of these signals is 'unwanted' or beneficial) is currently the topic of intense research.

Dissecting the determinants of light sensitivity in amphioxus microvillar photoreceptors: possible evolutionary implications for melanopsin signaling.

PubMed

Ferrer, Camilo; Malagón, Gerardo; Gomez, María Del Pilar; Nasi, Enrico

2012-12-12

Melanopsin, a photopigment related to the rhodopsin of microvillar photoreceptors of invertebrates, evolved in vertebrates to subserve nonvisual light-sensing functions, such as the pupillary reflex and entrainment of circadian rhythms. However, vertebrate circadian receptors display no hint of a microvillar specialization and show an extremely low light sensitivity and sluggish kinetics. Recently in amphioxus, the most basal chordate, melanopsin-expressing photoreceptors were characterized; these cells share salient properties with both rhabdomeric photoreceptors of invertebrates and circadian receptors of vertebrates. We used electrophysiology to dissect the gain of the light-transduction process in amphioxus and examine key features that help outline the evolutionary transition toward a sensor optimized to report mean ambient illumination rather than mediating spatial vision. By comparing the size of current fluctuations attributable to single photon melanopsin isomerizations with the size of single-channels activated by light, we concluded that the gain of the transduction cascade is lower than in rhabdomeric receptors. In contrast, the expression level of melanopsin (gauged by measuring charge displacements during photo-induced melanopsin isomerization) is comparable with that of canonical visual receptors. A modest amplification in melanopsin-using receptors is therefore apparent in early chordates; the decrease in photopigment expression-and loss of the anatomical correlates-observed in vertebrates subsequently enabled them to attain the low photosensitivity tailored to the role of circadian receptors.

Spatial modeling of the membrane-cytosolic interface in protein kinase signal transduction

PubMed Central

Schröder, Andreas

2018-01-01

The spatial architecture of signaling pathways and the interaction with cell size and morphology are complex, but little understood. With the advances of single cell imaging and single cell biology, it becomes crucial to understand intracellular processes in time and space. Activation of cell surface receptors often triggers a signaling cascade including the activation of membrane-attached and cytosolic signaling components, which eventually transmit the signal to the cell nucleus. Signaling proteins can form steep gradients in the cytosol, which cause strong cell size dependence. We show that the kinetics at the membrane-cytosolic interface and the ratio of cell membrane area to the enclosed cytosolic volume change the behavior of signaling cascades significantly. We suggest an estimate of average concentration for arbitrary cell shapes depending on the cell volume and cell surface area. The normalized variance, known from image analysis, is suggested as an alternative measure to quantify the deviation from the average concentration. A mathematical analysis of signal transduction in time and space is presented, providing analytical solutions for different spatial arrangements of linear signaling cascades. Quantification of signaling time scales reveals that signal propagation is faster at the membrane than at the nucleus, while this time difference decreases with the number of signaling components in the cytosol. Our investigations are complemented by numerical simulations of non-linear cascades with feedback and asymmetric cell shapes. We conclude that intracellular signal propagation is highly dependent on cell geometry and, thereby, conveys information on cell size and shape to the nucleus. PMID:29630597

Aluminum stress and its role in the phospholipid signaling pathway in plants and possible biotechnological applications.

PubMed

Poot-Poot, Wilberth; Hernandez-Sotomayor, Soledad M Teresa

2011-10-01

An early response of plants to environmental signals or abiotic stress suggests that the phospholipid signaling pathway plays a pivotal role in these mechanisms. The phospholipid signaling cascade is one of the main systems of cellular transduction and is related to other signal transduction mechanisms. These other mechanisms include the generation of second messengers and their interactions with various proteins, such as ion channels. This phospholipid signaling cascade is activated by changes in the environment, such as phosphate starvation, water, metals, saline stres, and plant-pathogen interactions. One important factor that impacts agricultural crops is metal-induced stress. Because aluminum has been considered to be a major toxic factor for agriculture conducted in acidic soils, many researchers have focused on understanding the mechanisms of aluminum toxicity in plants. We have contributed the last fifteen years in this field by studying the effects of aluminum on phospholipid signaling in coffee, one of the Mexico's primary crops. We have focused our research on aluminum toxicity mechanisms in Coffea arabica suspension cells as a model for developing future contributions to the biotechnological transformation of coffee crops such that they can be made resistant to aluminum toxicity. We conclude that aluminum is able to not only generate a signal cascade in plants but also modulate other signal cascades generated by other types of stress in plants. The aim of this review is to discuss possible involvement of the phospholipid signaling pathway in the aluminum toxicity response of plant cells. Copyright © 2011 Wiley Periodicals, Inc.

Piracy of Decay-Accelerating Factor (CD55) Signal Transduction by the Diffusely Adhering Strain Escherichia coli C1845 Promotes Cytoskeletal F-Actin Rearrangements in Cultured Human Intestinal INT407 Cells

PubMed Central

Peiffer, Isabelle; Servin, Alain L.; Bernet-Camard, Marie-Françoise

1998-01-01

Diffusely adhering Escherichia coli (DAEC) C1845 (clinical isolate) harboring the fimbrial adhesin F1845 can infect cultured human differentiated intestinal epithelial cells; this process is followed by the disassembly of the actin network in the apical domain. The aim of this study was to examine the mechanism by which DAEC C1845 promotes F-actin rearrangements. For this purpose, we used a human embryonic intestinal cell line (INT407) expressing the membrane-associated glycosylphosphatidylinositol (GPI) protein-anchored decay-accelerating factor (DAF), the receptor of the F1845 adhesin. We show here that infection of INT407 cells by DAEC C1845 can provoke dramatic F-actin rearrangements without cell entry. Clustering of phosphotyrosines was observed, revealing that the DAEC C1845-DAF interaction involves the recruitment of signal transduction molecules. A pharmacological approach with a subset of inhibitors of signal transduction molecules was used to identify the cascade of signal transduction molecules that are coupled to the DAF, that are activated upon infection, and that promote the F-actin rearrangements. DAEC C1845-induced F-actin rearrangements can be blocked dose dependently by protein tyrosine kinase, phospholipase Cγ, phosphatidylinositol 3-kinase, protein kinase C, and Ca2+ inhibitors. F-actin rearrangements and blocking by inhibitors were observed after infection of the cells with two E. coli recombinants carrying the plasmids containing the fimbrial adhesin F1845 or the fimbrial hemagglutinin Dr, belonging to the same family of adhesins. These findings show that the DAEC Dr family of pathogens promotes alterations in the intestinal cell cytoskeleton by piracy of the DAF-GPI signal cascade without bacterial cell entry. PMID:9712744

[Signal transduction mechanisms of hormones through membrane receptors].

PubMed

Yasufuku-Takano, Junko; Takano, Koji

2002-02-01

Hormones exert their effect on cells either via membrane receptors or intracellular receptors. This paper aims to review membrane receptors and the intracellular signal transduction mechanisms. Membrane receptors could be classified according to their structural characteristics and the way they initiate the intracellular signal transduction. These include 1) Seven transmembrane(or G-protein coupled) receptors--heterotrimeric G-proteins--effector, system, 2) Receptor tyrosine kinases--protein-protein interaction through SH2, SH3, and PTB domain--MAP kinase cascades and PI3-kinase pathways, 3) Cytokine receptors--JAK--STAT pathways, 4) Receptors of the TGF- beta superfamily--SMAD pathways, 5) Apoptosis-related receptors--caspase pathways, and 6) ligand-gated ion channels. There are growing knowledge of cross-talks between these pathways. It is being recognized that steroid hormones have distinct membrane receptors, which mediate rapid, nongenomic effect.

The styryl dye FM1-43 suppresses odorant responses in a subset of olfactory neurons by blocking cyclic nucleotide-gated (CNG) channels.

PubMed

Breunig, Esther; Kludt, Eugen; Czesnik, Dirk; Schild, Detlev

2011-08-12

Many olfactory receptor neurons use a cAMP-dependent transduction mechanism to transduce odorants into depolarizations. This signaling cascade is characterized by a sequence of two currents: a cation current through cyclic nucleotide-gated channels followed by a chloride current through calcium-activated chloride channels. To date, it is not possible to interfere with these generator channels under physiological conditions with potent and specific blockers. In this study we identified the styryl dye FM1-43 as a potent blocker of native olfactory cyclic nucleotide-gated channels. Furthermore, we characterized this substance to stain olfactory receptor neurons that are endowed with cAMP-dependent transduction. This allows optical differentiation and pharmacological interference with olfactory receptor neurons at the level of the signal transduction.

Blocking adenylyl cyclase inhibits olfactory generator currents induced by "IP(3)-odors".

PubMed

Chen, S; Lane, A P; Bock, R; Leinders-Zufall, T; Zufall, F

2000-07-01

Vertebrate olfactory receptor neurons (ORNs) transduce odor stimuli into electrical signals by means of an adenylyl cyclase/cAMP second messenger cascade, but it remains widely debated whether this cAMP cascade mediates transduction for all odorants or only certain odor classes. To address this problem, we have analyzed the generator currents induced by odors that failed to produce cAMP in previous biochemical assays but instead produced IP(3) ("IP(3)-odors"). We show that in single salamander ORNs, sensory responses to "cAMP-odors" and IP(3)-odors are not mutually exclusive but coexist in the same cells. The currents induced by IP(3)-odors exhibit identical biophysical properties as those induced by cAMP odors or direct activation of the cAMP cascade. By disrupting adenylyl cyclase to block cAMP formation using two potent antagonists of adenylyl cyclase, SQ22536 and MDL12330A, we show that this molecular step is necessary for the transduction of both odor classes. To assess whether these results are also applicable to mammals, we examine the electrophysiological responses to IP(3)-odors in intact mouse main olfactory epithelium (MOE) by recording field potentials. The results show that inhibition of adenylyl cyclase prevents EOG responses to both odor classes in mouse MOE, even when "hot spots" with heightened sensitivity to IP(3)-odors are examined.

Gqalpha-linked PLCbeta and PLCgamma are essential components of the pheromone biosynthesis activating neuropeptide (PBAN) signal transduction cascade

USDA-ARS?s Scientific Manuscript database

Sex pheromone production for most moths is regulated by pheromone biosynthesis activating neuropeptide (PBAN). In Bombyx mori, PBAN binding triggers the opening of store-operated Ca2+ channels, suggesting the involvement of a receptor-activated phospholipase C (PLC). In this study, we found that P...

Calcium signaling during the plant-plant interaction of parasitic Cuscuta reflexa with its hosts.

PubMed

Albert, Markus; Kaiser, Bettina; van der Krol, Sander; Kaldenhoff, Ralf

2010-09-01

The plant parasite Cuscuta reflexa induces various responses in compatible and incompatible host plants. The visual reactions of both types of host plants including obvious morphological changes require the recognition of Cuscuta ssp. A consequently initiated signaling cascade is triggered which leads to a tolerance of the infection or, in the case of some incompatible host plants, to resistance. Calcium (Ca(2+)) release is the major second messenger during signal transduction. Therefore, we have studied Ca(2+) spiking in tomato and tobacco during infection with C. reflexa. In our recently published study Ca(2+) signals were monitored as bioluminescence in aequorin-expressing tomato plants after the onset of C. reflexa infestation. Signals at the attachment sites were observed from 30 to 48 h after infection. In an assay with leaf disks of aequorin-expressing tomato which were treated with different C. reflexa plant extracts it turned out that the substance that induced Ca(2+) release in the host plant was closely linked to the parasite's haustoria.

Calcium signaling during the plant-plant interaction of parasitic Cuscuta reflexa with its hosts

PubMed Central

Kaiser, Bettina; van der Krol, Sander; Kaldenhoff, Ralf

2010-01-01

The plant parasite Cuscuta reflexa induces various responses in compatible and incompatible host plants. The visual reactions of both types of host plants including obvious morphological changes require the recognition of Cuscuta ssp. A consequently initiated signaling cascade is triggered which leads to a tolerance of the infection or, in the case of some incompatible host plants, to resistance. Calcium (Ca2+) release is the major second messenger during signal transduction. Therefore, we have studied Ca2+ spiking in tomato and tobacco during infection with C. reflexa. In our recently published study1 Ca2+ signals were monitored as bioluminescence in aequorin-expressing tomato plants after the onset of C. reflexa infestation. Signals at the attachment sites were observed from 30 to 48 h after infection. In an assay with leaf disks of aequorin-expressing tomato which were treated with different C. reflexa plant extracts it turned out that the substance that induced Ca2+ release in the host plant was closely linked to the parasite's haustoria. PMID:20818172

Expression of taste receptors in Solitary Chemosensory Cells of rodent airways

PubMed Central

2011-01-01

Background Chemical irritation of airway mucosa elicits a variety of reflex responses such as coughing, apnea, and laryngeal closure. Inhaled irritants can activate either chemosensitive free nerve endings, laryngeal taste buds or solitary chemosensory cells (SCCs). The SCC population lies in the nasal respiratory epithelium, vomeronasal organ, and larynx, as well as deeper in the airway. The objective of this study is to map the distribution of SCCs within the airways and to determine the elements of the chemosensory transduction cascade expressed in these SCCs. Methods We utilized a combination of immunohistochemistry and molecular techniques (rtPCR and in situ hybridization) on rats and transgenic mice where the Tas1R3 or TRPM5 promoter drives expression of green fluorescent protein (GFP). Results Epithelial SCCs specialized for chemoreception are distributed throughout much of the respiratory tree of rodents. These cells express elements of the taste transduction cascade, including Tas1R and Tas2R receptor molecules, α-gustducin, PLCβ2 and TrpM5. The Tas2R bitter taste receptors are present throughout the entire respiratory tract. In contrast, the Tas1R sweet/umami taste receptors are expressed by numerous SCCs in the nasal cavity, but decrease in prevalence in the trachea, and are absent in the lower airways. Conclusions Elements of the taste transduction cascade including taste receptors are expressed by SCCs distributed throughout the airways. In the nasal cavity, SCCs, expressing Tas1R and Tas2R taste receptors, mediate detection of irritants and foreign substances which trigger trigeminally-mediated protective airway reflexes. Lower in the respiratory tract, similar chemosensory cells are not related to the trigeminal nerve but may still trigger local epithelial responses to irritants. In total, SCCs should be considered chemoreceptor cells that help in preventing damage to the respiratory tract caused by inhaled irritants and pathogens. PMID:21232137

Expression of taste receptors in solitary chemosensory cells of rodent airways.

PubMed

Tizzano, Marco; Cristofoletti, Mirko; Sbarbati, Andrea; Finger, Thomas E

2011-01-13

Chemical irritation of airway mucosa elicits a variety of reflex responses such as coughing, apnea, and laryngeal closure. Inhaled irritants can activate either chemosensitive free nerve endings, laryngeal taste buds or solitary chemosensory cells (SCCs). The SCC population lies in the nasal respiratory epithelium, vomeronasal organ, and larynx, as well as deeper in the airway. The objective of this study is to map the distribution of SCCs within the airways and to determine the elements of the chemosensory transduction cascade expressed in these SCCs. We utilized a combination of immunohistochemistry and molecular techniques (rtPCR and in situ hybridization) on rats and transgenic mice where the Tas1R3 or TRPM5 promoter drives expression of green fluorescent protein (GFP). Epithelial SCCs specialized for chemoreception are distributed throughout much of the respiratory tree of rodents. These cells express elements of the taste transduction cascade, including Tas1R and Tas2R receptor molecules, α-gustducin, PLCβ2 and TrpM5. The Tas2R bitter taste receptors are present throughout the entire respiratory tract. In contrast, the Tas1R sweet/umami taste receptors are expressed by numerous SCCs in the nasal cavity, but decrease in prevalence in the trachea, and are absent in the lower airways. Elements of the taste transduction cascade including taste receptors are expressed by SCCs distributed throughout the airways. In the nasal cavity, SCCs, expressing Tas1R and Tas2R taste receptors, mediate detection of irritants and foreign substances which trigger trigeminally-mediated protective airway reflexes. Lower in the respiratory tract, similar chemosensory cells are not related to the trigeminal nerve but may still trigger local epithelial responses to irritants. In total, SCCs should be considered chemoreceptor cells that help in preventing damage to the respiratory tract caused by inhaled irritants and pathogens.

Cascade exciton-pumping engines with manipulated speed and efficiency in light-harvesting porous π-network films

PubMed Central

Gu, Cheng; Huang, Ning; Xu, Fei; Gao, Jia; Jiang, Donglin

2015-01-01

Light-harvesting antennae are the machinery for exciton pumping in natural photosynthesis, whereas cascade energy transfer through chlorophyll is key to long-distance, efficient energy transduction. Numerous artificial antennae have been developed. However, they are limited in their cascade energy-transfer abilities because of a lack of control over complex chromophore aggregation processes, which has impeded their advancement. Here we report a viable approach for addressing this issue by using a light-harvesting porous polymer film in which a three-dimensional π-network serves as the antenna and micropores segregate multiple dyes to prevent aggregation. Cascade energy-transfer engines are integrated into the films; the rate and efficiency of the energy-funneling engines are precisely manipulated by tailoring the dye components and contents. The nanofilms allow accurate and versatile luminescence engineering, resulting in the production of thirty emission hues, including blue, green, red and white. This advance may open new pathways for realising photosynthesis and photoenergy conversion. PMID:25746459

Activation of the yeast Hippo pathway by phosphorylation-dependent assembly of signaling complexes.

PubMed

Rock, Jeremy M; Lim, Daniel; Stach, Lasse; Ogrodowicz, Roksana W; Keck, Jamie M; Jones, Michele H; Wong, Catherine C L; Yates, John R; Winey, Mark; Smerdon, Stephen J; Yaffe, Michael B; Amon, Angelika

2013-05-17

Scaffold-assisted signaling cascades guide cellular decision-making. In budding yeast, one such signal transduction pathway called the mitotic exit network (MEN) governs the transition from mitosis to the G1 phase of the cell cycle. The MEN is conserved and in metazoans is known as the Hippo tumor-suppressor pathway. We found that signaling through the MEN kinase cascade was mediated by an unusual two-step process. The MEN kinase Cdc15 first phosphorylated the scaffold Nud1. This created a phospho-docking site on Nud1, to which the effector kinase complex Dbf2-Mob1 bound through a phosphoserine-threonine binding domain, in order to be activated by Cdc15. This mechanism of pathway activation has implications for signal transmission through other kinase cascades and might represent a general principle in scaffold-assisted signaling.

Molecular and functional identification of a novel photopigment in Pecten ciliary photoreceptors.

PubMed

Arenas, Oscar; Osorno, Tomás; Malagón, Gerardo; Pulido, Camila; Gomez, María Del Pilar; Nasi, Enrico

2018-01-26

The two basic animal photoreceptor types, ciliary and microvillar, use different light-transduction schemes: their photopigments couple to G t versus G q proteins, respectively, to either mobilize cyclic nucleotides or trigger a lipid signaling cascade. A third class of photoreceptors has been described in the dual retina of some marine invertebrates; these present a ciliary morphology but operate via radically divergent mechanisms, prompting the suggestion that they comprise a novel lineage of light sensors. In one of these organisms, an uncommon putative opsin was uncovered that was proposed to signal through G o Orthologues subsequently emerged in diverse phyla, including mollusks, echinoderms, and chordates, but the cells in which they express have not been identified, and no studies corroborated their function as visual pigments or their suggested signaling mode. Conversely, in only one invertebrate species, Pecten irradians , have the ciliary photoreceptors been physiologically characterized, but their photopigment has not been identified molecularly. We used the transcriptome of Pecten retina to guide the cloning by polymerase chain reaction (PCR) and rapid amplification of cDNA ends (RACE) extensions of a new member of this group of putative opsins. In situ hybridization shows selective transcription in the distal retina, and specific antibodies identify a single band of the expected molecular mass in Western blots and distinctly label ciliary photoreceptors in retina sections. RNA interference knockdown resulted in a reduction in the early receptor current-the first manifestation of light transduction-and prevented the prolonged aftercurrent, which requires a large buildup of activated rhodopsin. We also obtained a full-length clone of the α-subunit of a G o from Pecten retina complementary DNA and localized it by in situ hybridization to the distal photoreceptors. Small interfering RNA targeting this G o caused a specific depression of the photocurrent. These results establish this novel putative opsin as a bona fide visual pigment that couples to G o to convey the light signal. © 2018 Arenas et al.

The Antarctic Krill Euphausia superba Shows Diurnal Cycles of Transcription under Natural Conditions

PubMed Central

Albiero, Alessandro; Sales, Gabriele; Millino, Caterina; Mazzotta, Gabriella M.; Bertolucci, Cristiano; Costa, Rodolfo

2013-01-01

Background Polar environments are characterized by extreme seasonal changes in day length, light intensity and spectrum, the extent of sea ice during the winter, and food availability. A key species of the Southern Ocean ecosystem, the Antarctic krill (Euphausia superba) has evolved rhythmic physiological and behavioral mechanisms to adapt to daily and seasonal changes. The molecular organization of the clockwork underlying these biological rhythms is, nevertheless, still only partially understood. Methodology/Principal Findings The genome sequence of the Antarctic krill is not yet available. A normalized cDNA library was produced and pyrosequenced in the attempt to identify large numbers of transcripts. All available E. superba sequences were then assembled to create the most complete existing oligonucleotide microarray platform with a total of 32,217 probes. Gene expression signatures of specimens collected in the Ross Sea at five different time points over a 24-hour cycle were defined, and 1,308 genes differentially expressed were identified. Of the corresponding transcripts, 609 showed a significant sinusoidal expression pattern; about 40% of these exibithed a 24-hour periodicity while the other 60% was characterized by a shorter (about 12-hour) rhythm. We assigned the differentially expressed genes to functional categories and noticed that those concerning translation, proteolysis, energy and metabolic process, redox regulation, visual transduction and stress response, which are most likely related to daily environmental changes, were significantly enriched. Two transcripts of peroxiredoxin, thought to represent the ancestral timekeeping system that evolved about 2.5 billion years ago, were also identified as were two isoforms of the EsRh1 opsin and two novel arrestin1 sequences involved in the visual transduction cascade. Conclusions Our work represents the first characterization of the krill diurnal transcriptome under natural conditions and provides a first insight into the genetic regulation of physiological changes, which occur around the clock during an Antarctic summer day. PMID:23874706

Cellular Signaling Networks Function as Generalized Wiener-Kolmogorov Filters to Suppress Noise

NASA Astrophysics Data System (ADS)

Hinczewski, Michael; Thirumalai, D.

2014-10-01

Cellular signaling involves the transmission of environmental information through cascades of stochastic biochemical reactions, inevitably introducing noise that compromises signal fidelity. Each stage of the cascade often takes the form of a kinase-phosphatase push-pull network, a basic unit of signaling pathways whose malfunction is linked with a host of cancers. We show that this ubiquitous enzymatic network motif effectively behaves as a Wiener-Kolmogorov optimal noise filter. Using concepts from umbral calculus, we generalize the linear Wiener-Kolmogorov theory, originally introduced in the context of communication and control engineering, to take nonlinear signal transduction and discrete molecule populations into account. This allows us to derive rigorous constraints for efficient noise reduction in this biochemical system. Our mathematical formalism yields bounds on filter performance in cases important to cellular function—such as ultrasensitive response to stimuli. We highlight features of the system relevant for optimizing filter efficiency, encoded in a single, measurable, dimensionless parameter. Our theory, which describes noise control in a large class of signal transduction networks, is also useful both for the design of synthetic biochemical signaling pathways and the manipulation of pathways through experimental probes such as oscillatory input.

Bitter triggers acetylcholine release from polymodal urethral chemosensory cells and bladder reflexes.

PubMed

Deckmann, Klaus; Filipski, Katharina; Krasteva-Christ, Gabriela; Fronius, Martin; Althaus, Mike; Rafiq, Amir; Papadakis, Tamara; Renno, Liane; Jurastow, Innokentij; Wessels, Lars; Wolff, Miriam; Schütz, Burkhard; Weihe, Eberhard; Chubanov, Vladimir; Gudermann, Thomas; Klein, Jochen; Bschleipfer, Thomas; Kummer, Wolfgang

2014-06-03

Chemosensory cells in the mucosal surface of the respiratory tract ("brush cells") use the canonical taste transduction cascade to detect potentially hazardous content and trigger local protective and aversive respiratory reflexes on stimulation. So far, the urogenital tract has been considered to lack this cell type. Here we report the presence of a previously unidentified cholinergic, polymodal chemosensory cell in the mammalian urethra, the potential portal of entry for bacteria and harmful substances into the urogenital system, but not in further centrally located parts of the urinary tract, such as the bladder, ureter, and renal pelvis. Urethral brush cells express bitter and umami taste receptors and downstream components of the taste transduction cascade; respond to stimulation with bitter (denatonium), umami (monosodium glutamate), and uropathogenic Escherichia coli; and release acetylcholine to communicate with other cells. They are approached by sensory nerve fibers expressing nicotinic acetylcholine receptors, and intraurethral application of denatonium reflexively increases activity of the bladder detrusor muscle in anesthetized rats. We propose a concept of urinary bladder control involving a previously unidentified cholinergic chemosensory cell monitoring the chemical composition of the urethral luminal microenvironment for potential hazardous content.

Ultrasensitivity in signaling cascades revisited: Linking local and global ultrasensitivity estimations

PubMed Central

Altszyler, Edgar; Ventura, Alejandra C.; Colman-Lerner, Alejandro; Chernomoretz, Ariel

2017-01-01

Ultrasensitive response motifs, capable of converting graded stimuli into binary responses, are well-conserved in signal transduction networks. Although it has been shown that a cascade arrangement of multiple ultrasensitive modules can enhance the system’s ultrasensitivity, how a given combination of layers affects a cascade’s ultrasensitivity remains an open question for the general case. Here, we introduce a methodology that allows us to determine the presence of sequestration effects and to quantify the relative contribution of each module to the overall cascade’s ultrasensitivity. The proposed analysis framework provides a natural link between global and local ultrasensitivity descriptors and it is particularly well-suited to characterize and understand mathematical models used to study real biological systems. As a case study, we have considered three mathematical models introduced by O’Shaughnessy et al. to study a tunable synthetic MAPK cascade, and we show how our methodology can help modelers better understand alternative models. PMID:28662096

Ras regulates assembly of mitogenic signalling complexes through the effector protein IMP.

PubMed

Matheny, Sharon A; Chen, Chiyuan; Kortum, Robert L; Razidlo, Gina L; Lewis, Robert E; White, Michael A

2004-01-15

The signal transduction cascade comprising Raf, mitogen-activated protein (MAP) kinase kinase (MEK) and MAP kinase is a Ras effector pathway that mediates diverse cellular responses to environmental cues and contributes to Ras-dependent oncogenic transformation. Here we report that the Ras effector protein Impedes Mitogenic signal Propagation (IMP) modulates sensitivity of the MAP kinase cascade to stimulus-dependent activation by limiting functional assembly of the core enzymatic components through the inactivation of KSR, a scaffold/adaptor protein that couples activated Raf to its substrate MEK. IMP is a Ras-responsive E3 ubiquitin ligase that, on activation of Ras, is modified by auto-polyubiquitination, which releases the inhibition of Raf-MEK complex formation. Thus, Ras activates the MAP kinase cascade through simultaneous dual effector interactions: induction of Raf kinase activity and derepression of Raf-MEK complex formation. IMP depletion results in increased stimulus-dependent MEK activation without alterations in the timing or duration of the response. These observations suggest that IMP functions as a threshold modulator, controlling sensitivity of the cascade to stimulus and providing a mechanism to allow adaptive behaviour of the cascade in chronic or complex signalling environments.

Cellular Prion Protein and Caveolin-1 Interaction in a Neuronal Cell Line Precedes Fyn/Erk 1/2 Signal Transduction

PubMed Central

Toni, Mattia; Spisni, Enzo; Griffoni, Cristiana; Santi, Spartaco; Riccio, Massimo; Lenaz, Patrizia; Tomasi, Vittorio

2006-01-01

It has been reported that cellular prion protein (PrPc) is enriched in caveolae or caveolae-like domains with caveolin-1 (Cav-1) participating to signal transduction events by Fyn kinase recruitment. By using the Glutathione-S-transferase (GST)-fusion proteins assay, we observed that PrPc strongly interacts in vitro with Cav-1. Thus, we ascertained the PrPc caveolar localization in a hypothalamic neuronal cell line (GN11), by confocal microscopy analysis, flotation on density gradient, and coimmunoprecipitation experiments. Following the anti-PrPc antibody-mediated stimulation of live GN11 cells, we observed that PrPc clustered on plasma membrane domains rich in Cav-1 in which Fyn kinase converged to be activated. After these events, a signaling cascade through p42/44 MAP kinase (Erk 1/2) was triggered, suggesting that following translocations from rafts to caveolae or caveolaelike domains PrPc could interact with Cav-1 and induce signal transduction events. PMID:17489019

The Attention Cascade Model and Attentional Blink

ERIC Educational Resources Information Center

Shih, Shui-I

2008-01-01

An attention cascade model is proposed to account for attentional blinks in rapid serial visual presentation (RSVP) of stimuli. Data were collected using single characters in a single RSVP stream at 10 Hz [Shih, S., & Reeves, A. (2007). "Attentional capture in rapid serial visual presentation." "Spatial Vision", 20(4), 301-315], and single words,…

Microscopic insight into thermodynamics of conformational changes of SAP-SLAM complex in signal transduction cascade

NASA Astrophysics Data System (ADS)

Samanta, Sudipta; Mukherjee, Sanchita

2017-04-01

The signalling lymphocytic activation molecule (SLAM) family of receptors, expressed by an array of immune cells, associate with SLAM-associated protein (SAP)-related molecules, composed of single SH2 domain architecture. SAP activates Src-family kinase Fyn after SLAM ligation, resulting in a SLAM-SAP-Fyn complex, where, SAP binds the Fyn SH3 domain that does not involve canonical SH3 or SH2 interactions. This demands insight into this SAP mediated signalling cascade. Thermodynamics of the conformational changes are extracted from the histograms of dihedral angles obtained from the all-atom molecular dynamics simulations of this structurally well characterized SAP-SLAM complex. The results incorporate the binding induced thermodynamic changes of individual amino acid as well as the secondary structural elements of the protein and the solvent. Stabilization of the peptide partially comes through a strong hydrogen bonding network with the protein, while hydrophobic interactions also play a significant role where the peptide inserts itself into a hydrophobic cavity of the protein. SLAM binding widens SAP's second binding site for Fyn, which is the next step in the signal transduction cascade. The higher stabilization and less fluctuation of specific residues of SAP in the Fyn binding site, induced by SAP-SLAM complexation, emerge as the key structural elements to trigger the recognition of SAP by the SH3 domain of Fyn. The thermodynamic quantification of the protein due to complexation not only throws deeper understanding in the established mode of SAP-SLAM interaction but also assists in the recognition of the relevant residues of the protein responsible for alterations in its activity.

Microscopic insight into thermodynamics of conformational changes of SAP-SLAM complex in signal transduction cascade.

PubMed

Samanta, Sudipta; Mukherjee, Sanchita

2017-04-28

The signalling lymphocytic activation molecule (SLAM) family of receptors, expressed by an array of immune cells, associate with SLAM-associated protein (SAP)-related molecules, composed of single SH2 domain architecture. SAP activates Src-family kinase Fyn after SLAM ligation, resulting in a SLAM-SAP-Fyn complex, where, SAP binds the Fyn SH3 domain that does not involve canonical SH3 or SH2 interactions. This demands insight into this SAP mediated signalling cascade. Thermodynamics of the conformational changes are extracted from the histograms of dihedral angles obtained from the all-atom molecular dynamics simulations of this structurally well characterized SAP-SLAM complex. The results incorporate the binding induced thermodynamic changes of individual amino acid as well as the secondary structural elements of the protein and the solvent. Stabilization of the peptide partially comes through a strong hydrogen bonding network with the protein, while hydrophobic interactions also play a significant role where the peptide inserts itself into a hydrophobic cavity of the protein. SLAM binding widens SAP's second binding site for Fyn, which is the next step in the signal transduction cascade. The higher stabilization and less fluctuation of specific residues of SAP in the Fyn binding site, induced by SAP-SLAM complexation, emerge as the key structural elements to trigger the recognition of SAP by the SH3 domain of Fyn. The thermodynamic quantification of the protein due to complexation not only throws deeper understanding in the established mode of SAP-SLAM interaction but also assists in the recognition of the relevant residues of the protein responsible for alterations in its activity.

Identification and Analysis of Mitogen-Activated Protein Kinase (MAPK) Cascades in Fragaria vesca.

PubMed

Zhou, Heying; Ren, Suyue; Han, Yuanfang; Zhang, Qing; Qin, Ling; Xing, Yu

2017-08-13

Mitogen-activated protein kinase (MAPK) cascades are highly conserved signaling modules in eukaryotes, including yeasts, plants and animals. MAPK cascades are responsible for protein phosphorylation during signal transduction events, and typically consist of three protein kinases: MAPK, MAPK kinase, and MAPK kinase kinase. In this current study, we identified a total of 12 FvMAPK , 7 FvMAPKK , 73 FvMAPKKK , and one FvMAPKKKK genes in the recently published Fragaria vesca genome sequence. This work reported the classification, annotation and phylogenetic evaluation of these genes and an assessment of conserved motifs and the expression profiling of members of the gene family were also analyzed here. The expression profiles of the MAPK and MAPKK genes in different organs and fruit developmental stages were further investigated using quantitative real-time reverse transcription PCR (qRT-PCR). Finally, the MAPK and MAPKK expression patterns in response to hormone and abiotic stresses (salt, drought, and high and low temperature) were investigated in fruit and leaves of F. vesca . The results provide a platform for further characterization of the physiological and biochemical functions of MAPK cascades in strawberry.

A respiratory chain controlled signal transduction cascade in the mitochondrial intermembrane space mediates hydrogen peroxide signaling

PubMed Central

Patterson, Heide Christine; Gerbeth, Carolin; Thiru, Prathapan; Vögtle, Nora F.; Knoll, Marko; Shahsafaei, Aliakbar; Samocha, Kaitlin E.; Huang, Cher X.; Harden, Mark Michael; Song, Rui; Chen, Cynthia; Kao, Jennifer; Shi, Jiahai; Salmon, Wendy; Shaul, Yoav D.; Stokes, Matthew P.; Silva, Jeffrey C.; Bell, George W.; MacArthur, Daniel G.; Ruland, Jürgen; Meisinger, Chris; Lodish, Harvey F.

2015-01-01

Reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) govern cellular homeostasis by inducing signaling. H2O2 modulates the activity of phosphatases and many other signaling molecules through oxidation of critical cysteine residues, which led to the notion that initiation of ROS signaling is broad and nonspecific, and thus fundamentally distinct from other signaling pathways. Here, we report that H2O2 signaling bears hallmarks of a regular signal transduction cascade. It is controlled by hierarchical signaling events resulting in a focused response as the results place the mitochondrial respiratory chain upstream of tyrosine-protein kinase Lyn, Lyn upstream of tyrosine-protein kinase SYK (Syk), and Syk upstream of numerous targets involved in signaling, transcription, translation, metabolism, and cell cycle regulation. The active mediators of H2O2 signaling colocalize as H2O2 induces mitochondria-associated Lyn and Syk phosphorylation, and a pool of Lyn and Syk reside in the mitochondrial intermembrane space. Finally, the same intermediaries control the signaling response in tissues and species responsive to H2O2 as the respiratory chain, Lyn, and Syk were similarly required for H2O2 signaling in mouse B cells, fibroblasts, and chicken DT40 B cells. Consistent with a broad role, the Syk pathway is coexpressed across tissues, is of early metazoan origin, and displays evidence of evolutionary constraint in the human. These results suggest that H2O2 signaling is under control of a signal transduction pathway that links the respiratory chain to the mitochondrial intermembrane space-localized, ubiquitous, and ancient Syk pathway in hematopoietic and nonhematopoietic cells. PMID:26438848

A respiratory chain controlled signal transduction cascade in the mitochondrial intermembrane space mediates hydrogen peroxide signaling.

PubMed

Patterson, Heide Christine; Gerbeth, Carolin; Thiru, Prathapan; Vögtle, Nora F; Knoll, Marko; Shahsafaei, Aliakbar; Samocha, Kaitlin E; Huang, Cher X; Harden, Mark Michael; Song, Rui; Chen, Cynthia; Kao, Jennifer; Shi, Jiahai; Salmon, Wendy; Shaul, Yoav D; Stokes, Matthew P; Silva, Jeffrey C; Bell, George W; MacArthur, Daniel G; Ruland, Jürgen; Meisinger, Chris; Lodish, Harvey F

2015-10-20

Reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) govern cellular homeostasis by inducing signaling. H2O2 modulates the activity of phosphatases and many other signaling molecules through oxidation of critical cysteine residues, which led to the notion that initiation of ROS signaling is broad and nonspecific, and thus fundamentally distinct from other signaling pathways. Here, we report that H2O2 signaling bears hallmarks of a regular signal transduction cascade. It is controlled by hierarchical signaling events resulting in a focused response as the results place the mitochondrial respiratory chain upstream of tyrosine-protein kinase Lyn, Lyn upstream of tyrosine-protein kinase SYK (Syk), and Syk upstream of numerous targets involved in signaling, transcription, translation, metabolism, and cell cycle regulation. The active mediators of H2O2 signaling colocalize as H2O2 induces mitochondria-associated Lyn and Syk phosphorylation, and a pool of Lyn and Syk reside in the mitochondrial intermembrane space. Finally, the same intermediaries control the signaling response in tissues and species responsive to H2O2 as the respiratory chain, Lyn, and Syk were similarly required for H2O2 signaling in mouse B cells, fibroblasts, and chicken DT40 B cells. Consistent with a broad role, the Syk pathway is coexpressed across tissues, is of early metazoan origin, and displays evidence of evolutionary constraint in the human. These results suggest that H2O2 signaling is under control of a signal transduction pathway that links the respiratory chain to the mitochondrial intermembrane space-localized, ubiquitous, and ancient Syk pathway in hematopoietic and nonhematopoietic cells.

Odorants selectively activate distinct G protein subtypes in olfactory cilia.

PubMed

Schandar, M; Laugwitz, K L; Boekhoff, I; Kroner, C; Gudermann, T; Schultz, G; Breer, H

1998-07-03

Chemoelectrical signal transduction in olfactory neurons appears to involve intracellular reaction cascades mediated by heterotrimeric GTP-binding proteins. In this study attempts were made to identify the G protein subtype(s) in olfactory cilia that are activated by the primary (odorant) signal. Antibodies directed against the alpha subunits of distinct G protein subtypes interfered specifically with second messenger reponses elicited by defined subsets of odorants; odor-induced cAMP-formation was attenuated by Galphas antibodies, whereas Galphao antibodies blocked odor-induced inositol 1,4, 5-trisphosphate (IP3) formation. Activation-dependent photolabeling of Galpha subunits with [alpha-32P]GTP azidoanilide followed by immunoprecipitation using subtype-specific antibodies enabled identification of particular individual G protein subtypes that were activated upon stimulation of isolated olfactory cilia by chemically distinct odorants. For example odorants that elicited a cAMP response resulted in labeling of a Galphas-like protein, whereas odorants that elicited an IP3 response led to the labeling of a Galphao-like protein. Since odorant-induced IP3 formation was also blocked by Gbeta antibodies, activation of olfactory phospholipase C might be mediated by betagamma subunits of a Go-like G protein. These results indicate that different subsets of odorants selectively trigger distinct reaction cascades and provide evidence for dual transduction pathways in olfactory signaling.

Various abiotic stresses rapidly activate Arabidopsis MAP kinases ATMPK4 and ATMPK6.

PubMed

Ichimura, K; Mizoguchi, T; Yoshida, R; Yuasa, T; Shinozaki, K

2000-12-01

Mitogen-activated protein kinase (MAP kinase, MAPK) cascades play pivotal roles in signal transduction of extracellular stimuli, such as environmental stresses and growth regulators, in various organisms. Arabidopsis thaliana MAP kinases constitute a gene family, but stimulatory signals for each MAP kinase have not been elucidated. Here we show that environmental stresses such as low temperature, low humidity, hyper-osmolarity, touch and wounding induce rapid and transient activation of the Arabidopsis MAP kinases ATMPK4 and ATMPK6. Activation of ATMPK4 and ATMPK6 was associated with tyrosine phosphorylation but not with the amounts of mRNA or protein. Kinetics during activation differ between these two MAP kinases. These results suggest that ATMPK4 and ATMPK6 are involved in distinct signal transduction pathways responding to these environmental stresses.

Using Cascading Style Sheets to Design a Fly-Out Menu with Microsoft Visual Studio

ERIC Educational Resources Information Center

Liu, Chang; Downing, Charles

2010-01-01

The menu has become an integrated component within nearly all professionally designed websites. This teaching tip presents a no-code way to design either a vertical or a horizontal fly-out menu by using Cascading Style Sheets (CSS) within Microsoft Visual Studio 2008. The approach described in this tip helps students fully understand how to…

Signal Transduction Pathways of TNAP: Molecular Network Analyses.

PubMed

Négyessy, László; Györffy, Balázs; Hanics, János; Bányai, Mihály; Fonta, Caroline; Bazsó, Fülöp

2015-01-01

Despite the growing body of evidence pointing on the involvement of tissue non-specific alkaline phosphatase (TNAP) in brain function and diseases like epilepsy and Alzheimer's disease, our understanding about the role of TNAP in the regulation of neurotransmission is severely limited. The aim of our study was to integrate the fragmented knowledge into a comprehensive view regarding neuronal functions of TNAP using objective tools. As a model we used the signal transduction molecular network of a pyramidal neuron after complementing with TNAP related data and performed the analysis using graph theoretic tools. The analyses show that TNAP is in the crossroad of numerous pathways and therefore is one of the key players of the neuronal signal transduction network. Through many of its connections, most notably with molecules of the purinergic system, TNAP serves as a controller by funnelling signal flow towards a subset of molecules. TNAP also appears as the source of signal to be spread via interactions with molecules involved among others in neurodegeneration. Cluster analyses identified TNAP as part of the second messenger signalling cascade. However, TNAP also forms connections with other functional groups involved in neuronal signal transduction. The results indicate the distinct ways of involvement of TNAP in multiple neuronal functions and diseases.

Activity Dependent Signal Transduction in Skeletal Muscle

NASA Technical Reports Server (NTRS)

Hamilton, Susan L.

1999-01-01

The overall goals of this project are: 1) to define the initial signal transduction events whereby the removal of gravitational load from antigravity muscles, such as the soleus, triggers muscle atrophy, and 2) to develop countermeasures to prevent this from happening. Our rationale for this approach is that, if countermeasures can be developed to regulate these early events, we could avoid having to deal with the multiple cascades of events that occur downstream from the initial event. One of our major findings is that hind limb suspension causes an early and sustained increase in intracellular Ca(2+) concentration ([Ca (2+)](sub i)). In most cells the consequences of changes in ([Ca (2+)](sub i))depend on the amplitude, frequency and duration of the Ca(2+) signal and on other factors in the intracellular environment. We propose that muscle remodeling in microgravity represents a change in the balance among several CA(2+) regulated signal transduction pathways, in particular those involving the transcription factors NFAT and NFkB and the pro-apoptotic protein BAD. Other Ca(2+) sensitive pathways involving PKC, ras, rac, and CaM kinase II may also contribute to muscle remodeling.

CasCADe: A Novel 4D Visualization System for Virtual Construction Planning.

PubMed

Ivson, Paulo; Nascimento, Daniel; Celes, Waldemar; Barbosa, Simone Dj

2018-01-01

Building Information Modeling (BIM) provides an integrated 3D environment to manage large-scale engineering projects. The Architecture, Engineering and Construction (AEC) industry explores 4D visualizations over these datasets for virtual construction planning. However, existing solutions lack adequate visual mechanisms to inspect the underlying schedule and make inconsistencies readily apparent. The goal of this paper is to apply best practices of information visualization to improve 4D analysis of construction plans. We first present a review of previous work that identifies common use cases and limitations. We then consulted with AEC professionals to specify the main design requirements for such applications. These guided the development of CasCADe, a novel 4D visualization system where task sequencing and spatio-temporal simultaneity are immediately apparent. This unique framework enables the combination of diverse analytical features to create an information-rich analysis environment. We also describe how engineering collaborators used CasCADe to review the real-world construction plans of an Oil & Gas process plant. The system made evident schedule uncertainties, identified work-space conflicts and helped analyze other constructability issues. The results and contributions of this paper suggest new avenues for future research in information visualization for the AEC industry.

Non-independent quantum bumps in Limulus ventral nerve photoreceptors--a new insight in the light transduction mechanism.

PubMed

Nagy, K

1992-09-14

Single photon-induced transient currents, called quantum bumps were stimulated by short flashes in dark-adapted ventral nerve photoreceptors of Limulus. Flash intensities were set to activate 3 or more bumps. In most cases, current bumps were activated with a constant rate. The frequency of bump occurrence was between 9 and 17 Hz. Results show that consecutive bumps are not independent and that some of them are not activated by a photon. The periodic bump activation indicates a molecular mechanism which quantifies the transmitter release not only by a light quantum, but also by a late phase of the transduction cascade. A model is proposed, in which Ca2+ ions released from intracellular stores transiently block the further Ca2+ release by inositol trisphosphate in an all-or-none manner.

Endocytosis and Signaling during Development

PubMed Central

Bökel, Christian

2014-01-01

The development of multicellular organisms relies on an intricate choreography of intercellular communication events that pattern the embryo and coordinate the formation of tissues and organs. It is therefore not surprising that developmental biology, especially using genetic model organisms, has contributed significantly to the discovery and functional dissection of the associated signal-transduction cascades. At the same time, biophysical, biochemical, and cell biological approaches have provided us with insights into the underlying cell biological machinery. Here we focus on how endocytic trafficking of signaling components (e.g., ligands or receptors) controls the generation, propagation, modulation, reception, and interpretation of developmental signals. A comprehensive enumeration of the links between endocytosis and signal transduction would exceed the limits of this review. We will instead use examples from different developmental pathways to conceptually illustrate the various functions provided by endocytic processes during key steps of intercellular signaling. PMID:24591521

The dysbindin gene in major depression: an association study.

PubMed

Zill, Peter; Baghai, Thomas C; Engel, Rolf; Zwanzger, Peter; Schüle, Cornelius; Eser, Daniela; Behrens, Stefanie; Rupprecht, Rainer; Möller, Hans-Jürgen; Ackenheil, Manfred; Bondy, Brigitta

2004-08-15

The pathophysiological mechanisms, as well as the molecular loci of antidepressant drug action have not yet been established, but recent models proposed that several adaptive mechanisms in signal transduction cascades beyond the receptor and reuptake systems are involved in antidepressant action and play an important role in the etiology of affective disorders. In this context, the dysbindin gene (dystrobrevin-binding-protein 1, DTNBP1), which was recently reported to be associated with schizophrenia seems to be an interesting candidate gene for affective disorders. Dysbindin is widely expressed in the human brain and binds to the dystrophin-associated protein complex (DPC) which appears to be involved in signal transduction pathways, which have been repeatedly investigated and described as altered or disturbed in affective disorders [McLeod et al. [2003: Psychopharmacol Bull 35:24-41]; Brambilla et al. [2003: Mol Psychiatry 8:721-737

Geometric analysis of Arabidopsis root apex reveals a new aspect of the ethylene signal transduction pathway in development

NASA Technical Reports Server (NTRS)

Cervantes, Emilio; Tocino, Angel

2005-01-01

Structurally, ethylene is the simplest phytohormone and regulates multiple aspects of plant growth and development. Its effects are mediated by a signal transduction cascade involving receptors, MAP kinases and transcription factors. Many morphological effects of ethylene in plant development, including root size, have been previously described. In this article a combined geometric and algebraic approach has been used to analyse the shape and the curvature in the root apex of Arabidopsis seedlings. The process requires the fitting of Bezier curves that reproduce the root apex shape, and the calculation of the corresponding curvatures. The application of the method has allowed us to identify significant differences in the root curvatures of ethylene insensitive mutants (ein2-1 and etr1-1) with respect to the wild-type Columbia.

Ras signaling in aging and metabolic regulation.

PubMed

Slack, Cathy

2017-12-07

Aberrant signal transduction downstream of the Ras GTPase has a well-established role in tumorigenesis. Mutations that result in hyperactivation of Ras are responsible for a third of all human cancers. Hence, small molecule inhibitors of the Ras signal transduction cascade have been under intense focus as potential cancer treatments. In both invertebrate and mammalian models, emerging evidence has also implicated components of the Ras signaling pathway in aging and metabolic regulation. Here, I review the current evidence for Ras signaling in these newly discovered roles highlighting the interactions between the Ras pathway and other longevity assurance mechanisms. Defining the role of Ras signaling in maintaining age-related health may have important implications for the development of interventions that could not only increase lifespan but also delay the onset and/or progression of age-related functional decline.

Genetic control of an epigenetic cell degeneration syndrome in Podospora anserina.

PubMed

Haedens, Vicki; Malagnac, Fabienne; Silar, Philippe

2005-06-01

Filamentous fungi frequently present degenerative processes, whose molecular basis is very often unknown. Here, we present three mutant screens that result in the identification of 29 genes that directly or indirectly control Crippled Growth (CG), an epigenetic cell degeneration of the filamentous ascomycete Podospora anserina. Two of these genes were previously shown to encode a MAP kinase kinase kinase and an NADPH oxidase involved in a signal transduction cascade that participates in stationary phase differentiations, fruiting body development and defence against fungal competitors. The numerous genes identified can be incorporated in a model in which CG results from the sustained activation of the MAP kinase cascade. Our data also emphasize the complex regulatory network underlying three interconnected processes in P. anserina: sexual reproduction, defence against competitors, and cell degeneration.

Opsin evolution and expression in arthropod compound eyes and ocelli: insights from the cricket Gryllus bimaculatus.

PubMed

Henze, Miriam J; Dannenhauer, Kara; Kohler, Martin; Labhart, Thomas; Gesemann, Matthias

2012-08-30

Opsins are key proteins in animal photoreception. Together with a light-sensitive group, the chromophore, they form visual pigments which initiate the visual transduction cascade when photoactivated. The spectral absorption properties of visual pigments are mainly determined by their opsins, and thus opsins are crucial for understanding the adaptations of animal eyes. Studies on the phylogeny and expression pattern of opsins have received considerable attention, but our knowledge about insect visual opsins is still limited. Up to now, researchers have focused on holometabolous insects, while general conclusions require sampling from a broader range of taxa. We have therefore investigated visual opsins in the ocelli and compound eyes of the two-spotted cricket Gryllus bimaculatus, a hemimetabolous insect. Phylogenetic analyses place all identified cricket sequences within the three main visual opsin clades of insects. We assign three of these opsins to visual pigments found in the compound eyes with peak absorbances in the green (515 nm), blue (445 nm) and UV (332 nm) spectral range. Their expression pattern divides the retina into distinct regions: (1) the polarization-sensitive dorsal rim area with blue- and UV-opsin, (2) a newly-discovered ventral band of ommatidia with blue- and green-opsin and (3) the remainder of the compound eye with UV- and green-opsin. In addition, we provide evidence for two ocellar photopigments with peak absorbances in the green (511 nm) and UV (350 nm) spectral range, and with opsins that differ from those expressed in the compound eyes. Our data show that cricket eyes are spectrally more specialized than has previously been assumed, suggesting that similar adaptations in other insect species might have been overlooked. The arrangement of spectral receptor types within some ommatidia of the cricket compound eyes differs from the generally accepted pattern found in holometabolous insect taxa and awaits a functional explanation. From the opsin phylogeny, we conclude that gene duplications, which permitted differential opsin expression in insect ocelli and compound eyes, occurred independently in several insect lineages and are recent compared to the origin of the eyes themselves.

Opsin evolution and expression in Arthropod compound Eyes and Ocelli: Insights from the cricket Gryllus bimaculatus

PubMed Central

2012-01-01

Background Opsins are key proteins in animal photoreception. Together with a light-sensitive group, the chromophore, they form visual pigments which initiate the visual transduction cascade when photoactivated. The spectral absorption properties of visual pigments are mainly determined by their opsins, and thus opsins are crucial for understanding the adaptations of animal eyes. Studies on the phylogeny and expression pattern of opsins have received considerable attention, but our knowledge about insect visual opsins is still limited. Up to now, researchers have focused on holometabolous insects, while general conclusions require sampling from a broader range of taxa. We have therefore investigated visual opsins in the ocelli and compound eyes of the two-spotted cricket Gryllus bimaculatus, a hemimetabolous insect. Results Phylogenetic analyses place all identified cricket sequences within the three main visual opsin clades of insects. We assign three of these opsins to visual pigments found in the compound eyes with peak absorbances in the green (515 nm), blue (445 nm) and UV (332 nm) spectral range. Their expression pattern divides the retina into distinct regions: (1) the polarization-sensitive dorsal rim area with blue- and UV-opsin, (2) a newly-discovered ventral band of ommatidia with blue- and green-opsin and (3) the remainder of the compound eye with UV- and green-opsin. In addition, we provide evidence for two ocellar photopigments with peak absorbances in the green (511 nm) and UV (350 nm) spectral range, and with opsins that differ from those expressed in the compound eyes. Conclusions Our data show that cricket eyes are spectrally more specialized than has previously been assumed, suggesting that similar adaptations in other insect species might have been overlooked. The arrangement of spectral receptor types within some ommatidia of the cricket compound eyes differs from the generally accepted pattern found in holometabolous insect taxa and awaits a functional explanation. From the opsin phylogeny, we conclude that gene duplications, which permitted differential opsin expression in insect ocelli and compound eyes, occurred independently in several insect lineages and are recent compared to the origin of the eyes themselves. PMID:22935102

Expression of Recombinant Phosphoproteins for Signal Transduction Studies.

PubMed

Barber, Karl W; Rinehart, Jesse

2017-01-01

Complex signaling cascades are difficult to study in vitro without phosphorylated proteins. Here, we describe a technique for the routine production of recombinant phosphoproteins by directly incorporating phosphoserine as a nonstandard amino acid. This protocol utilizes an optimized phosphoserine orthogonal translation system and an engineered strain of E. coli containing no genomic amber codons. This approach has been used to generate a variety of phosphorylated proteins to understand the role of protein phosphorylation in cell signaling.

Identification of a Novel Gnao-Mediated Alternate Olfactory Signaling Pathway in Murine OSNs.

PubMed

Scholz, Paul; Mohrhardt, Julia; Jansen, Fabian; Kalbe, Benjamin; Haering, Claudia; Klasen, Katharina; Hatt, Hanns; Osterloh, Sabrina

2016-01-01

It is generally agreed that in olfactory sensory neurons (OSNs), the binding of odorant molecules to their specific olfactory receptor (OR) triggers a cAMP-dependent signaling cascade, activating cyclic-nucleotide gated (CNG) channels. However, considerable controversy dating back more than 20 years has surrounded the question of whether alternate signaling plays a role in mammalian olfactory transduction. In this study, we demonstrate a specific alternate signaling pathway in Olfr73-expressing OSNs. Methylisoeugenol (MIEG) and at least one other known weak Olfr73 agonist (Raspberry Ketone) trigger a signaling cascade independent from the canonical pathway, leading to the depolarization of the cell. Interestingly, this pathway is mediated by Gnao activation, leading to Cl(-) efflux; however, the activation of adenylyl cyclase III (ACIII), the recruitment of Ca(2+) from extra-or intracellular stores, and phosphatidylinositol 3-kinase-dependent signaling (PI signaling) are not involved. Furthermore, we demonstrated that our newly identified pathway coexists with the canonical olfactory cAMP pathway in the same OSN and can be triggered by the same OR in a ligand-selective manner. We suggest that this pathway might reflect a mechanism for odor recognition predominantly used in early developmental stages before olfactory cAMP signaling is fully developed. Taken together, our findings support the existence of at least one odor-induced alternate signal transduction pathway in native OSNs mediated by Olfr73 in a ligand-selective manner.

Response to Hyperosmotic Stress

PubMed Central

Saito, Haruo; Posas, Francesc

2012-01-01

An appropriate response and adaptation to hyperosmolarity, i.e., an external osmolarity that is higher than the physiological range, can be a matter of life or death for all cells. It is especially important for free-living organisms such as the yeast Saccharomyces cerevisiae. When exposed to hyperosmotic stress, the yeast initiates a complex adaptive program that includes temporary arrest of cell-cycle progression, adjustment of transcription and translation patterns, and the synthesis and retention of the compatible osmolyte glycerol. These adaptive responses are mostly governed by the high osmolarity glycerol (HOG) pathway, which is composed of membrane-associated osmosensors, an intracellular signaling pathway whose core is the Hog1 MAP kinase (MAPK) cascade, and cytoplasmic and nuclear effector functions. The entire pathway is conserved in diverse fungal species, while the Hog1 MAPK cascade is conserved even in higher eukaryotes including humans. This conservation is illustrated by the fact that the mammalian stress-responsive p38 MAPK can rescue the osmosensitivity of hog1Δ mutations in response to hyperosmotic challenge. As the HOG pathway is one of the best-understood eukaryotic signal transduction pathways, it is useful not only as a model for analysis of osmostress responses, but also as a model for mathematical analysis of signal transduction pathways. In this review, we have summarized the current understanding of both the upstream signaling mechanism and the downstream adaptive responses to hyperosmotic stress in yeast. PMID:23028184

Release of GTP Exchange Factor Mediated Down-Regulation of Abscisic Acid Signal Transduction through ABA-Induced Rapid Degradation of RopGEFs

PubMed Central

Waadt, Rainer; Schroeder, Julian I.

2016-01-01

The phytohormone abscisic acid (ABA) is critical to plant development and stress responses. Abiotic stress triggers an ABA signal transduction cascade, which is comprised of the core components PYL/RCAR ABA receptors, PP2C-type protein phosphatases, and protein kinases. Small GTPases of the ROP/RAC family act as negative regulators of ABA signal transduction. However, the mechanisms by which ABA controls the behavior of ROP/RACs have remained unclear. Here, we show that an Arabidopsis guanine nucleotide exchange factor protein RopGEF1 is rapidly sequestered to intracellular particles in response to ABA. GFP-RopGEF1 is sequestered via the endosome-prevacuolar compartment pathway and is degraded. RopGEF1 directly interacts with several clade A PP2C protein phosphatases, including ABI1. Interestingly, RopGEF1 undergoes constitutive degradation in pp2c quadruple abi1/abi2/hab1/pp2ca mutant plants, revealing that active PP2C protein phosphatases protect and stabilize RopGEF1 from ABA-mediated degradation. Interestingly, ABA-mediated degradation of RopGEF1 also plays an important role in ABA-mediated inhibition of lateral root growth. The presented findings point to a PP2C-RopGEF-ROP/RAC control loop model that is proposed to aid in shutting off ABA signal transduction, to counteract leaky ABA signal transduction caused by “monomeric” PYL/RCAR ABA receptors in the absence of stress, and facilitate signaling in response to ABA. PMID:27192441

Spectral Tuning of Killer Whale (Orcinus orca) Rhodopsin: Evidence for Positive Selection and Functional Adaptation in a Cetacean Visual Pigment.

PubMed

Dungan, Sarah Z; Kosyakov, Alexander; Chang, Belinda S W

2016-02-01

Cetaceans have undergone a remarkable evolutionary transition that was accompanied by many sensory adaptations, including modification of the visual system for underwater environments. Recent sequencing of cetacean genomes has made it possible to begin exploring the molecular basis of these adaptations. In this study we use in vitro expression methods to experimentally characterize the first step of the visual transduction cascade, the light activation of rhodopsin, for the killer whale. To investigate the spectral effects of amino acid substitutions thought to correspond with absorbance shifts relative to terrestrial mammals, we used the orca gene as a background for the first site-directed mutagenesis experiments in a cetacean rhodopsin. The S292A mutation had the largest effect, and was responsible for the majority of the spectral difference between killer whale and bovine (terrestrial) rhodopsin. Using codon-based likelihood models, we also found significant evidence for positive selection in cetacean rhodopsin sequences, including on spectral tuning sites we experimentally mutated. We then investigated patterns of ecological divergence that may be correlated with rhodopsin functional variation by using a series of clade models that partitioned the data set according to phylogeny, habitat, and foraging depth zone. Only the model partitioning according to depth was significant. This suggests that foraging dives might be a selective regime influencing cetacean rhodopsin divergence, and our experimental results indicate that spectral tuning may be playing an adaptive role in this process. Our study demonstrates that combining computational and experimental methods is crucial for gaining insight into the selection pressures underlying molecular evolution. © The Author 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

A Visualization Study of Secondary Flows in Cascades

NASA Technical Reports Server (NTRS)

Herzig, Howard Z; Hansen, Arthur G; Costello, George R

1954-01-01

Flow-visualization techniques are employed to ascertain the streamline patterns of the nonpotential secondary flows in the boundary layers of cascades, and thereby to provide a basis for more extended analyses in turbomachines. The three-dimensional deflection of the end-wall boundary layer results in the formation of a vortex within each cascade passage. The size and tightness of the vortex generated depend upon the main-flow turning in the cascade passage. Once formed, a vortex resists turning in subsequent blade rows, with consequent unfavorable angles of attack and possible flow disturbances on the pressure surfaces of subsequent blade rows when the vortices impinge on these surfaces. Two major tip-clearance effects are observed, the formation of a tip-clearance vortex and the scraping effect of a blade with relative motion past the wall boundary layer. The flow patterns indicate methods for improving the blade tip-loading characteristics of compressors and of low- and high-speed turbulence.

Repulsive Guidance Molecules (RGMs) and Neogenin in Bone Morphogenetic Protein (BMP) signaling

PubMed Central

Tian, Chenxi; Liu, Jun

2015-01-01

Summary Bone morphogenetic proteins (BMPs) belong to the transforming growth factor-beta (TGFβ) superfamily. BMPs mediate a highly conserved signal transduction cascade through the type I and type II serine/threonine kinase receptors and intracellular Smad proteins. The BMP pathway regulates multiple developmental and homeostatic processes. Mutations in this pathway can cause various diseases in humans, such as skeletal disorders, cardiovascular diseases and various cancers. Multiple levels of regulation, including extracellular regulation, help to ensure proper spatiotemporal control of BMP signaling in the right cellular context. The family of repulsive guidance molecules (RGMs) and the type I trans-membrane protein neogenin, a paralog of DCC (Deleted in Colorectal Cancer), have been implicated in modulating the BMP pathway. In this review, we discuss the properties and functions of RGM proteins and neogenin, focusing on their roles in the modulation of BMP signal transduction. PMID:23740870

Campylobacter jejuni transducer like proteins: Chemotaxis and beyond

PubMed Central

Chandrashekhar, Kshipra; Kassem, Issmat I.; Rajashekara, Gireesh

2017-01-01

ABSTRACT Chemotaxis, a process that mediates directional motility toward or away from chemical stimuli (chemoeffectors/ligands that can be attractants or repellents) in the environment, plays an important role in the adaptation of Campylobacter jejuni to disparate niches. The chemotaxis system consists of core signal transduction proteins and methyl-accepting-domain-containing Transducer like proteins (Tlps). Ligands binding to Tlps relay a signal to chemotaxis proteins in the cytoplasm which initiate a signal transduction cascade, culminating into a directional flagellar movement. Tlps facilitate substrate-specific chemotaxis in C. jejuni, which plays an important role in the pathogen's adaptation, pathobiology and colonization of the chicken gastrointestinal tract. However, the role of Tlps in C. jejuni's host tissue specific colonization, physiology and virulence remains not completely understood. Based on recent studies, it can be predicted that Tlps might be important targets for developing strategies to control C. jejuni via vaccines and antimicrobials. PMID:28080213

Campylobacter jejuni transducer like proteins: Chemotaxis and beyond.

PubMed

Chandrashekhar, Kshipra; Kassem, Issmat I; Rajashekara, Gireesh

2017-07-04

Chemotaxis, a process that mediates directional motility toward or away from chemical stimuli (chemoeffectors/ligands that can be attractants or repellents) in the environment, plays an important role in the adaptation of Campylobacter jejuni to disparate niches. The chemotaxis system consists of core signal transduction proteins and methyl-accepting-domain-containing Transducer like proteins (Tlps). Ligands binding to Tlps relay a signal to chemotaxis proteins in the cytoplasm which initiate a signal transduction cascade, culminating into a directional flagellar movement. Tlps facilitate substrate-specific chemotaxis in C. jejuni, which plays an important role in the pathogen's adaptation, pathobiology and colonization of the chicken gastrointestinal tract. However, the role of Tlps in C. jejuni's host tissue specific colonization, physiology and virulence remains not completely understood. Based on recent studies, it can be predicted that Tlps might be important targets for developing strategies to control C. jejuni via vaccines and antimicrobials.

Controlled membrane translocation provides a mechanism for signal transduction and amplification

NASA Astrophysics Data System (ADS)

Langton, Matthew J.; Keymeulen, Flore; Ciaccia, Maria; Williams, Nicholas H.; Hunter, Christopher A.

2017-05-01

Transmission and amplification of chemical signals across lipid bilayer membranes is of profound significance in many biological processes, from the development of multicellular organisms to information processing in the nervous system. In biology, membrane-spanning proteins are responsible for the transmission of chemical signals across membranes, and signal transduction is often associated with an amplified signalling cascade. The ability to reproduce such processes in artificial systems has potential applications in sensing, controlled drug delivery and communication between compartments in tissue-like constructs of synthetic vesicles. Here we describe a mechanism for transmitting a chemical signal across a membrane based on the controlled translocation of a synthetic molecular transducer from one side of a lipid bilayer membrane to the other. The controlled molecular motion has been coupled to the activation of a catalyst on the inside of a vesicle, which leads to a signal-amplification process analogous to the biological counterpart.

Mechanistic Insights in Ethylene Perception and Signal Transduction1

PubMed Central

Ju, Chuanli; Chang, Caren

2015-01-01

The gaseous hormone ethylene profoundly affects plant growth, development, and stress responses. Ethylene perception occurs at the endoplasmic reticulum membrane, and signal transduction leads to a transcriptional cascade that initiates diverse responses, often in conjunction with other signals. Recent findings provide a more complete picture of the components and mechanisms in ethylene signaling, now rendering a more dynamic view of this conserved pathway. This includes newly identified protein-protein interactions at the endoplasmic reticulum membrane, as well as the major discoveries that the central regulator ETHYLENE INSENSITIVE2 (EIN2) is the long-sought phosphorylation substrate for the CONSTITUTIVE RESPONSE1 protein kinase, and that cleavage of EIN2 transmits the signal to the nucleus. In the nucleus, hundreds of potential gene targets of the EIN3 master transcription factor have been identified and found to be induced in transcriptional waves, and transcriptional coregulation has been shown to be a mechanism of ethylene cross talk. PMID:26246449

Taste transductions in taste receptor cells: basic tastes and moreover.

PubMed

Iwata, Shusuke; Yoshida, Ryusuke; Ninomiya, Yuzo

2014-01-01

In the oral cavity, taste receptor cells dedicate to detecting chemical compounds in foodstuffs and transmitting their signals to gustatory nerve fibers. Heretofore, five taste qualities (sweet, umami, bitter, salty and sour) are generally accepted as basic tastes. Each of these may have a specific role in the detection of nutritious and poisonous substances; sweet for carbohydrate sources of calories, umami for protein and amino acid contents, bitter for harmful compounds, salty for minerals and sour for ripeness of fruits and spoiled foods. Recent studies have revealed molecular mechanisms for reception and transduction of these five basic tastes. Sweet, umami and bitter tastes are mediated by G-protein coupled receptors (GPCRs) and second-messenger signaling cascades. Salty and sour tastes are mediated by channel-type receptors. In addition to five basic tastes, taste receptor cells may have the ability to detect fat taste, which is elicited by fatty acids, and calcium taste, which is elicited by calcium. Taste compounds eliciting either fat taste or calcium taste may be detected by specific GPCRs expressed in taste receptor cells. This review will focus on transduction mechanisms and cellular characteristics responsible for each of basic tastes, fat taste and calcium taste.

The stimulatory Gα(s) protein is involved in olfactory signal transduction in Drosophila.

PubMed

Deng, Ying; Zhang, Weiyi; Farhat, Katja; Oberland, Sonja; Gisselmann, Günter; Neuhaus, Eva M

2011-04-07

Seven-transmembrane receptors typically mediate olfactory signal transduction by coupling to G-proteins. Although insect odorant receptors have seven transmembrane domains like G-protein coupled receptors, they have an inverted membrane topology, constituting a key difference between the olfactory systems of insects and other animals. While heteromeric insect ORs form ligand-activated non-selective cation channels in recombinant expression systems, the evidence for an involvement of cyclic nucleotides and G-proteins in odor reception is inconsistent. We addressed this question in vivo by analyzing the role of G-proteins in olfactory signaling using electrophysiological recordings. We found that Gα(s) plays a crucial role for odorant induced signal transduction in OR83b expressing olfactory sensory neurons, but not in neurons expressing CO₂ responsive proteins GR21a/GR63a. Moreover, signaling of Drosophila ORs involved Gα(s) also in a heterologous expression system. In agreement with these observations was the finding that elevated levels of cAMP result in increased firing rates, demonstrating the existence of a cAMP dependent excitatory signaling pathway in the sensory neurons. Together, we provide evidence that Gα(s) plays a role in the OR mediated signaling cascade in Drosophila.

Integrins engage mitochondrial function for signal transduction by a mechanism dependent on Rho GTPases

PubMed Central

Werner, Erica; Werb, Zena

2002-01-01

We show here the transient activation of the small GTPase Rac, followed by a rise in reactive oxygen species (ROS), as necessary early steps in a signal transduction cascade that lead to NFκB activation and collagenase-1 (CL-1)/matrix metalloproteinase-1 production after integrin-mediated cell shape changes. We show evidence indicating that this constitutes a new mechanism for ROS production mediated by small GTPases. Activated RhoA also induced ROS production and up-regulated CL-1 expression. A Rac mutant (L37) that prevents reorganization of the actin cytoskeleton prevented integrin-induced CL-1 expression, whereas mutations that abrogate Rac binding to the neutrophil NADPH membrane oxidase in vitro (H26 and N130) did not. Instead, ROS were produced by integrin-induced changes in mitochondrial function, which were inhibited by Bcl-2 and involved transient membrane potential loss. The cells showing this transient decrease in mitochondrial membrane potential were already committed to CL-1 expression. These results unveil a new molecular mechanism of signal transduction triggered by integrin engagement where a global mitochondrial metabolic response leads to gene expression rather than apoptosis. PMID:12119354

Integrins engage mitochondrial function for signal transduction by a mechanism dependent on Rho GTPases.

PubMed

Werner, Erica; Werb, Zena

2002-07-22

We show here the transient activation of the small GTPase Rac, followed by a rise in reactive oxygen species (ROS), as necessary early steps in a signal transduction cascade that lead to NFkappaB activation and collagenase-1 (CL-1)/matrix metalloproteinase-1 production after integrin-mediated cell shape changes. We show evidence indicating that this constitutes a new mechanism for ROS production mediated by small GTPases. Activated RhoA also induced ROS production and up-regulated CL-1 expression. A Rac mutant (L37) that prevents reorganization of the actin cytoskeleton prevented integrin-induced CL-1 expression, whereas mutations that abrogate Rac binding to the neutrophil NADPH membrane oxidase in vitro (H26 and N130) did not. Instead, ROS were produced by integrin-induced changes in mitochondrial function, which were inhibited by Bcl-2 and involved transient membrane potential loss. The cells showing this transient decrease in mitochondrial membrane potential were already committed to CL-1 expression. These results unveil a new molecular mechanism of signal transduction triggered by integrin engagement where a global mitochondrial metabolic response leads to gene expression rather than apoptosis.

Protein receptor-independent plasma membrane remodeling by HAMLET: a tumoricidal protein-lipid complex

PubMed Central

Nadeem, Aftab; Sanborn, Jeremy; Gettel, Douglas L.; James, Ho C. S.; Rydström, Anna; Ngassam, Viviane N.; Klausen, Thomas Kjær; Pedersen, Stine Falsig; Lam, Matti; Parikh, Atul N.; Svanborg, Catharina

2015-01-01

A central tenet of signal transduction in eukaryotic cells is that extra-cellular ligands activate specific cell surface receptors, which orchestrate downstream responses. This ‘’protein-centric” view is increasingly challenged by evidence for the involvement of specialized membrane domains in signal transduction. Here, we propose that membrane perturbation may serve as an alternative mechanism to activate a conserved cell-death program in cancer cells. This view emerges from the extraordinary manner in which HAMLET (Human Alpha-lactalbumin Made LEthal to Tumor cells) kills a wide range of tumor cells in vitro and demonstrates therapeutic efficacy and selectivity in cancer models and clinical studies. We identify a ‘’receptor independent” transformation of vesicular motifs in model membranes, which is paralleled by gross remodeling of tumor cell membranes. Furthermore, we find that HAMLET accumulates within these de novo membrane conformations and define membrane blebs as cellular compartments for direct interactions of HAMLET with essential target proteins such as the Ras family of GTPases. Finally, we demonstrate lower sensitivity of healthy cell membranes to HAMLET challenge. These features suggest that HAMLET-induced curvature-dependent membrane conformations serve as surrogate receptors for initiating signal transduction cascades, ultimately leading to cell death. PMID:26561036

Protein receptor-independent plasma membrane remodeling by HAMLET: a tumoricidal protein-lipid complex.

PubMed

Nadeem, Aftab; Sanborn, Jeremy; Gettel, Douglas L; James, Ho C S; Rydström, Anna; Ngassam, Viviane N; Klausen, Thomas Kjær; Pedersen, Stine Falsig; Lam, Matti; Parikh, Atul N; Svanborg, Catharina

2015-11-12

A central tenet of signal transduction in eukaryotic cells is that extra-cellular ligands activate specific cell surface receptors, which orchestrate downstream responses. This ''protein-centric" view is increasingly challenged by evidence for the involvement of specialized membrane domains in signal transduction. Here, we propose that membrane perturbation may serve as an alternative mechanism to activate a conserved cell-death program in cancer cells. This view emerges from the extraordinary manner in which HAMLET (Human Alpha-lactalbumin Made LEthal to Tumor cells) kills a wide range of tumor cells in vitro and demonstrates therapeutic efficacy and selectivity in cancer models and clinical studies. We identify a ''receptor independent" transformation of vesicular motifs in model membranes, which is paralleled by gross remodeling of tumor cell membranes. Furthermore, we find that HAMLET accumulates within these de novo membrane conformations and define membrane blebs as cellular compartments for direct interactions of HAMLET with essential target proteins such as the Ras family of GTPases. Finally, we demonstrate lower sensitivity of healthy cell membranes to HAMLET challenge. These features suggest that HAMLET-induced curvature-dependent membrane conformations serve as surrogate receptors for initiating signal transduction cascades, ultimately leading to cell death.

THE RGM/DRAGON FAMILY OF BMP CO-RECEPTORS

PubMed Central

Corradini, Elena; Babitt, Jodie L.; Lin, Herbert Y.

2013-01-01

The BMP signaling pathway controls a number of cell processes during development and in adult tissues. At the cellular level, ligands of the BMP family act by binding a hetero-tetrameric signaling complex, composed of two type I and two type II receptors. BMP ligands make use of a limited number of receptors, which in turn activate a common signal transduction cascade at the intracellular level. A complex regulatory network is required in order to activate the signaling cascade at proper times and locations, and to generate specific downstream effects in the appropriate cellular context. One such regulatory mechanism is the repulsive guidance molecule (RGM) family of BMP co-receptors. This article reviews the current knowledge regarding the structure, regulation, and function of RGMs, focusing on known and potential roles of RGMs in physiology and pathophysiology. PMID:19897400

G-protein-mediated inhibition of the Trp channel TRPM1 requires the Gβγ dimer.

PubMed

Shen, Yin; Rampino, Melissa Ann F; Carroll, Reed C; Nawy, Scott

2012-05-29

ON bipolar cells are critical for the function of the ON pathway in the visual system. They express a metabotropic glutamate receptor (mGluR6) that, when activated, couples to the G(o) class of G protein. The channel that is primarily responsible for the synaptic response has been recently identified as the transient receptor potential cation channel subfamily M member 1 (TRPM1); TRPM1 is negatively coupled to the mGluR6/Go cascade such that activation of the cascade results in closure of the channel. Light indirectly opens TRPM1 by reducing transmitter release from presynaptic photoreceptors, resulting in a decrease in mGluR6 activation. Conversely, in the dark, binding of synaptic glutamate to mGluR6 inhibits TRPM1 current. Closure of TRPM1 by G-protein activation in the dark is a critical step in the process of ON bipolar cell signal transduction, but the precise pathway linking these two events is not understood. To address this question, we measured TRPM1 activity in retinal bipolar cells, in human ependymal melanocytes (HEMs) that endogenously express TRPM1, and in HEK293 cells transfected with TRPM1. Dialysis of the Gβγ subunit dimer, but not Gα(o), closed TRPM1 channels in every cell type that we tested. In addition, activation of an endogenous G-protein-coupled receptor pathway in HEK293 cells that releases Gβγ without activating Go protein also closed TRPM1 channels. These results suggest a model in which the Gβγ dimer that is released as a result of the dissociation from Gα(o) upon activation of mGluR6 closes the TRPM1 channel, perhaps via a direct interaction.

Guanine nucleotide-binding protein (Gα) endocytosis by a cascade of ubiquitin binding domain proteins is required for sustained morphogenesis and proper mating in yeast.

PubMed

Dixit, Gauri; Baker, Rachael; Sacks, Carly M; Torres, Matthew P; Dohlman, Henrik G

2014-05-23

Heterotrimeric G proteins are well known to transmit signals from cell surface receptors to intracellular effector proteins. There is growing appreciation that G proteins are also present at endomembrane compartments, where they can potentially interact with a distinct set of signaling proteins. Here, we examine the cellular trafficking function of the G protein α subunit in yeast, Gpa1. Gpa1 contains a unique 109-amino acid insert within the α-helical domain that undergoes a variety of posttranslational modifications. Among these is monoubiquitination, catalyzed by the NEDD4 family ubiquitin ligase Rsp5. Using a newly optimized method for G protein purification together with biophysical measures of structure and function, we show that the ubiquitination domain does not influence enzyme activity. By screening a panel of 39 gene deletion mutants, each lacking a different ubiquitin binding domain protein, we identify seven that are necessary to deliver Gpa1 to the vacuole compartment including four proteins (Ede1, Bul1, Ddi1, and Rup1) previously not known to be involved in this process. Finally, we show that proper endocytosis of the G protein is needed for sustained cellular morphogenesis and mating in response to pheromone stimulation. We conclude that a cascade of ubiquitin-binding proteins serves to deliver the G protein to its final destination within the cell. In this instance and in contrast to the previously characterized visual system, endocytosis from the plasma membrane is needed for proper signal transduction rather than for signal desensitization. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Simultaneous Loss of NCKX4 and CNG Channel Desensitization Impairs Olfactory Sensitivity.

PubMed

Ferguson, Christopher H; Zhao, Haiqing

2017-01-04

In vertebrate olfactory sensory neurons (OSNs), Ca 2+ plays key roles in both mediating and regulating the olfactory response. Ca 2+ enters OSN cilia during the response through the olfactory cyclic nucleotide-gated (CNG) channel and stimulates a depolarizing chloride current by opening the olfactory Ca 2+ -activated chloride channel to amplify the response. Ca 2+ also exerts negative regulation on the olfactory transduction cascade, through mechanisms that include reducing the CNG current by desensitizing the CNG channel via Ca 2+ /calmodulin (CaM), to reduce the response. Ca 2+ is removed from the cilia primarily by the K + -dependent Na + /Ca 2+ exchanger 4 (NCKX4), and the removal of Ca 2+ leads to closure of the chloride channel and response termination. In this study, we investigate how two mechanisms conventionally considered negative regulatory mechanisms of olfactory transduction, Ca 2+ removal by NCKX4, and desensitization of the CNG channel by Ca 2+ /CaM, interact to regulate the olfactory response. We performed electro-olfactogram (EOG) recordings on the double-mutant mice, NCKX4 -/- ;CNGB1 ΔCaM , which are simultaneously lacking NCKX4 (NCKX4 -/- ) and Ca 2+ /CaM-mediated CNG channel desensitization (CNGB1 ΔCaM ). Despite exhibiting alterations in various response attributes, including termination kinetics and adaption properties, OSNs in either NCKX4 -/- mice or CNGB1 ΔCaM mice show normal resting sensitivity, as determined by their unchanged EOG response amplitude. We found that OSNs in NCKX4 -/- ;CNGB1 ΔCaM mice displayed markedly reduced EOG amplitude accompanied by alterations in other response attributes. This study suggests that what are conventionally considered negative regulatory mechanisms of olfactory transduction also play a role in setting the resting sensitivity in OSNs. Sensory receptor cells maintain high sensitivity at rest. Although the mechanisms responsible for setting the resting sensitivity of sensory receptor cells are not well understood, it has generally been assumed that the sensitivity is set primarily by how effectively the components in the activation cascade of sensory transduction can be stimulated. Our findings in mouse olfactory sensory neurons suggest that mechanisms that are primarily responsible for terminating the olfactory response are also critical for proper resting sensitivity. Copyright © 2017 the authors 0270-6474/17/370110-10$15.00/0.

Elementary signaling modes predict the essentiality of signal transduction network components

PubMed Central

2011-01-01

Background Understanding how signals propagate through signaling pathways and networks is a central goal in systems biology. Quantitative dynamic models help to achieve this understanding, but are difficult to construct and validate because of the scarcity of known mechanistic details and kinetic parameters. Structural and qualitative analysis is emerging as a feasible and useful alternative for interpreting signal transduction. Results In this work, we present an integrative computational method for evaluating the essentiality of components in signaling networks. This approach expands an existing signaling network to a richer representation that incorporates the positive or negative nature of interactions and the synergistic behaviors among multiple components. Our method simulates both knockout and constitutive activation of components as node disruptions, and takes into account the possible cascading effects of a node's disruption. We introduce the concept of elementary signaling mode (ESM), as the minimal set of nodes that can perform signal transduction independently. Our method ranks the importance of signaling components by the effects of their perturbation on the ESMs of the network. Validation on several signaling networks describing the immune response of mammals to bacteria, guard cell abscisic acid signaling in plants, and T cell receptor signaling shows that this method can effectively uncover the essentiality of components mediating a signal transduction process and results in strong agreement with the results of Boolean (logical) dynamic models and experimental observations. Conclusions This integrative method is an efficient procedure for exploratory analysis of large signaling and regulatory networks where dynamic modeling or experimental tests are impractical. Its results serve as testable predictions, provide insights into signal transduction and regulatory mechanisms and can guide targeted computational or experimental follow-up studies. The source codes for the algorithms developed in this study can be found at http://www.phys.psu.edu/~ralbert/ESM. PMID:21426566

Targeting signal transduction pathways of cancer stem cells for therapeutic opportunities of metastasis.

PubMed

Iqbal, Waqas; Alkarim, Saleh; AlHejin, Ahmed; Mukhtar, Hasan; Saini, Kulvinder S

2016-11-15

Tumor comprises of heterogeneous population of cells where not all the disseminated cancer cells have the prerogative and "in-build genetic cues" to form secondary tumors. Cells with stem like properties complemented by key signaling molecules clearly have shown to exhibit selective growth advantage to form tumors at distant metastatic sites. Thus, defining the role of cancer stem cells (CSC) in tumorigenesis and metastasis is emerging as a major thrust area for therapeutic intervention. Precise relationship and regulatory mechanisms operating in various signal transduction pathways during cancer dissemination, extravasation and angiogenesis still remain largely enigmatic. How the crosstalk amongst circulating tumor cells (CTC), epithelial mesenchymal transition (EMT) process and CSC is coordinated for initiating the metastasis at secondary tissues, and during cancer relapse could be of great therapeutic interest. The signal transduction mechanisms facilitating the dissemination, infiltration of CSC into blood stream, extravasations, progression of metastasis phenotype and angiogenesis, at distant organs, are the key pathologically important vulnerabilities being elucidated. Therefore, current new drug discovery focus has shifted towards finding "key driver genes" operating in parallel signaling pathways, during quiescence, survival and maintenance of stemness in CSC. Understanding these mechanisms could open new horizons for tackling the issue of cancer recurrence and metastasis-the cause of ~90% cancer associated mortality. To design futuristic & targeted therapies, we propose a multi-pronged strategy involving small molecules, RNA interference, vaccines, antibodies and other biotechnological modalities against CSC and the metastatic signal transduction cascade.

Caenorhabditis elegans TRPV Channels Function in a Modality-Specific Pathway to Regulate Response to Aberrant Sensory Signaling

PubMed Central

Ezak , Meredith J.; Hong , Elizabeth; Chaparro-Garcia , Angela; Ferkey , Denise M.

2010-01-01

Olfaction and some forms of taste (including bitter) are mediated by G protein-coupled signal transduction pathways. Olfactory and gustatory ligands bind to chemosensory G protein-coupled receptors (GPCRs) in specialized sensory cells to activate intracellular signal transduction cascades. G protein-coupled receptor kinases (GRKs) are negative regulators of signaling that specifically phosphorylate activated GPCRs to terminate signaling. Although loss of GRK function usually results in enhanced cellular signaling, Caenorhabditis elegans lacking GRK-2 function are not hypersensitive to chemosensory stimuli. Instead, grk-2 mutant animals do not chemotax toward attractive olfactory stimuli or avoid aversive tastes and smells. We show here that loss-of-function mutations in the transient receptor potential vanilloid (TRPV) channels OSM-9 and OCR-2 selectively restore grk-2 behavioral avoidance of bitter tastants, revealing modality-specific mechanisms for TRPV channel function in the regulation of C. elegans chemosensation. Additionally, a single amino acid point mutation in OCR-2 that disrupts TRPV channel-mediated gene expression, but does not decrease channel function in chemosensory primary signal transduction, also restores grk-2 bitter taste avoidance. Thus, loss of GRK-2 function may lead to changes in gene expression, via OSM-9/OCR-2, to selectively alter the levels of signaling components that transduce or regulate bitter taste responses. Our results suggest a novel mechanism and multiple modality-specific pathways that sensory cells employ in response to aberrant signal transduction. PMID:20176974

Elucidation of Cross-Talk and Specificity of Early Response Mechanisms to Salt and PEG-Simulated Drought Stresses in Brassica napus Using Comparative Proteomic Analysis

PubMed Central

Luo, Junling; Tang, Shaohua; Peng, Xiaojue; Yan, Xiaohong; Zeng, Xinhua; Li, Jun; Li, Xiaofei; Wu, Gang

2015-01-01

To understand the cross-talk and specificity of the early responses of plants to salt and drought, we performed physiological and proteome analyses of Brassica napus seedlings pretreated with 245 mM NaCl or 25% polyethylene glycol (PEG) 6000 under identical osmotic pressure (-1.0 MPa). Significant decreases in water content and photosynthetic rate and excessive accumulation of compatible osmolytes and oxidative damage were observed in response to both stresses. Unexpectedly, the drought response was more severe than the salt response. We further identified 45 common differentially expressed proteins (DEPs), 143 salt-specific DEPs and 160 drought-specific DEPs by isobaric tags for relative and absolute quantitation (iTRAQ) analysis. The proteome quantitative data were then confirmed by multiple reaction monitoring (MRM). The differences in the proteomic profiles between drought-treated and salt-treated seedlings exceeded the similarities in the early stress responses. Signal perception and transduction, transport and membrane trafficking, and photosynthesis-related proteins were enriched as part of the molecular cross-talk and specificity mechanism in the early responses to the two abiotic stresses. The Ca2+ signaling, G protein-related signaling, 14-3-3 signaling pathway and phosphorylation cascades were the common signal transduction pathways shared by both salt and drought stress responses; however, the proteins with executive functions varied. These results indicate functional specialization of family proteins in response to different stresses, i.e., CDPK21, TPR, and CTR1 specific to phosphorylation cascades under early salt stress, whereas STN7 and BSL were specific to phosphorylation cascades under early drought stress. Only the calcium-binding EF-hand family protein and ZKT were clearly identified as signaling proteins that acted as cross-talk nodes for salt and drought signaling pathways. Our study provides new clues and insights for developing strategies to improve the tolerance of crops to complex, multiple environmental stresses. PMID:26448643

Molecular Steps in the Immune Signaling Pathway Evoked by Plant Elicitor Peptides: Ca2+-Dependent Protein Kinases, Nitric Oxide, and Reactive Oxygen Species Are Downstream from the Early Ca2+ Signal1[OPEN

PubMed Central

Ma, Yi; Zhao, Yichen; Walker, Robin K.; Berkowitz, Gerald A.

2013-01-01

Endogenous plant elicitor peptides (Peps) can act to facilitate immune signaling and pathogen defense responses. Binding of these peptides to the Arabidopsis (Arabidopsis thaliana) plasma membrane-localized Pep receptors (PEPRs) leads to cytosolic Ca2+ elevation, an early event in a signaling cascade that activates immune responses. This immune response includes the amplification of signaling evoked by direct perception of pathogen-associated molecular patterns by plant cells under assault. Work included in this report further characterizes the Pep immune response and identifies new molecular steps in the signal transduction cascade. The PEPR coreceptor BRASSINOSTEROID-INSENSITIVE1 Associated Kinase1 contributes to generation of the Pep-activated Ca2+ signal and leads to increased defense gene expression and resistance to a virulent bacterial pathogen. Ca2+-dependent protein kinases (CPKs) decode the Ca2+ signal, also facilitating defense gene expression and enhanced resistance to the pathogen. Nitric oxide and reduced nicotinamide adenine dinucleotide phosphate oxidase-dependent reactive oxygen species generation (due to the function of Respiratory Burst Oxidase Homolog proteins D and F) are also involved downstream from the Ca2+ signal in the Pep immune defense signal transduction cascade, as is the case with BRASSINOSTEROID-INSENSITIVE1 Associated Kinase1 and CPK5, CPK6, and CPK11. These steps of the pathogen defense response are required for maximal Pep immune activation that limits growth of a virulent bacterial pathogen in the plant. We find a synergism between function of the PEPR and Flagellin Sensing2 receptors in terms of both nitric oxide and reactive oxygen species generation. Presented results are also consistent with the involvement of the secondary messenger cyclic GMP and a cyclic GMP-activated Ca2+-conducting channel in the Pep immune signaling pathway. PMID:24019427

PrP(C) signalling in neurons: from basics to clinical challenges.

PubMed

Hirsch, Théo Z; Hernandez-Rapp, Julia; Martin-Lannerée, Séverine; Launay, Jean-Marie; Mouillet-Richard, Sophie

2014-09-01

The cellular prion protein PrP(C) was identified over twenty-five years ago as the normal counterpart of the scrapie prion protein PrP(Sc), itself the main if not the sole component of the infectious agent at the root of Transmissible Spongiform Encephalopathies (TSEs). PrP(C) is a ubiquitous cell surface protein, abundantly expressed in neurons, which constitute the targets of PrP(Sc)-mediated toxicity. Converging evidence have highlighted that neuronal, GPI-anchored PrP(C) is absolutely required for prion-induced neuropathogenesis, which warrants investigating into the normal function exerted by PrP(C) in a neuronal context. It is now well-established that PrP(C) can serve as a cell signalling molecule, able to mobilize transduction cascades in response to interactions with partners. This function endows PrP(C) with the capacity to participate in multiple neuronal processes, ranging from survival to synaptic plasticity. A diverse array of data have allowed to shed light on how this function is corrupted by PrP(Sc). Recently, amyloid Aβ oligomers, whose accumulation is associated with Alzheimer's disease (AD), were shown to similarly instigate toxic events by deviating PrP(C)-mediated signalling. Here, we provide an overview of the various signal transduction cascades ascribed to PrP(C) in neurons, summarize how their subversion by PrP(Sc) or Aβ oligomers contributes to TSE or AD neuropathogenesis and discuss the ensuing clinical implications. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Oxygen-sensitive potassium channels in chemoreceptor cell physiology: making a virtue of necessity.

PubMed

Gonzalez, Constancio; Vaquero, Luis M; López-López, José Ramón; Pérez-García, M Teresa

2009-10-01

The characterization of the molecular mechanisms involved in low-oxygen chemotransduction has been an active field of research since the first description of an oxygen-sensitive K(+) channel in rabbit carotid body (CB) chemoreceptor cells. As a result, a large number of components of the transduction cascade, from O(2) sensors to O(2)-sensitive ion channels, have been found. Although the endpoints of the process are analogous, the heterogeneity of the elements involved in the different chemoreceptor tissues precludes a unifying theory of hypoxic signaling, and it has been a source of controversy. However, when these molecular constituents of the hypoxic cascade are brought back to their physiological context, it becomes clear that the diversity of mechanisms is necessary to build up an integrated cellular response that demands the concerted action of several O(2) sensors and several effectors.

Gene regulation by the VirS/VirR system in Clostridium perfringens.

PubMed

Ohtani, Kaori

2016-10-01

The Gram-positive anaerobic spore-forming rod, Clostridium perfringens, is widely distributed in nature, especially in soil and the gastrointestinal tract of humans and animals. C. perfringens produces many secreted toxins and enzymes that are involved in the pathogenesis of gas gangrane and gastrointestinal disease. One of the most important systems regulating the production of these proteins in C. perfringens is the VirS/VirR-VR-RNA signal transduction cascade. The Agr system also important for the regulation of toxin genes. VirS appears to sense the peptide produced by the Agr (accessory gene regulator) system. The VirS/VirR-VR-RNA cascade controls the pathogenesis of C. perfringens infections by regulating virulence related genes and genes for energy metabolism. These systems are important for the host cell-induced upregulation of toxin production. Copyright © 2016 Elsevier Ltd. All rights reserved.

Mitogen-Activated Protein Kinase Signaling in Plant-Interacting Fungi: Distinct Messages from Conserved Messengers[W

PubMed Central

Hamel, Louis-Philippe; Nicole, Marie-Claude; Duplessis, Sébastien; Ellis, Brian E.

2012-01-01

Mitogen-activated protein kinases (MAPKs) are evolutionarily conserved proteins that function as key signal transduction components in fungi, plants, and mammals. During interaction between phytopathogenic fungi and plants, fungal MAPKs help to promote mechanical and/or enzymatic penetration of host tissues, while plant MAPKs are required for activation of plant immunity. However, new insights suggest that MAPK cascades in both organisms do not operate independently but that they mutually contribute to a highly interconnected molecular dialogue between the plant and the fungus. As a result, some pathogenesis-related processes controlled by fungal MAPKs lead to the activation of plant signaling, including the recruitment of plant MAPK cascades. Conversely, plant MAPKs promote defense mechanisms that threaten the survival of fungal cells, leading to a stress response mediated in part by fungal MAPK cascades. In this review, we make use of the genomic data available following completion of whole-genome sequencing projects to analyze the structure of MAPK protein families in 24 fungal taxa, including both plant pathogens and mycorrhizal symbionts. Based on conserved patterns of sequence diversification, we also propose the adoption of a unified fungal MAPK nomenclature derived from that established for the model species Saccharomyces cerevisiae. Finally, we summarize current knowledge of the functions of MAPK cascades in phytopathogenic fungi and highlight the central role played by MAPK signaling during the molecular dialogue between plants and invading fungal pathogens. PMID:22517321

Signaling intermediates (MAPK and PI3K) as therapeutic targets in NSCLC.

PubMed

Ciuffreda, Ludovica; Incani, Ursula Cesta; Steelman, Linda S; Abrams, Stephen L; Falcone, Italia; Curatolo, Anais Del; Chappell, William H; Franklin, Richard A; Vari, Sabrina; Cognetti, Francesco; McCubrey, James A; Milella, Michele

2014-01-01

The RAS/RAF/MEK/ ERK and the PI3K/AKT/mTOR pathways govern fundamental physiological processes, such as cell proliferation, differentiation, metabolism, cytoskeleton reorganization and cell death and survival. Constitutive activation of these signal transduction pathways is a required hallmark of cancer and dysregulation, on either genetic or epigenetic grounds, of these pathways has been implicated in the initiation, progression and metastastic spread of lung cances. Targeting components of the MAPK and PI3K cascades is thus an attractive strategy in the development of novel therapeutic approaches to treat lung cancer, although the use of single pathway inhibitors has met with limited clinical success so far. Indeed, the presence of intra- and inter-pathway compensatory loops that re-activate the very same cascade, either upstream or downstream the point of pharmacological blockade, or activate the alternate pathway following the blockade of one signaling cascade has been demonstrated, potentially driving preclinical (and possibly clinical) resistance. Therefore, the blockade of both pathways with combinations of signaling inhibitors might result in a more efficient anti-tumor effect, and thus potentially overcome and/or delay clinical resistance, as compared with single agent. The current review aims at summarizing the current status of preclinical and clinical research with regard to pathway crosstalks between the MAPK and PI3K cascades in NSCLC and the rationale for combined therapeutic pathway targeting.

ERBB oncogene proteins as targets for monoclonal antibodies.

PubMed

Polanovski, O L; Lebedenko, E N; Deyev, S M

2012-03-01

General properties of the family of tyrosine kinase ERBB receptors are considered in connection with their role in the generation of cascades of signal transduction in normal and tumor cells. Causes of acquisition of oncogene features by genes encoding these receptors and their role in tumorigenesis are analyzed. Anti-ERBB monoclonal antibodies approved for therapy are described in detail, and mechanisms of their antitumor activity and development of resistance to them are reviewed. The existing and the most promising strategies for creating and using monoclonal antibodies and their derivatives for therapy of cancer are discussed.

Protein receptor-independent plasma membrane remodeling by HAMLET: A tumoricidal protein-lipid complex

DOE PAGES

Nadeem, Aftab; Sanborn, Jeremy; Gettel, Douglas L.; ...

2015-11-12

A central tenet of signal transduction in eukaryotic cells is that extra-cellular ligands activate specific cell surface receptors, which orchestrate downstream responses. This ‘’protein-centric” view is increasingly challenged by evidence for the involvement of specialized membrane domains in signal transduction. Here, we propose that membrane perturbation may serve as an alternative mechanism to activate a conserved cell-death program in cancer cells. This view emerges from the extraordinary manner in which HAMLET (Human Alpha-lactalbumin Made LEthal to Tumor cells) kills a wide range of tumor cells in vitro and demonstrates therapeutic efficacy and selectivity in cancer models and clinical studies. Wemore » identify a ‘’receptor independent” transformation of vesicular motifs in model membranes, which is paralleled by gross remodeling of tumor cell membranes. Furthermore, we find that HAMLET accumulates within these de novo membrane conformations and define membrane blebs as cellular compartments for direct interactions of HAMLET with essential target proteins such as the Ras family of GTPases. In conclusion, we demonstrate lower sensitivity of healthy cell membranes to HAMLET challenge. These features suggest that HAMLET-induced curvature-dependent membrane conformations serve as surrogate receptors for initiating signal transduction cascades, ultimately leading to cell death.« less

Plasma membrane order and fluidity are diversely triggered by elicitors of plant defence

PubMed Central

Sandor, Roman; Der, Christophe; Grosjean, Kevin; Anca, Iulia; Noirot, Elodie; Leborgne-Castel, Nathalie; Lochman, Jan; Simon-Plas, Françoise; Gerbeau-Pissot, Patricia

2016-01-01

Although plants are exposed to a great number of pathogens, they usually defend themselves by triggering mechanisms able to limit disease development. Alongside signalling events common to most such incompatible interactions, modifications of plasma membrane (PM) physical properties could be new players in the cell transduction cascade. Different pairs of elicitors (cryptogein, oligogalacturonides, and flagellin) and plant cells (tobacco and Arabidopsis) were used to address the issue of possible modifications of plant PM biophysical properties induced by elicitors and their links to other events of the defence signalling cascade. We observed an increase of PM order whatever the elicitor/plant cell pair used, provided that a signalling cascade was induced. Such membrane modification is dependent on the NADPH oxidase-mediated reactive oxygen species production. Moreover, cryptogein, which is the sole elicitor able to trap sterols, is also the only one able to trigger an increase in PM fluidity. The use of cryptogein variants with altered sterol-binding properties confirms the strong correlation between sterol removal from the PM and PM fluidity enhancement. These results propose PM dynamics as a player in early signalling processes triggered by elicitors of plant defence. PMID:27604805

Cytotoxic Effect of Nano-SiO2 in Human Breast Cancer Cells via Modulation of EGFR Signaling Cascades.

PubMed

Jeon, Donghwan; Kim, Hyungjoo; Nam, Keesoo; Oh, Sunhwa; Son, Seog-Ho; Shin, Incheol

2017-11-01

Silica nanoparticles (nano-SiO 2 ) are widely used in many industrial areas and there is much controversy surrounding cytotoxic effects of such nanoparticles. In order to determine the toxicity and possible molecular mechanisms involved, we conducted several tests with two breast cancer cell lines, MDA-MB-231 and Hs578T. After exposure to nano-SiO 2 , growth, apoptosis, motility of breast cancer cells were monitored. In addition, modulation of signal transduction induced by nano-SiO 2 was detected through western blot analysis. Treatment of nano-SiO 2 repressed the growth of breast cancer cell lines. It also increased apoptosis and reduced cell motility. Moreover, exposure to nano-SiO 2 significantly disturbed the dimerization of epidermal growth factor receptor (EGFR), followed by down-regulation of its downstream cellular sarcoma kinase (c-SRC) and signal transducer and activator of transcription 3 (STAT3) signaling cascades. Nano-SiO 2 has a cytotoxic effect on MDA-MB-231 and Hs578T breast cancer cells via modulation of EGFR signaling cascades. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Visual Cortex Plasticity: A Complex Interplay of Genetic and Environmental Influences

PubMed Central

Maya-Vetencourt, José Fernando; Origlia, Nicola

2012-01-01

The central nervous system architecture is highly dynamic and continuously modified by sensory experience through processes of neuronal plasticity. Plasticity is achieved by a complex interplay of environmental influences and physiological mechanisms that ultimately activate intracellular signal transduction pathways regulating gene expression. In addition to the remarkable variety of transcription factors and their combinatorial interaction at specific gene promoters, epigenetic mechanisms that regulate transcription have emerged as conserved processes by which the nervous system accomplishes the induction of plasticity. Experience-dependent changes of DNA methylation patterns and histone posttranslational modifications are, in fact, recruited as targets of plasticity-associated signal transduction mechanisms. Here, we shall concentrate on structural and functional consequences of early sensory deprivation in the visual system and discuss how intracellular signal transduction pathways associated with experience regulate changes of chromatin structure and gene expression patterns that underlie these plastic phenomena. Recent experimental evidence for mechanisms of cross-modal plasticity following congenital or acquired sensory deprivation both in human and animal models will be considered as well. We shall also review different experimental strategies that can be used to achieve the recovery of sensory functions after long-term deprivation in humans. PMID:22852098

Teaching an old hormone new tricks: cytosolic Ca2+ elevation involvement in plant brassinosteroid signal transduction cascades.

PubMed

Zhao, Yichen; Qi, Zhi; Berkowitz, Gerald A

2013-10-01

Brassinosteroids (BRs) are hormones that control many aspects of plant growth and development, acting at the cell level to promote division and expansion. BR regulation of plant and plant cell function occurs through altered expression of many genes. Transcriptional reprogramming downstream from cell perception of this hormone is currently known to be mediated by a phosphorylation/dephosphorylation ("phosphorelay") cascade that alters the stability of two master transcription regulators. Here, we provide evidence that BR perception by their receptor also causes an elevation in cytosolic Ca(2+), initiating a Ca(2+) signaling cascade in Arabidopsis (Arabidopsis thaliana) cell cytosol. BR-dependent increases in the expression of some genes (INDOLE-3-ACETIC ACID-INDUCIBLE1 and PHYTOCHROME B ACTIVATION-TAGGED SUPPRESSOR1) were impaired in wild-type plants by a Ca(2+) channel blocker and also in the defense-no-death (dnd1) mutant, which lacks a functional cyclic GMP-activated cell membrane Ca(2+)-conducting channel. Alternatively, mutations that impair the BR phosphorelay cascade did not much affect the BR-dependent expression of these genes. Similar effects of the Ca(2+) channel blocker and dnd1 mutation were observed on a BR plant growth phenotype, deetiolation of the seedling hypocotyl. Further evidence presented in this report suggests that a BR-dependent elevation in cyclic GMP may be involved in the Ca(2+) signaling cascade initiated by this hormone. The work presented here leads to a new model of the molecular steps that mediate some of the cell responses to this plant hormone.

Loss of CDKL5 disrupts kinome profile and event-related potentials leading to autistic-like phenotypes in mice.

PubMed

Wang, I-Ting Judy; Allen, Megan; Goffin, Darren; Zhu, Xinjian; Fairless, Andrew H; Brodkin, Edward S; Siegel, Steve J; Marsh, Eric D; Blendy, Julie A; Zhou, Zhaolan

2012-12-26

Mutations in the X-linked cyclin-dependent kinase-like 5 (CDKL5) gene have been identified in neurodevelopmental disorders including atypical Rett syndrome (RTT), autism spectrum disorders (ASDs), and early infantile epileptic encephalopathy. The biological function of CDKL5 and its role in the etiology of these disorders, however, remain unclear. Here we report the development of a unique knockout mouse model of CDKL5-related disorders and demonstrate that mice lacking CDKL5 show autistic-like deficits in social interaction, as well as impairments in motor control and fear memory. Neurophysiological recordings reveal alterations in event-related potentials (ERPs) similar to those observed in RTT and ASDs. Moreover, kinome profiling uncovers disruption of multiple signal transduction pathways, including the AKT-mammalian target of rapamycin (mTOR) cascade, upon Cdkl5 loss-of-function. These data demonstrate that CDKL5 regulates signal transduction pathways and mediates autistic-like phenotypes and together establish a causal role for Cdkl5 loss-of-function in neurodevelopmental disorders.

RIP4 is a target of multiple signal transduction pathways in keratinocytes: Implications for epidermal differentiation and cutaneous wound repair

DOE Office of Scientific and Technical Information (OSTI.GOV)

Adams, Stephanie; Munz, Barbara, E-mail: barbara.munz@charite.de

2010-01-01

Receptor interacting protein 4 (RIP4) is an important regulator of epidermal morphogenesis during embryonic development. We could previously show that expression of the rip4 gene is strongly downregulated in cutaneous wound repair, which might be initiated by a broad variety of growth factors and cytokines. Here, we demonstrate that in keratinocytes, rip4 expression is controlled by a multitude of different signal transduction pathways, such as the p38 mitogen-activated protein kinase (MAPK) and the nuclear factor kappa B (NF-{kappa}B) cascade, in a unique and specific manner. Furthermore, we show that the steroid dexamethasone abolishes the physiological rip4 downregulation after injury andmore » might thus contribute to the phenotype of reduced and delayed wound reepithelialization seen in glucocorticoid-treated patients. As a whole, our data indicate that rip4 expression is regulated in a complex manner, which might have therapeutic implications.« less

Receptor for Advanced Glycation End Products and Its Involvement in Inflammatory Diseases

PubMed Central

Talib, Herni; Tie, Tung Hing; Nordin, Norshariza

2013-01-01

The receptor for advanced glycation end products (RAGE) is a transmembrane receptor of the immunoglobulin superfamily, capable of binding a broad repertoire of ligands. RAGE-ligands interaction induces a series of signal transduction cascades and lead to the activation of transcription factor NF-κB as well as increased expression of cytokines, chemokines, and adhesion molecules. These effects endow RAGE with the role in the signal transduction from pathogen substrates to cell activation during the onset and perpetuation of inflammation. RAGE signaling and downstream pathways have been implicated in a wide spectrum of inflammatory-related pathologic conditions such as arteriosclerosis, Alzheimer's disease, arthritis, acute respiratory failure, and sepsis. Despite the significant progress in other RAGE studies, the functional importance of the receptor in clinical situations and inflammatory diseases still remains to be fully realized. In this review, we will summarize current understandings and lines of evidence on the molecular mechanisms through which RAGE signaling contributes to the pathogenesis of the aforementioned inflammation-associated conditions. PMID:24102034

The signal transduction pathways controlling in planta tuberization in potato: an emerging synthesis.

PubMed

Sarkar, Debabrata

2008-01-01

Tuberization is one of the multiple outputs of a single-input phytochrome B sensory system, involving several regulatory genes. Phytochrome B- and GA-mediated photoperiodic perception occurs in the leaf, and then the RNA acts as a systemic signal in the long-distance signaling pathway to initiate tuberization in the subapical region of an underground stolon. There is good evidence that flowering and tuberizing signals might be similar. Is there a cross-talk with an oxidative burst-mediated redox signaling pathway during tuberization? Is the lipoxygenase cascade involved in the formation of the perimedullary tissue in a growing tuber? Do aquaporins regulate cell division, expansion and elongation during stolon growth and tuber induction in potato? Is the adaptive diversity for tuberization under varying photoperiods a micro-evolutionary indicator of differential transduction of cell-to-cell signal molecules under spatial and temporal expression of regulatory genes encoding transcriptional activators? Taking these views into consideration, the review presents an interim synthesis of a signaling network regulating in planta tuberization in potato.

Investigation of Flow Separation in a Transonic-fan Linear Cascade Using Visualization Methods

NASA Technical Reports Server (NTRS)

Lepicovsky, Jan; Chima, Rodrick V.; Jett, Thomas A.; Bencic, Timothy J.; Weiland, Kenneth E.

2000-01-01

An extensive study into the nature of the separated flows on the suction side of modem transonic fan airfoils at high incidence is described in the paper. Suction surface.flow separation is an important flow characteristic that may significantly contribute to stall flutter in transonic fans. Flutter in axial turbomachines is a highly undesirable and dangerous self-excited mode of blade oscillations that can result in high cycle fatigue blade failure. The study basically focused on two visualization techniques: surface flow visualization using dye oils, and schlieren (and shadowgraph) flow visualization. The following key observations were made during the study. For subsonic inlet flow, the flow on the suction side of the blade is separated over a large portion of the blade, and the separated area increases with increasing inlet Mach number. For the supersonic inlet flow condition, the flow is attached from the leading edge up to the point where a bow shock from the upper neighboring blade hits the blade surface. Low cascade solidity, for the subsonic inlet flow, results in an increased area of separated flow. For supersonic flow conditions, a low solidity results in an improvement in flow over the suction surface. Finally, computational results modeling the transonic cascade flowfield illustrate our ability to simulate these flows numerically.

Novel Molecular Strategies and Targets for Opioid Drug Discovery for the Treatment of Chronic Pain

PubMed Central

Olson, Keith M.; Lei, Wei; Keresztes, Attila; LaVigne, Justin; Streicher, John M.

2017-01-01

Opioid drugs like morphine and fentanyl are the gold standard for treating moderate to severe acute and chronic pain. However, opioid drug use can be limited by serious side effects, including constipation, tolerance, respiratory suppression, and addiction. For more than 100 years, we have tried to develop opioids that decrease or eliminate these liabilities, with little success. Recent advances in understanding opioid receptor signal transduction have suggested new possibilities to activate the opioid receptors to cause analgesia, while reducing or eliminating unwanted side effects. These new approaches include designing functionally selective ligands, which activate desired signaling cascades while avoiding signaling cascades that are thought to provoke side effects. It may also be possible to directly modulate downstream signaling through the use of selective activators and inhibitors. Separate from downstream signal transduction, it has also been found that when the opioid system is stimulated, various negative feedback systems are upregulated to compensate, which can drive side effects. This has led to the development of multi-functional molecules that simultaneously activate the opioid receptor while blocking various negative feedback receptor systems including cholecystokinin and neurokinin-1. Other novel approaches include targeting heterodimers of the opioid and other receptor systems which may drive side effects, and making endogenous opioid peptides druggable, which may also reduce opioid mediated side effects. Taken together, these advances in our molecular understanding provide a path forward to break the barrier in producing an opioid with reduced or eliminated side effects, especially addiction, which may provide relief for millions of patients. PMID:28356897

Cholinergic epithelial cell with chemosensory traits in murine thymic medulla.

PubMed

Panneck, Alexandra Regina; Rafiq, Amir; Schütz, Burkhard; Soultanova, Aichurek; Deckmann, Klaus; Chubanov, Vladimir; Gudermann, Thomas; Weihe, Eberhard; Krasteva-Christ, Gabriela; Grau, Veronika; del Rey, Adriana; Kummer, Wolfgang

2014-12-01

Specialized epithelial cells with a tuft of apical microvilli ("brush cells") sense luminal content and initiate protective reflexes in response to potentially harmful substances. They utilize the canonical taste transduction cascade to detect "bitter" substances such as bacterial quorum-sensing molecules. In the respiratory tract, most of these cells are cholinergic and are approached by cholinoceptive sensory nerve fibers. Utilizing two different reporter mouse strains for the expression of choline acetyltransferase (ChAT), we observed intense labeling of a subset of thymic medullary cells. ChAT expression was confirmed by in situ hybridization. These cells showed expression of villin, a brush cell marker protein, and ultrastructurally exhibited lateral microvilli. They did not express neuroendocrine (chromogranin A, PGP9.5) or thymocyte (CD3) markers but rather thymic epithelial (CK8, CK18) markers and were immunoreactive for components of the taste transduction cascade such as Gα-gustducin, transient receptor potential melastatin-like subtype 5 channel (TRPM5), and phospholipase Cβ2. Reverse transcription and polymerase chain reaction confirmed the expression of Gα-gustducin, TRPM5, and phospholipase Cβ2. Thymic "cholinergic chemosensory cells" were often in direct contact with medullary epithelial cells expressing the nicotinic acetylcholine receptor subunit α3. These cells have recently been identified as terminally differentiated epithelial cells (Hassall's corpuscle-like structures in mice). Contacts with nerve fibers (identified by PGP9.5 and CGRP antibodies), however, were not observed. Our data identify, in the thymus, a previously unrecognized presumptive chemosensitive cell that probably utilizes acetylcholine for paracrine signaling. This cell might participate in intrathymic infection-sensing mechanisms.

Cilia- and Flagella-Associated Protein 69 Regulates Olfactory Transduction Kinetics in Mice

PubMed Central

Dong, Frederick N.

2017-01-01

Animals detect odorous chemicals through specialized olfactory sensory neurons (OSNs) that transduce odorants into neural electrical signals. We identified a novel and evolutionarily conserved protein, cilia- and flagella-associated protein 69 (CFAP69), in mice that regulates olfactory transduction kinetics. In the olfactory epithelium, CFAP69 is enriched in OSN cilia, where olfactory transduction occurs. Bioinformatic analysis suggests that a large portion of CFAP69 can form Armadillo-type α-helical repeats, which may mediate protein–protein interactions. OSNs lacking CFAP69, remarkably, displayed faster kinetics in both the on and off phases of electrophysiological responses at both the neuronal ensemble level as observed by electroolfactogram and the single-cell level as observed by single-cell suction pipette recordings. In single-cell analysis, OSNs lacking CFAP69 showed faster response integration and were able to fire APs more faithfully to repeated odor stimuli. Furthermore, both male and female mutant mice that specifically lack CFAP69 in OSNs exhibited attenuated performance in a buried food pellet test when a background of the same odor to the food pellet was present even though they should have better temporal resolution of coding olfactory stimulation at the peripheral. Therefore, the role of CFAP69 in the olfactory system seems to be to allow the olfactory transduction machinery to work at a precisely regulated range of response kinetics for robust olfactory behavior. SIGNIFICANCE STATEMENT Sensory receptor cells are generally thought to evolve to respond to sensory cues as fast as they can. This idea is consistent with mutational analyses in various sensory systems, where mutations of sensory receptor cells often resulted in reduced response size and slowed response kinetics. Contrary to this idea, we have found that there is a kinetic “damper” present in the olfactory transduction cascade of the mouse that slows down the response kinetics and, by doing so, it reduces the peripheral temporal resolution in coding odor stimuli and allows for robust olfactory behavior. This study should trigger a rethinking of the significance of the intrinsic speed of sensory transduction and the pattern of the peripheral coding of sensory stimuli. PMID:28495971

Lateral distortions of electromagnetic cascades in emulsion chambers

NASA Technical Reports Server (NTRS)

Porter, L. G.; Levit, L. B.; Jones, W. V.; Huggett, R. W.; Barrowes, S. C.

1975-01-01

Electromagnetic cascades in a lead-emulsion chamber have been studied to determine the effect of air gaps on the upstream sides of the emulsions. Such air gaps cause a change in the form of the radial distribution of electron tracks, making cascades appear older and giving incorrect energy estimates. The number of tracks remaining within a radius r was found to vary as exp(-g/G), where g is the gap thickness. The characteristic gap thickness in mm is G = 3.04 + 1.30 ln (Err per GeV per sq mm) where E is the energy of the initiating gamma ray. Use of this relation provides a significant correction to cascade-energy estimates and allows one to calculate the effect of different gap thicknesses on the energy threshold for visual detection of cascades.

CRT--Cascade Routing Tool to define and visualize flow paths for grid-based watershed models

USGS Publications Warehouse

Henson, Wesley R.; Medina, Rose L.; Mayers, C. Justin; Niswonger, Richard G.; Regan, R.S.

2013-01-01

The U.S. Geological Survey Cascade Routing Tool (CRT) is a computer application for watershed models that include the coupled Groundwater and Surface-water FLOW model, GSFLOW, and the Precipitation-Runoff Modeling System (PRMS). CRT generates output to define cascading surface and shallow subsurface flow paths for grid-based model domains. CRT requires a land-surface elevation for each hydrologic response unit (HRU) of the model grid; these elevations can be derived from a Digital Elevation Model raster data set of the area containing the model domain. Additionally, a list is required of the HRUs containing streams, swales, lakes, and other cascade termination features along with indices that uniquely define these features. Cascade flow paths are determined from the altitudes of each HRU. Cascade paths can cross any of the four faces of an HRU to a stream or to a lake within or adjacent to an HRU. Cascades can terminate at a stream, lake, or HRU that has been designated as a watershed outflow location.

Signaling mechanisms of apoptosis-like programmed cell death in unicellular eukaryotes.

PubMed

Shemarova, Irina V

2010-04-01

In unicellular eukaryotes, apoptosis-like cell death occurs during development, aging and reproduction, and can be induced by environmental stresses and exposure to toxic agents. The essence of the apoptotic machinery in unicellular organisms is similar to that in mammals, but the apoptotic signal network is less complex and of more ancient origin. The review summarizes current data about key apoptotic proteins and mechanisms of the transduction of apoptotic signals by caspase-like proteases and mitochondrial apoptogenic proteins in unicellular eukaryotes. The roles of receptor-dependent and receptor-independent caspase cascades are reviewed. 2010 Elsevier Inc. All rights reserved.

The JAK-STAT signaling pathway: input and output integration.

PubMed

Murray, Peter J

2007-03-01

Universal and essential to cytokine receptor signaling, the JAK-STAT pathway is one of the best understood signal transduction cascades. Almost 40 cytokine receptors signal through combinations of four JAK and seven STAT family members, suggesting commonality across the JAK-STAT signaling system. Despite intense study, there remain substantial gaps in understanding how the cascades are activated and regulated. Using the examples of the IL-6 and IL-10 receptors, I will discuss how diverse outcomes in gene expression result from regulatory events that effect the JAK1-STAT3 pathway, common to both receptors. I also consider receptor preferences by different STATs and interpretive problems in the use of STAT-deficient cells and mice. Finally, I consider how the suppressor of cytokine signaling (SOCS) proteins regulate the quality and quantity of STAT signals from cytokine receptors. New data suggests that SOCS proteins introduce additional diversity into the JAK-STAT pathway by adjusting the output of activated STATs that alters downstream gene activation.

Modulation of learning and memory by cytokines: signaling mechanisms and long term consequences.

PubMed

Donzis, Elissa J; Tronson, Natalie C

2014-11-01

This review describes the role of cytokines and their downstream signaling cascades on the modulation of learning and memory. Immune proteins are required for many key neural processes and dysregulation of these functions by systemic inflammation can result in impairments of memory that persist long after the resolution of inflammation. Recent research has demonstrated that manipulations of individual cytokines can modulate learning, memory, and synaptic plasticity. The many conflicting findings, however, have prevented a clear understanding of the precise role of cytokines in memory. Given the complexity of inflammatory signaling, understanding its modulatory role requires a shift in focus from single cytokines to a network of cytokine interactions and elucidation of the cytokine-dependent intracellular signaling cascades. Finally, we propose that whereas signal transduction and transcription may mediate short-term modulation of memory, long-lasting cellular and molecular mechanisms such as epigenetic modifications and altered neurogenesis may be required for the long lasting impact of inflammation on memory and cognition. Copyright © 2014 Elsevier Inc. All rights reserved.

Raf kinase inhibitory protein: a signal transduction modulator and metastasis suppressor.

PubMed

Granovsky, Alexey E; Rosner, Marsha Rich

2008-04-01

Cells have a multitude of controls to maintain their integrity and prevent random switching from one biological state to another. Raf Kinase Inhibitory Protein (RKIP), a member of the phosphatidylethanolamine binding protein (PEBP) family, is representative of a new class of modulators of signaling cascades that function to maintain the "yin yang" or balance of biological systems. RKIP inhibits MAP kinase (Raf-MEK-ERK), G protein-coupled receptor (GPCR) kinase and NFkappaB signaling cascades. Because RKIP targets different kinases dependent upon its state of phosphorylation, RKIP also acts to integrate crosstalk initiated by multiple environmental stimuli. Loss or depletion of RKIP results in disruption of the normal cellular stasis and can lead to chromosomal abnormalities and disease states such as cancer. Since RKIP and the PEBP family have been reviewed previously, the goal of this analysis is to provide an update and highlight some of the unique features of RKIP that make it a critical player in the regulation of cellular signaling processes.

Ubiquitin-Modifying Enzymes and Regulation of the Inflammasome.

PubMed

Kattah, Michael G; Malynn, Barbara A; Ma, Averil

2017-11-10

Ubiquitin and ubiquitin-modifying enzymes play critical roles in a wide variety of intracellular signaling pathways. Inflammatory signaling cascades downstream of TNF, TLR agonists, antigen receptor cross-linking, and cytokine receptors, all rely on ubiquitination events to direct subsequent immune responses. In the past several years, inflammasome activation and subsequent signal transduction have emerged as an excellent example of how ubiquitin signals control inflammatory responses. Inflammasomes are multiprotein signaling complexes that ultimately lead to caspase activation and release of the interleukin-1 (IL-1) family members, IL-1β and IL-18. Inflammasome activation is critical for the host's defense against pathogens, but dysregulation of inflammasomes may contribute to the pathogenesis of multiple diseases. Ultimately, understanding how various ubiquitin interacting proteins control inflammatory signaling cascades could provide new pathways for therapeutic intervention. Here we review specific ubiquitin-modifying enzymes and ubiquitination events that orchestrate inflammatory responses, with an emphasis on the NLRP3 inflammasome. Copyright © 2017 Elsevier Ltd. All rights reserved.

Identification of specific gravity sensitive signal transduction pathways in human A431 carcinoma cells

NASA Astrophysics Data System (ADS)

Rijken, P. J.; de Groot, R. P.; Kruijer, W.; de Laat, S. W.; Verkleij, A. J.; Boonstra, J.

Epidermal growth factor (EGF) activates a well characterized signal transduction cascade in human A431 epidermoid carcinoma cells. The influence of gravity on EGF-induced EGF-receptor clustering and early gene expression as well as on actin polymerization and actin organization have been investigated. Different signalling pathways induced by the agents TPA, forskolin and A23187 that activate gene expression were tested for sensitivity to gravity. EGF-induced c-fos and c-jun expression were decreased in microgravity. However, constitutive β-2 microglobulin expression remained unaltered. Under simulated weightlessness conditions EGF- and TPA-induced c-fos expression was decreased, while forskolin- and A23187-induced c-fos expression was independent of the gravity conditions. These results suggest that gravity affects specific signalling pathways. Preliminary results indicate that EGF-induced EGF-receptor clustering remained unaltered irrespective of the gravity conditions. Furthermore, the relative filamentous actin content of steady state A431 cells was enhanced under microgravity conditions and actin filament organization was altered. Under simulated weightlessness actin filament organization in steady state cells as well as in EGF-treated cells was altered as compared to the 1 G reference experiment. Interestingly the microtubule and keratin organization in untreated cells showed no difference with the normal gravity samples. This indicates that gravity may affect specific components of the signal transduction circuitry.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stoiber, Marcus; Celniker, Susan; Cherbas, Lucy

Steroid hormones induce cascades of gene activation and repression with transformative effects on cell fate . Steroid transduction plays a major role in the development and physiology of nearly all metazoan species, and in the progression of the most common forms of cancer. Despite the paramount importance of steroids in developmental and translational biology, a complete map of transcriptional response has not been developed for any hormone . In the case of 20-hydroxyecdysone (ecdysone) in Drosophila melanogaster, these trajectories range from apoptosis to immortalization. We mapped the ecdysone transduction network in a cohort of 41 cell lines, the largest suchmore » atlas yet assembled. We found that the early transcriptional response mirrors the distinctiveness of physiological origins: genes respond in restricted patterns, conditional on the expression levels of dozens of transcription factors. Only a small cohort of genes is constitutively modulated independent of initial cell state. Ecdysone-responsive genes tend to organize into directional same-stranded units, with consecutive genes induced from the same strand. Here, we identify half of the ecdysone receptor heterodimer as the primary rate-limiting step in the response, and find that initial receptor isoform levels modulate the activated cohort of target transcription factors. In conclusion, this atlas of steroid response reveals organizing principles of gene regulation by a model type II nuclear receptor and lays the foundation for comprehensive and predictive understanding of the ecdysone transduction network in the fruit fly.« less

The attentive brain: insights from developmental cognitive neuroscience.

PubMed

Amso, Dima; Scerif, Gaia

2015-10-01

Visual attention functions as a filter to select environmental information for learning and memory, making it the first step in the eventual cascade of thought and action systems. Here, we review studies of typical and atypical visual attention development and explain how they offer insights into the mechanisms of adult visual attention. We detail interactions between visual processing and visual attention, as well as the contribution of visual attention to memory. Finally, we discuss genetic mechanisms underlying attention disorders and how attention may be modified by training.

Fast response and high sensitivity to microsaccades in a cascading-adaptation neural network with short-term synaptic depression

NASA Astrophysics Data System (ADS)

Yuan, Wu-Jie; Zhou, Jian-Fang; Zhou, Changsong

2016-04-01

Microsaccades are very small eye movements during fixation. Experimentally, they have been found to play an important role in visual information processing. However, neural responses induced by microsaccades are not yet well understood and are rarely studied theoretically. Here we propose a network model with a cascading adaptation including both retinal adaptation and short-term depression (STD) at thalamocortical synapses. In the neural network model, we compare the microsaccade-induced neural responses in the presence of STD and those without STD. It is found that the cascading with STD can give rise to faster and sharper responses to microsaccades. Moreover, STD can enhance response effectiveness and sensitivity to microsaccadic spatiotemporal changes, suggesting improved detection of small eye movements (or moving visual objects). We also explore the mechanism of the response properties in the model. Our studies strongly indicate that STD plays an important role in neural responses to microsaccades. Our model considers simultaneously retinal adaptation and STD at thalamocortical synapses in the study of microsaccade-induced neural activity, and may be useful for further investigation of the functional roles of microsaccades in visual information processing.

Partial reconstitution of photoreceptor cGMP phosphodiesterase characteristics in cGMP phosphodiesterase-5.

PubMed

Granovsky, A E; Artemyev, N O

2001-06-15

Photoreceptor cGMP phosphodiesterases (PDE6) are uniquely qualified to serve as effector enzymes in the vertebrate visual transduction cascade. In the dark-adapted photoreceptors, the activity of PDE6 is blocked via tight association with the inhibitory gamma-subunits (Pgamma). The Pgamma block is removed in the light-activated PDE6 by the visual G protein, transducin. Transducin-activated PDE6 exhibits an exceptionally high catalytic rate of cGMP hydrolysis ensuring high signal amplification. To identify the structural determinants for the inhibitory interaction with Pgamma and the remarkable cGMP hydrolytic ability, we sought to reproduce the PDE6 characteristics by mutagenesis of PDE5, a related cyclic GMP-specific, cGMP-binding PDE. PDE5 is insensitive to Pgamma and has a more than 100-fold lower k(cat) for cGMP hydrolysis. Our mutational analysis of chimeric PDE5/PDE6alpha' enzymes revealed that the inhibitory interaction of cone PDE6 catalytic subunits (PDE6alpha') with Pgamma is mediated primarily by three hydrophobic residues at the entry to the catalytic pocket, Met(758), Phe(777), and Phe(781). The maximal catalytic rate of PDE5 was enhanced by at least 10-fold with substitutions of PDE6alpha'-specific glycine residues for the corresponding PDE5 alanine residues, Ala(608) and Ala(612). The Gly residues are adjacent to the highly conserved metal binding motif His-Asn-X-X-His, which is essential for cGMP hydrolysis. Our results suggest that the unique Gly residues allow the PDE6 metal binding site to adopt a more favorable conformation for cGMP hydrolysis.

IGF-1 signaling mediated cell-specific skeletal mechano-transduction.

PubMed

Tian, Faming; Wang, Yongmei; Bikle, Daniel D

2018-02-01

Mechanical loading preserves bone mass and stimulates bone formation, whereas skeletal unloading leads to bone loss. In addition to osteocytes, which are considered the primary sensor of mechanical load, osteoblasts, and bone specific mesenchymal stem cells also are involved. The skeletal response to mechanical signals is a complex process regulated by multiple signaling pathways including that of insulin-like growth factor-1 (IGF-1). Conditional osteocyte deletion of IGF-1 ablates the osteogenic response to mechanical loading. Similarly, osteocyte IGF-1 receptor (IGF-1R) expression is necessary for reloading-induced periosteal bone formation. Transgenic overexpression of IGF-1 in osteoblasts results in enhanced responsiveness to in vivo mechanical loading in mice, a response which is eliminated by osteoblastic conditional disruption of IGF-1 in vivo. Bone marrow derived stem cells (BMSC) from unloaded bone fail to respond to IGF-1 in vitro. IGF-1R is required for the transduction of a mechanical stimulus to downstream effectors, transduction which is lost when the IGF-1R is deleted. Although the molecular mechanisms are not yet fully elucidated, the IGF signaling pathway and its interactions with potentially interlinked signaling cascades involving integrins, the estrogen receptor, and wnt/β-catenin play an important role in regulating adaptive response of cancer bone cells to mechanical stimuli. In this review, we discuss recent advances investigating how IGF-1 and other interlinked molecules and signaling pathways regulate skeletal mechano-transduction involving different bone cells, providing an overview of the IGF-1 signaling mediated cell-specific response to mechanical stimuli. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:576-583, 2018. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

Biologically active leptin-related synthetic peptides activate STAT3 via phosphorylation of ERK1/2 and PI-3K.

PubMed

Lin, Hung-Yun; Yang, Sheng-Huei; Tang, Heng-Yuan; Cheng, Guei-Yun; Davis, Paul J; Grasso, Patricia

2014-07-01

The effects of leptin-related synthetic peptides [d-Leu-4]-OB3 and OB3 on energy balance and glucose homeostasis in ob/ob and db/db mice have been confirmed. The molecular basis of these effects, however, remains unclear. In the present study, we examined the ability of these peptides to activate signal transduction pathways known to be involved in transduction of the leptin signal. In a specific and concentration-dependent manner, [d-Leu-4]-OB3 induced phosphorylation of ERK1/2, PI-3K, Ser-727 STAT3, and Tyr-705 of STAT3. OB3 also induced activation of STAT3 via phosphorylation of ERK1/2, STAT3 Ser-727, STAT3 Tyr-705 and PI-3K p85, but to a lesser degree. Using PD98059 and LY294002, specific inhibitors of MEK and PI-3K, respectively, we were able to identify the signal transduction pathways involved in peptide-induced STAT3 activation. [d-Leu-4]-OB3 induced serine phosphorylation of STAT3 primarily through activation of ERK1/2. Tyrosine phosphorylation of STAT3, however, was induced primarily through activation of PI-3K. Our data suggest that in db/db mice, [d-Leu-4]-OB3 binding to short isoforms of the leptin receptor induces intracellular signaling cascades which do not require OB-Rb activation. These signals may ultimately result in peptide effects on transcriptional and translational events associated with energy balance and glycemic regulation. In summary, we have shown for the first time that, similar to leptin, bioactive leptin-related synthetic peptide analogs activate STAT3 via phosphorylation of serine and tyrosine residues by multiple signal transduction pathways. Copyright © 2014 Elsevier Inc. All rights reserved.

Plasma membrane order and fluidity are diversely triggered by elicitors of plant defence.

PubMed

Sandor, Roman; Der, Christophe; Grosjean, Kevin; Anca, Iulia; Noirot, Elodie; Leborgne-Castel, Nathalie; Lochman, Jan; Simon-Plas, Françoise; Gerbeau-Pissot, Patricia

2016-09-01

Although plants are exposed to a great number of pathogens, they usually defend themselves by triggering mechanisms able to limit disease development. Alongside signalling events common to most such incompatible interactions, modifications of plasma membrane (PM) physical properties could be new players in the cell transduction cascade. Different pairs of elicitors (cryptogein, oligogalacturonides, and flagellin) and plant cells (tobacco and Arabidopsis) were used to address the issue of possible modifications of plant PM biophysical properties induced by elicitors and their links to other events of the defence signalling cascade. We observed an increase of PM order whatever the elicitor/plant cell pair used, provided that a signalling cascade was induced. Such membrane modification is dependent on the NADPH oxidase-mediated reactive oxygen species production. Moreover, cryptogein, which is the sole elicitor able to trap sterols, is also the only one able to trigger an increase in PM fluidity. The use of cryptogein variants with altered sterol-binding properties confirms the strong correlation between sterol removal from the PM and PM fluidity enhancement. These results propose PM dynamics as a player in early signalling processes triggered by elicitors of plant defence. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Effect of the synthetic cannabinoid HU-210 on quorum sensing and on the production of quorum sensing-mediated virulence factors by Vibrio harveyi.

PubMed

Soni, Divya; Smoum, Reem; Breuer, Aviva; Mechoulam, Raphael; Steinberg, Doron

2015-08-12

Bacterial populations communicate through the cell density-dependent mechanism of quorum sensing (QS). Vibrio harveyi, one of the best studied model organisms for QS, was used to explore effects of the synthetic cannabinoid HU-210 on QS and different QS-regulated physiological processes in bacteria. Analysis of QS-regulated bioluminescence in wild-type and mutant strains of V. harveyi revealed that HU-210 affects the autoinducer-2 (AI-2) pathway, one of three known QS cascades of V. harveyi. Furthermore, QS-mediated biofilm formation and swimming motility in the mutant strain BB152 (AI-1(-), AI-2(+)) were significantly reduced in the presence of HU-210. HU-210 inhibited QS-mediated virulence factor production without any inhibitory effect on bacterial growth. It also alters the expression of several genes, which are regulated by QS, specifically downregulating the genes of the AI-2 QS cascade. First evidence is being provided for interference of bacterial signal-transduction systems by a synthetic cannabinoid. The effect of HU-210 was specific to the AI-2 cascade in V. harveyi. AI-2 is known as a "universal autoinducer" and interference with its activity opens a broad spectrum of applications for synthetic cannabinoids in future research as a potential anti-QS agent.

Induction of the unfolded protein response by constitutive G-protein signaling in rod photoreceptor cells.

PubMed

Wang, Tian; Chen, Jeannie

2014-10-17

Phototransduction is a G-protein signal transduction cascade that converts photon absorption to a change in current at the plasma membrane. Certain genetic mutations affecting the proteins in the phototransduction cascade cause blinding disorders in humans. Some of these mutations serve as a genetic source of "equivalent light" that activates the cascade, whereas other mutations lead to amplification of the light response. How constitutive phototransduction causes photoreceptor cell death is poorly understood. We showed that persistent G-protein signaling, which occurs in rod arrestin and rhodopsin kinase knock-out mice, caused a rapid and specific induction of the PERK pathway of the unfolded protein response. These changes were not observed in the cGMP-gated channel knock-out rods, an equivalent light condition that mimics light-stimulated channel closure. Thus transducin signaling, but not channel closure, triggers rapid cell death in light damage caused by constitutive phototransduction. Additionally, we show that in the albino light damage model cell death was not associated with increase in global protein ubiquitination or unfolded protein response induction. Taken together, these observations provide novel mechanistic insights into the cell death pathway caused by constitutive phototransduction and identify the unfolded protein response as a potential target for therapeutic intervention. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Evolution of phototransduction, vertebrate photoreceptors and retina.

PubMed

Lamb, Trevor D

2013-09-01

Evidence is reviewed from a wide range of studies relevant to the evolution of vertebrate photoreceptors and phototransduction, in order to permit the synthesis of a scenario for the major steps that occurred during the evolution of cones, rods and the vertebrate retina. The ancestral opsin originated more than 700 Mya (million years ago) and duplicated to form three branches before cnidarians diverged from our own lineage. During chordate evolution, ciliary opsins (C-opsins) underwent multiple stages of improvement, giving rise to the 'bleaching' opsins that characterise cones and rods. Prior to the '2R' rounds of whole genome duplication near the base of the vertebrate lineage, 'cone' photoreceptors already existed; they possessed a transduction cascade essentially the same as in modern cones, along with two classes of opsin: SWS and LWS (short- and long-wave-sensitive). These cones appear to have made synaptic contact directly onto ganglion cells, in a two-layered retina that resembled the pineal organ of extant non-mammalian vertebrates. Interestingly, those ganglion cells appear to be descendants of microvillar photoreceptor cells. No lens was associated with this two-layered retina, and it is likely to have mediated circadian timing rather than spatial vision. Subsequently, retinal bipolar cells evolved, as variants of ciliary photoreceptors, and greatly increased the computational power of the retina. With the advent of a lens and extraocular muscles, spatial imaging information became available for central processing, and gave rise to vision in vertebrates more than 500 Mya. The '2R' genome duplications permitted the refinement of cascade components suitable for both rods and cones, and also led to the emergence of five visual opsins. The exact timing of the emergence of 'true rods' is not yet clear, but it may not have occurred until after the divergence of jawed and jawless vertebrates. Copyright © 2013 The Author. Published by Elsevier Ltd.. All rights reserved.

Evaluation of diffuse-illumination holographic cinematography in a flutter cascade

NASA Technical Reports Server (NTRS)

Decker, A. J.

1986-01-01

Since 1979, the Lewis Research Center has examined holographic cinematography for three-dimensional flow visualization. The Nd:YAG lasers used were Q-switched, double-pulsed, and frequency-doubled, operating at 20 pulses per second. The primary subjects for flow visualization were the shock waves produced in two flutter cascades. Flow visualization was by diffuse-illumination, double-exposure, and holographic interferometry. The performances of the lasers, holography, and diffuse-illumination interferometry are evaluated in single-window wind tunnels. The fringe-contrast factor is used to evaluate the results. The effects of turbulence on shock-wave visualization in a transonic flow are discussed. The depth of field for visualization of a turbulent structure is demonstrated to be a measure of the relative density and scale of that structure. Other items discussed are the holographic emulsion, tests of coherence and polarization, effects of windows and diffusers, hologram bleaching, laser configurations, influence and handling of specular reflections, modes of fringe localization, noise sources, and coherence requirements as a function of the pulse energy. Holography and diffuse illumination interferometry are also reviewed.

A visual resource management study of alternative dams, reservoirs and highway and transmission line corridors near Copper Creek, Washington

Treesearch

John Ady; Brian A. Gray; Grant R. Jones

1979-01-01

Three alternative dams have been considered by Seattle City Light for the Skagit River Narrows in the North Cascades National Recreation Area, Washington. The authors assessed the area's existing visual resources, identified three alternative highway and transmission line realignments, evaluated changes in visual character and quality for 13 different combinations...

The histone acetyltransferase p300 inhibitor C646 reduces pro-inflammatory gene expression and inhibits histone deacetylases

PubMed Central

van den Bosch, Thea; Boichenko, Alexander; Leus, Niek G. J.; Eleni Ourailidou, Maria; Wapenaar, Hannah; Rotili, Dante; Mai, Antonello; Imhof, Axel; Bischoff, Rainer; Haisma, Hidde J.; Dekker, Frank J.

2016-01-01

Lysine acetylations are reversible posttranslational modifications of histone and non-histone proteins that play important regulatory roles in signal transduction cascades and gene expression. Lysine acetylations are regulated by histone acetyltransferases as writers and histone deacetylases as erasers. Because of their role in signal transduction cascades, these enzymes are important players in inflammation. Therefore, applications of histone acetyltransferase inhibitors to reduce inflammatory responses are interesting. Among the few histone acetyltransferase inhibitors described, C646 is one of the most potent (Ki of 0.4 μM for histone acetyltransferase p300). C646 was described to regulate the NF-κB pathway; an important pathway in inflammatory responses, which is regulated by acetylation. Interestingly, this pathway has been implicated in asthma and COPD. Therefore we hypothesized that via regulation of the NF-κB signaling pathway, C646 can inhibit pro-inflammatory gene expression, and have potential for the treatment of inflammatory lung diseases. In line with this, here we demonstrate that C646 reduces pro-inflammatory gene expression in RAW264.7 murine macrophages and murine precision-cut lung slices. To unravel its effects on cellular substrates we applied mass spectrometry and found, counterintuitively, a slight increase in acetylation of histone H3. Based on this finding, and structural features of C646, we presumed inhibitory activity of C646 on histone deacetylases, and indeed found inhibition of histone deacetylases from 7 μM and higher concentrations. This indicates that C646 has potential for further development towards applications in the treatment of inflammation, however, its newly discovered lack of selectivity at higher concentrations needs to be taken into account. PMID:26718586

Deciphering complex dynamics of water counteraction around secondary structural elements of allosteric protein complex: Case study of SAP-SLAM system in signal transduction cascade

NASA Astrophysics Data System (ADS)

Samanta, Sudipta; Mukherjee, Sanchita

2018-01-01

The first hydration shell of a protein exhibits heterogeneous behavior owing to several attributes, majorly local polarity and structural flexibility as revealed by solvation dynamics of secondary structural elements. We attempt to recognize the change in complex water counteraction generated due to substantial alteration in flexibility during protein complex formation. The investigation is carried out with the signaling lymphocytic activation molecule (SLAM) family of receptors, expressed by an array of immune cells, and interacting with SLAM-associated protein (SAP), composed of one SH2 domain. All atom molecular dynamics simulations are employed to the aqueous solutions of free SAP and SLAM-peptide bound SAP. We observed that water dynamics around different secondary structural elements became highly affected as well as nicely correlated with the SLAM-peptide induced change in structural rigidity obtained by thermodynamic quantification. A few instances of contradictory dynamic features of water to the change in structural flexibility are explained by means of occluded polar residues by the peptide. For βD, EFloop, and BGloop, both structural flexibility and solvent accessibility of the residues confirm the obvious contribution. Most importantly, we have quantified enhanced restriction in water dynamics around the second Fyn-binding site of the SAP due to SAP-SLAM complexation, even prior to the presence of Fyn. This observation leads to a novel argument that SLAM induced more restricted water molecules could offer more water entropic contribution during the subsequent Fyn binding and provide enhanced stability to the SAP-Fyn complex in the signaling cascade. Finally, SLAM induced water counteraction around the second binding site of the SAP sheds light on the allosteric property of the SAP, which becomes an integral part of the underlying signal transduction mechanism.

Deciphering complex dynamics of water counteraction around secondary structural elements of allosteric protein complex: Case study of SAP-SLAM system in signal transduction cascade.

PubMed

Samanta, Sudipta; Mukherjee, Sanchita

2018-01-28

The first hydration shell of a protein exhibits heterogeneous behavior owing to several attributes, majorly local polarity and structural flexibility as revealed by solvation dynamics of secondary structural elements. We attempt to recognize the change in complex water counteraction generated due to substantial alteration in flexibility during protein complex formation. The investigation is carried out with the signaling lymphocytic activation molecule (SLAM) family of receptors, expressed by an array of immune cells, and interacting with SLAM-associated protein (SAP), composed of one SH2 domain. All atom molecular dynamics simulations are employed to the aqueous solutions of free SAP and SLAM-peptide bound SAP. We observed that water dynamics around different secondary structural elements became highly affected as well as nicely correlated with the SLAM-peptide induced change in structural rigidity obtained by thermodynamic quantification. A few instances of contradictory dynamic features of water to the change in structural flexibility are explained by means of occluded polar residues by the peptide. For βD, EFloop, and BGloop, both structural flexibility and solvent accessibility of the residues confirm the obvious contribution. Most importantly, we have quantified enhanced restriction in water dynamics around the second Fyn-binding site of the SAP due to SAP-SLAM complexation, even prior to the presence of Fyn. This observation leads to a novel argument that SLAM induced more restricted water molecules could offer more water entropic contribution during the subsequent Fyn binding and provide enhanced stability to the SAP-Fyn complex in the signaling cascade. Finally, SLAM induced water counteraction around the second binding site of the SAP sheds light on the allosteric property of the SAP, which becomes an integral part of the underlying signal transduction mechanism.

Signal transduction in primary human T lymphocytes in altered gravity during parabolic flight and clinostat experiments.

PubMed

Tauber, Svantje; Hauschild, Swantje; Paulsen, Katrin; Gutewort, Annett; Raig, Christiane; Hürlimann, Eva; Biskup, Josefine; Philpot, Claudia; Lier, Hartwin; Engelmann, Frank; Pantaleo, Antonella; Cogoli, Augusto; Pippia, Proto; Layer, Liliana E; Thiel, Cora S; Ullrich, Oliver

2015-01-01

Several limiting factors for human health and performance in microgravity have been clearly identified arising from the immune system, and substantial research activities are required in order to provide the basic information for appropriate integrated risk management. The gravity-sensitive nature of cells of the immune system renders them an ideal biological model in search for general gravity-sensitive mechanisms and to understand how the architecture and function of human cells is related to the gravitational force and therefore adapted to life on Earth. We investigated the influence of altered gravity in parabolic flight and 2D clinostat experiments on key proteins of activation and signaling in primary T lymphocytes. We quantified components of the signaling cascade 1.) in non-activated T lymphocytes to assess the "basal status" of the cascade and 2.) in the process of activation to assess the signal transduction. We found a rapid decrease of CD3 and IL-2R surface expression and reduced p-LAT after 20 seconds of altered gravity in non-activated primary T lymphocytes during parabolic flight. Furthermore, we observed decreased CD3 surface expression, reduced ZAP-70 abundance and increased histone H3-acetylation in activated T lymphocytes after 5 minutes of clinorotation and a transient downregulation of CD3 and stable downregulation of IL-2R during 60 minutes of clinorotation. CD3 and IL-2R are downregulated in primary T lymphocytes in altered gravity. We assume that a gravity condition around 1g is required for the expression of key surface receptors and appropriate regulation of signal molecules in T lymphocytes. © 2015 S. Karger AG, Basel.

Fluoro-Phenyl-Styrene-Sulfonamide, a Novel Inhibitor of σB Activity, Prevents the Activation of σB by Environmental and Energy Stresses in Bacillus subtilis

PubMed Central

Ringus, Daina L.; Gaballa, Ahmed; Helmann, John D.; Wiedmann, Martin

2013-01-01

Sigma B (σB) is an alternative sigma factor that regulates the general stress response in Bacillus subtilis and in many other Gram-positive organisms. σB activity in B. subtilis is tightly regulated via at least three distinct pathways within a complex signal transduction cascade in response to a variety of stresses, including environmental stress, energy stress, and growth at high or low temperatures. We probed the ability of fluoro-phenyl-styrene-sulfonamide (FPSS), a small-molecule inhibitor of σB activity in Listeria monocytogenes, to inhibit σB activity in B. subtilis through perturbation of signal transduction cascades under various stress conditions. FPSS inhibited the activation of σB in response to multiple categories of stress known to induce σB activity in B. subtilis. Specifically, FPSS prevented the induction of σB activity in response to energy stress, including entry into stationary phase, phosphate limitation, and azide stress. FPSS also inhibited chill induction of σB activity in a ΔrsbV strain, suggesting that FPSS does not exclusively target the RsbU and RsbP phosphatases or the anti–anti-sigma factor RsbV, all of which contribute to posttranslational regulation of σB activity. Genetic and biochemical experiments, including artificial induction of σB, analysis of the phosphorylation state of the anti–anti-sigma factor RsbV, and in vitro transcription assays, indicate that while FPSS does not bind directly to σB to inhibit activity, it appears to prevent the release of B. subtilis σB from its anti-sigma factor RsbW. PMID:23524614

Changes in Neuronal Signaling and Cell Stress Response Pathways are Associated with a Multigenic Response of Drosophila melanogaster to DDT Selection

PubMed Central

Coates, Brad S; Sun, Weilin; Clark, John M; Pittendrigh, Barry R

2017-01-01

Abstract The adaptation of insect populations to insecticidal control is a continual threat to human health and sustainable agricultural practices, but many complex genomic mechanisms involved in this adaption remain poorly understood. This study applied a systems approach to investigate the interconnections between structural and functional variance in response to dichlorodiphenyltrichloroethane (DDT) within the Drosophila melanogaster strain 91-R. Directional selection in 6 selective sweeps coincided with constitutive gene expression differences in DDT resistant flies, including the most highly upregulated transcript, Unc-115 b, which plays a central role in axon guidance, and the most highly downregulated transcript, the angiopoietin-like CG31832, which is involved in directing vascular branching and dendrite outgrowth but likely may be under trans-regulatory control. Direct functions and protein–protein interactions mediated by differentially expressed transcripts control changes in cell migration, signal transduction, and gene regulatory cascades that impact the nervous system. Although changes to cellular stress response pathways involve 8 different cytochrome P450s, stress response, and apoptosis is controlled by a multifacetted regulatory mechanism. These data demonstrate that DDT selection in 91-R may have resulted in genome-wide adaptations that impacts genetic and signal transduction pathways that converge to modify stress response, cell survival, and neurological functions. This study implicates the involvement of a multigenic mechanism in the adaptation to a chemical insecticide, which impact interconnected regulatory cascades. We propose that DDT selection within 91-R might act systemically, wherein pathway interactions function to reinforce the epistatic effects of individual adaptive changes on an additive or nonadditive basis. PMID:29211847

Loss of CDKL5 disrupts kinome profile and event-related potentials leading to autistic-like phenotypes in mice

PubMed Central

Wang, I-Ting Judy; Allen, Megan; Goffin, Darren; Zhu, Xinjian; Fairless, Andrew H.; Brodkin, Edward S.; Siegel, Steve J.; Marsh, Eric D.; Blendy, Julie A.; Zhou, Zhaolan

2012-01-01

Mutations in the X-linked cyclin-dependent kinase-like 5 (CDKL5) gene have been identified in neurodevelopmental disorders including atypical Rett syndrome (RTT), autism spectrum disorders (ASDs), and early infantile epileptic encephalopathy. The biological function of CDKL5 and its role in the etiology of these disorders, however, remain unclear. Here we report the development of a unique knockout mouse model of CDKL5-related disorders and demonstrate that mice lacking CDKL5 show autistic-like deficits in social interaction, as well as impairments in motor control and fear memory. Neurophysiological recordings reveal alterations in event-related potentials (ERPs) similar to those observed in RTT and ASDs. Moreover, kinome profiling uncovers disruption of multiple signal transduction pathways, including the AKT-mammalian target of rapamycin (mTOR) cascade, upon Cdkl5 loss-of-function. These data demonstrate that CDKL5 regulates signal transduction pathways and mediates autistic-like phenotypes and together establish a causal role for Cdkl5 loss-of-function in neurodevelopmental disorders. PMID:23236174

Role of ethylene receptors during senescence and ripening in horticultural crops

PubMed Central

Agarwal, Gaurav; Choudhary, Divya; Singh, Virendra P.; Arora, Ajay

2012-01-01

The past two decades have been rewarding in terms of deciphering the ethylene signal transduction and functional validation of the ethylene receptor and downstream genes involved in the cascade. Our knowledge of ethylene receptors and its signal transduction pathway provides us a robust platform where we can think of manipulating and regulating ethylene sensitivity by the use of genetic engineering and making transgenic. This review focuses on ethylene perception, receptor mediated regulation of ethylene biosynthesis, role of ethylene receptors in flower senescence, fruit ripening and other effects induced by ethylene. The expression behavior of the receptor and downstream molecules in climacteric and non climacteric crops is also elaborated upon. Possible strategies and recent advances in altering the ethylene sensitivity of plants using ethylene receptor genes in an attempt to modulate the regulation and sensitivity to ethylene have also been discussed. Not only will these transgenic plants be a boon to post-harvest physiology and crop improvement but, it will also help us in discovering the mechanism of regulation of ethylene sensitivity. PMID:22751331

Filamin 2 (Fln2)

PubMed Central

Thompson, Terri G.; Chan, Yiu-Mo; Hack, Andrew A.; Brosius, Melissa; Rajala, Michael; Lidov, Hart G.W.; McNally, Elizabeth M.; Watkins, Simon; Kunkel, Louis M.

2000-01-01

Mutations in genes encoding for the sarcoglycans, a subset of proteins within the dystrophin–glycoprotein complex, produce a limb-girdle muscular dystrophy phenotype; however, the precise role of this group of proteins in the skeletal muscle is not known. To understand the role of the sarcoglycan complex, we looked for sarcoglycan interacting proteins with the hope of finding novel members of the dystrophin–glycoprotein complex. Using the yeast two-hybrid method, we have identified a skeletal muscle-specific form of filamin, which we term filamin 2 (FLN2), as a γ- and δ-sarcoglycan interacting protein. In addition, we demonstrate that FLN2 protein localization in limb-girdle muscular dystrophy and Duchenne muscular dystrophy patients and mice is altered when compared with unaffected individuals. Previous studies of filamin family members have determined that these proteins are involved in actin reorganization and signal transduction cascades associated with cell migration, adhesion, differentiation, force transduction, and survival. Specifically, filamin proteins have been found essential in maintaining membrane integrity during force application. The finding that FLN2 interacts with the sarcoglycans introduces new implications for the pathogenesis of muscular dystrophy. PMID:10629222

Chemistry and biology of the initial steps in vision: the Friedenwald lecture.

PubMed

Palczewski, Krzysztof

2014-10-22

Visual transduction is the process in the eye whereby absorption of light in the retina is translated into electrical signals that ultimately reach the brain. The first challenge presented by visual transduction is to understand its molecular basis. We know that maintenance of vision is a continuous process requiring the activation and subsequent restoration of a vitamin A-derived chromophore through a series of chemical reactions catalyzed by enzymes in the retina and retinal pigment epithelium (RPE). Diverse biochemical approaches that identified key proteins and reactions were essential to achieve a mechanistic understanding of these visual processes. The three-dimensional arrangements of these enzymes' polypeptide chains provide invaluable insights into their mechanisms of action. A wealth of information has already been obtained by solving high-resolution crystal structures of both rhodopsin and the retinoid isomerase from pigment RPE (RPE65). Rhodopsin, which is activated by photoisomerization of its 11-cis-retinylidene chromophore, is a prototypical member of a large family of membrane-bound proteins called G protein-coupled receptors (GPCRs). RPE65 is a retinoid isomerase critical for regeneration of the chromophore. Electron microscopy (EM) and atomic force microscopy have provided insights into how certain proteins are assembled to form much larger structures such as rod photoreceptor cell outer segment membranes. A second challenge of visual transduction is to use this knowledge to devise therapeutic approaches that can prevent or reverse conditions leading to blindness. Imaging modalities like optical coherence tomography (OCT) and scanning laser ophthalmoscopy (SLO) applied to appropriate animal models as well as human retinal imaging have been employed to characterize blinding diseases, monitor their progression, and evaluate the success of therapeutic agents. Lately two-photon (2-PO) imaging, together with biochemical assays, are revealing functional aspects of vision at a new molecular level. These multidisciplinary approaches combined with suitable animal models and inbred mutant species can be especially helpful in translating provocative cell and tissue culture findings into therapeutic options for further development in animals and eventually in humans. A host of different approaches and techniques is required for substantial progress in understanding fundamental properties of the visual system. Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.

Influence of Additional Leading-Edge Surface Roughness on Performances in Highly Loaded Compressor Cascade

NASA Astrophysics Data System (ADS)

Chen, Shaowen; Xu, Hao; Sun, Shijun; Zhang, Longxin; Wang, Songtao

2015-05-01

Experimental research has been carried out at low speed to investigate the effect of additional leading-edge surface roughness on a highly-loaded axial compressor cascade. A 5-hole aerodynamic probe has been traversed across one pitch to obtain the distribution of total pressure loss coefficient, secondary flow vector, flow angles and other aerodynamic parameters at the exit section. Meanwhile, ink-trace flow visualization has been used to measure the flow fields on the walls of cascades and a detailed topology structure of the flow on the walls has been obtained. Aerodynamic parameters and flow characteristics are compared by arranging different levels of roughness on various parts of the leading edge. The results show that adding surface roughness at the leading edge and on the suction side obviously influences cascade performance. Aggravated 3-D flow separation significantly increases the loss in cascades, and the loss increases till 60% when the level of emery paper is 80 mm. Even there is the potential to improve cascade performance in local area of cascade passage. The influence of the length of surface roughness on cascade performance is not always adverse, and which depends on the position of surface roughness.

Visualization of deep ultraviolet photons based on Förster resonance energy transfer and cascade photon reabsorption in diphenylalanine-carbon nitrides composite film

NASA Astrophysics Data System (ADS)

Gan, Zhixing; Zhou, Weiping; Chen, Zhihui; Wang, Huan; Di, Yunsong; Huang, Shisong

2016-11-01

A diphenylalanine (L-Phe-L-Phe, FF)-carbon nitride composite film is designed and fabricated to visualize the deep ultraviolet (DUV, 245-290 nm) photons. The FF film, composed of diphenylalanine molecules, doped with carbon nitrides shows blue emission under excitation of DUV light, which makes the DUV beam observable. Both Förster resonance energy transfer and cascade photon reabsorption contribute to the conversion of photon energy. First, the FF is excited by the DUV photons. On one hand, the energy transfers to the embedded carbon nitrides through nonradiative dipole-dipole couplings. On the other hand, the 284 nm photons emitted from the FF would further excite the carbon nitrides, which will finally convert to blue fluorescence. Herein, the experimental demonstration of a simple device for the visualization of high DUV fluxes is reported.

A cascade model of information processing and encoding for retinal prosthesis.

PubMed

Pei, Zhi-Jun; Gao, Guan-Xin; Hao, Bo; Qiao, Qing-Li; Ai, Hui-Jian

2016-04-01

Retinal prosthesis offers a potential treatment for individuals suffering from photoreceptor degeneration diseases. Establishing biological retinal models and simulating how the biological retina convert incoming light signal into spike trains that can be properly decoded by the brain is a key issue. Some retinal models have been presented, ranking from structural models inspired by the layered architecture to functional models originated from a set of specific physiological phenomena. However, Most of these focus on stimulus image compression, edge detection and reconstruction, but do not generate spike trains corresponding to visual image. In this study, based on state-of-the-art retinal physiological mechanism, including effective visual information extraction, static nonlinear rectification of biological systems and neurons Poisson coding, a cascade model of the retina including the out plexiform layer for information processing and the inner plexiform layer for information encoding was brought forward, which integrates both anatomic connections and functional computations of retina. Using MATLAB software, spike trains corresponding to stimulus image were numerically computed by four steps: linear spatiotemporal filtering, static nonlinear rectification, radial sampling and then Poisson spike generation. The simulated results suggested that such a cascade model could recreate visual information processing and encoding functionalities of the retina, which is helpful in developing artificial retina for the retinally blind.

Receptor signaling clusters in the immune synapse(in eng)

DOE PAGES

Dustin, Michael L.; Groves, Jay T.

2012-02-23

Signaling processes between various immune cells involve large-scale spatial reorganization of receptors and signaling molecules within the cell-cell junction. These structures, now collectively referred to as immune synapses, interleave physical and mechanical processes with the cascades of chemical reactions that constitute signal transduction systems. Molecular level clustering, spatial exclusion, and long-range directed transport are all emerging as key regulatory mechanisms. The study of these processes is drawing researchers from physical sciences to join the effort and represents a rapidly growing branch of biophysical chemistry. Furthermore, recent advances in physical and quantitative analyses of signaling within the immune synapses are reviewedmore » here.« less

[Dysregulation of the mTOR signaling pathway in the pathogenesis of autism spectrum disorders].

PubMed

Gabryel, Bożena; Kapałka, Agata; Sobczyk, Wojciech; Łabuzek, Krzysztof; Gawęda, Agnieszka; Janas-Kozik, Małgorzata

2014-04-10

Mammalian target of rapamycin (mTor) plays multiple role in central nervous system and is involved in regulation of cell viability, differentiation, transcription, translation, protein degradation, actin cytoskeletal organization and autophagy. Recent experimental and clinical studies reveal that disturbances of mTOR signaling are involved in the pathogenesis of autism spectrum disorders (ASD). This article reviews current data on the alteration in the mTOR transduction cascade, which may contribute to common neurobehavioral disorders typical for ASD. Moreover, the results of the latest experimental studies on the potential of mTOR inhibitors for the treatment of ASD are reviewed.

Receptor signaling clusters in the immune synapse (in eng)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dustin, Michael L.; Groves, Jay T.

2012-02-23

Signaling processes between various immune cells involve large-scale spatial reorganization of receptors and signaling molecules within the cell-cell junction. These structures, now collectively referred to as immune synapses, interleave physical and mechanical processes with the cascades of chemical reactions that constitute signal transduction systems. Molecular level clustering, spatial exclusion, and long-range directed transport are all emerging as key regulatory mechanisms. The study of these processes is drawing researchers from physical sciences to join the effort and represents a rapidly growing branch of biophysical chemistry. Furthermore, recent advances in physical and quantitative analyses of signaling within the immune synapses are reviewedmore » here.« less

Diverse Hormone Response Networks in 41 Independent Drosophila Cell Lines

DOE PAGES

Stoiber, Marcus; Celniker, Susan; Cherbas, Lucy; ...

2016-01-15

Steroid hormones induce cascades of gene activation and repression with transformative effects on cell fate . Steroid transduction plays a major role in the development and physiology of nearly all metazoan species, and in the progression of the most common forms of cancer. Despite the paramount importance of steroids in developmental and translational biology, a complete map of transcriptional response has not been developed for any hormone . In the case of 20-hydroxyecdysone (ecdysone) in Drosophila melanogaster, these trajectories range from apoptosis to immortalization. We mapped the ecdysone transduction network in a cohort of 41 cell lines, the largest suchmore » atlas yet assembled. We found that the early transcriptional response mirrors the distinctiveness of physiological origins: genes respond in restricted patterns, conditional on the expression levels of dozens of transcription factors. Only a small cohort of genes is constitutively modulated independent of initial cell state. Ecdysone-responsive genes tend to organize into directional same-stranded units, with consecutive genes induced from the same strand. Here, we identify half of the ecdysone receptor heterodimer as the primary rate-limiting step in the response, and find that initial receptor isoform levels modulate the activated cohort of target transcription factors. In conclusion, this atlas of steroid response reveals organizing principles of gene regulation by a model type II nuclear receptor and lays the foundation for comprehensive and predictive understanding of the ecdysone transduction network in the fruit fly.« less

Nasal chemosensory cells use bitter taste signaling to detect irritants and bacterial signals.

PubMed

Tizzano, Marco; Gulbransen, Brian D; Vandenbeuch, Aurelie; Clapp, Tod R; Herman, Jake P; Sibhatu, Hiruy M; Churchill, Mair E A; Silver, Wayne L; Kinnamon, Sue C; Finger, Thomas E

2010-02-16

The upper respiratory tract is continually assaulted with harmful dusts and xenobiotics carried on the incoming airstream. Detection of such irritants by the trigeminal nerve evokes protective reflexes, including sneezing, apnea, and local neurogenic inflammation of the mucosa. Although free intra-epithelial nerve endings can detect certain lipophilic irritants (e.g., mints, ammonia), the epithelium also houses a population of trigeminally innervated solitary chemosensory cells (SCCs) that express T2R bitter taste receptors along with their downstream signaling components. These SCCs have been postulated to enhance the chemoresponsive capabilities of the trigeminal irritant-detection system. Here we show that transduction by the intranasal solitary chemosensory cells is necessary to evoke trigeminally mediated reflex reactions to some irritants including acyl-homoserine lactone bacterial quorum-sensing molecules, which activate the downstream signaling effectors associated with bitter taste transduction. Isolated nasal chemosensory cells respond to the classic bitter ligand denatonium as well as to the bacterial signals by increasing intracellular Ca(2+). Furthermore, these same substances evoke changes in respiration indicative of trigeminal activation. Genetic ablation of either G alpha-gustducin or TrpM5, essential elements of the T2R transduction cascade, eliminates the trigeminal response. Because acyl-homoserine lactones serve as quorum-sensing molecules for gram-negative pathogenic bacteria, detection of these substances by airway chemoreceptors offers a means by which the airway epithelium may trigger an epithelial inflammatory response before the bacteria reach population densities capable of forming destructive biofilms.

Nasal chemosensory cells use bitter taste signaling to detect irritants and bacterial signals

PubMed Central

Tizzano, Marco; Gulbransen, Brian D.; Vandenbeuch, Aurelie; Clapp, Tod R.; Herman, Jake P.; Sibhatu, Hiruy M.; Churchill, Mair E. A.; Silver, Wayne L.; Kinnamon, Sue C.; Finger, Thomas E.

2010-01-01

The upper respiratory tract is continually assaulted with harmful dusts and xenobiotics carried on the incoming airstream. Detection of such irritants by the trigeminal nerve evokes protective reflexes, including sneezing, apnea, and local neurogenic inflammation of the mucosa. Although free intra-epithelial nerve endings can detect certain lipophilic irritants (e.g., mints, ammonia), the epithelium also houses a population of trigeminally innervated solitary chemosensory cells (SCCs) that express T2R bitter taste receptors along with their downstream signaling components. These SCCs have been postulated to enhance the chemoresponsive capabilities of the trigeminal irritant-detection system. Here we show that transduction by the intranasal solitary chemosensory cells is necessary to evoke trigeminally mediated reflex reactions to some irritants including acyl–homoserine lactone bacterial quorum-sensing molecules, which activate the downstream signaling effectors associated with bitter taste transduction. Isolated nasal chemosensory cells respond to the classic bitter ligand denatonium as well as to the bacterial signals by increasing intracellular Ca2+. Furthermore, these same substances evoke changes in respiration indicative of trigeminal activation. Genetic ablation of either Gα-gustducin or TrpM5, essential elements of the T2R transduction cascade, eliminates the trigeminal response. Because acyl–homoserine lactones serve as quorum-sensing molecules for Gram-negative pathogenic bacteria, detection of these substances by airway chemoreceptors offers a means by which the airway epithelium may trigger an epithelial inflammatory response before the bacteria reach population densities capable of forming destructive biofilms. PMID:20133764

MassCascade: Visual Programming for LC-MS Data Processing in Metabolomics.

PubMed

Beisken, Stephan; Earll, Mark; Portwood, David; Seymour, Mark; Steinbeck, Christoph

2014-04-01

Liquid chromatography coupled to mass spectrometry (LC-MS) is commonly applied to investigate the small molecule complement of organisms. Several software tools are typically joined in custom pipelines to semi-automatically process and analyse the resulting data. General workflow environments like the Konstanz Information Miner (KNIME) offer the potential of an all-in-one solution to process LC-MS data by allowing easy integration of different tools and scripts. We describe MassCascade and its workflow plug-in for processing LC-MS data. The Java library integrates frequently used algorithms in a modular fashion, thus enabling it to serve as back-end for graphical front-ends. The functions available in MassCascade have been encapsulated in a plug-in for the workflow environment KNIME, allowing combined use with e.g. statistical workflow nodes from other providers and making the tool intuitive to use without knowledge of programming. The design of the software guarantees a high level of modularity where processing functions can be quickly replaced or concatenated. MassCascade is an open-source library for LC-MS data processing in metabolomics. It embraces the concept of visual programming through its KNIME plug-in, simplifying the process of building complex workflows. The library was validated using open data.

Signal Transduction Pathways through TRK‐A and TRK‐B Receptors in Human Neuroblastoma Cells

PubMed Central

Kuroda, Hiroshi; Horii, Yoshihiro; Moritake, Hiroshi; Tanaka, Takeo; Hattori, Seisuke

2001-01-01

Little is known about the signal transduction pathways of TRK family receptors in neuroblastoma (NB) cells. In this study, an NB cell line, designated MP‐N‐TS, was established from an adrenal tumor taken from a 2‐year‐old boy. This cell line expressed both TRK‐A and TRK‐B receptors, which is rare in a single NB cell line. Therefore, the MP‐N‐TS cell line was used to determine whether the signal transduction through these constitutive receptors is functional. Three neurotrophins, nerve growth factor (NGF), brain‐derived neurotrophic factor (BDNF) and neurotrophin‐4/ 5 (NT‐4/5), induced tyrosine phosphorylation of panTRK, and BDNF and NT‐4/5 induced tyrosine phosphorylation of TRK‐B. Tyrosine phosphorylation of panTRK and/or TRK‐B by the neurotro‐phins was inhibited in the presence of a tyrosine kinase inhibitor K252a. Tyrosine phosphorylation of Src homologous and collagen (She), extracellular signal‐regulated kinase (ERK)‐l and ERK‐2, and phospholipase C‐γl (PLC‐γl) was increased by the three neurotrophins and the increase was inhibited in the presence of K252a. Activation of Ras, detected as the GTP‐bound form of Ras, was induced by the three neurotrophins. The neurotrophins did not modulate the expressions of TRK‐A or TRK‐B mRNA, but they did induce the expression of c‐fos mRNA. Exogenous NGF induced weak neurite outgrowth, whereas exogenous BDNF and NT‐4/5 induced distinct neurite outgrowth. Exogenous BDNF and NT‐4/5 increased the number of viable cells, while NGF did not. Our results demonstrate that the signal transduction pathways through TRK‐A and TRK‐B in MP‐N‐TS cells are functional and similar, and the main downstream signaling pathways from the three neurotrophins are mitogen‐activated protein kinase (MAPK) cascades through She, activated Ras, ERK‐1 and ERK‐2, and the transduction pathway through PLC‐γl. Further, BDNF and NT‐4/5 increased cell viability. The MP‐N‐TS cell line should be useful for clarifying the TRK‐A and TRK‐B signaling pathways responsible for the different prognoses in patients with NB. PMID:11223544

Cascading reaction of arginase and urease on a graphene-based FET for ultrasensitive, real-time detection of arginine.

PubMed

Berninger, Teresa; Bliem, Christina; Piccinini, Esteban; Azzaroni, Omar; Knoll, Wolfgang

2018-09-15

Herein, a biosensor based on a reduced graphene oxide field effect transistor (rGO-FET) functionalized with the cascading enzymes arginase and urease was developed for the detection of L-arginine. Arginase and urease were immobilized on the rGO-FET sensing surface via electrostatic layer-by-layer assembly using polyethylenimine (PEI) as cationic building block. The signal transduction mechanism is based on the ability of the cascading enzymes to selectively perform chemical transformations and prompt local pH changes, that are sensitively detected by the rGO-FET. In the presence of L-arginine, the transistors modified with (PEI/urease(arginase)) multilayers showed a shift in the Dirac point due to the change in the local pH close to the graphene surface, produced by the catalyzed urea hydrolysis. The transistors were able to monitor L-arginine in the 10-1000 μM linear range with a LOD of 10 μM, displaying a fast response and a good long-term stability. The sensor showed stereospecificity and high selectivity in the presence of non-target amino acids. Taking into account the label-free, real-time measurement capabilities and the easily quantifiable, electronic output signal, this biosensor offers advantages over state-of-the-art L-arginine detection methods. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

RNA-Seq Analysis Reveals MAPKKK Family Members Related to Drought Tolerance in Maize

PubMed Central

Ren, Wen; Yang, Fengling; He, Hang; Zhao, Jiuran

2015-01-01

The mitogen-activated protein kinase (MAPK) cascade is an evolutionarily conserved signal transduction pathway that is involved in plant development and stress responses. As the first component of this phosphorelay cascade, mitogen-activated protein kinase kinase kinases (MAPKKKs) act as adaptors linking upstream signaling steps to the core MAPK cascade to promote the appropriate cellular responses; however, the functions of MAPKKKs in maize are unclear. Here, we identified 71 MAPKKK genes, of which 14 were novel, based on a computational analysis of the maize (Zea mays L.) genome. Using an RNA-seq analysis in the leaf, stem and root of maize under well-watered and drought-stress conditions, we identified 5,866 differentially expressed genes (DEGs), including 8 MAPKKK genes responsive to drought stress. Many of the DEGs were enriched in processes such as drought stress, abiotic stimulus, oxidation-reduction, and metabolic processes. The other way round, DEGs involved in processes such as oxidation, photosynthesis, and starch, proline, ethylene, and salicylic acid metabolism were clearly co-expressed with the MAPKKK genes. Furthermore, a quantitative real-time PCR (qRT-PCR) analysis was performed to assess the relative expression levels of MAPKKKs. Correlation analysis revealed that there was a significant correlation between expression levels of two MAPKKKs and relative biomass responsive to drought in 8 inbred lines. Our results indicate that MAPKKKs may have important regulatory functions in drought tolerance in maize. PMID:26599013

Minor isotope safeguards techniques (MIST): Analysis and visualization of gas centrifuge enrichment plant process data using the MSTAR model

NASA Astrophysics Data System (ADS)

Shephard, Adam M.; Thomas, Benjamin R.; Coble, Jamie B.; Wood, Houston G.

2018-05-01

This paper presents a development related to the use of minor isotope safeguards techniques (MIST) and the MSTAR cascade model as it relates to the application of international nuclear safeguards at gas centrifuge enrichment plants (GCEPs). The product of this paper is a derivation of the universal and dimensionless MSTAR cascade model. The new model can be used to calculate the minor uranium isotope concentrations in GCEP product and tails streams or to analyze, visualize, and interpret GCEP process data as part of MIST. Applications of the new model include the detection of undeclared feed and withdrawal streams at GCEPs when used in conjunction with UF6 sampling and/or other isotopic measurement techniques.

Prostaglandin signalling regulates ciliogenesis by modulating intraflagellar transport.

PubMed

Jin, Daqing; Ni, Terri T; Sun, Jianjian; Wan, Haiyan; Amack, Jeffrey D; Yu, Guangju; Fleming, Jonathan; Chiang, Chin; Li, Wenyan; Papierniak, Anna; Cheepala, Satish; Conseil, Gwenaëlle; Cole, Susan P C; Zhou, Bin; Drummond, Iain A; Schuetz, John D; Malicki, Jarema; Zhong, Tao P

2014-09-01

Cilia are microtubule-based organelles that mediate signal transduction in a variety of tissues. Despite their importance, the signalling cascades that regulate cilium formation remain incompletely understood. Here we report that prostaglandin signalling affects ciliogenesis by regulating anterograde intraflagellar transport (IFT). Zebrafish leakytail (lkt) mutants show ciliogenesis defects, and the lkt locus encodes an ATP-binding cassette transporter (ABCC4). We show that Lkt/ABCC4 localizes to the cell membrane and exports prostaglandin E2 (PGE2), a function that is abrogated by the Lkt/ABCC4(T804M) mutant. PGE2 synthesis enzyme cyclooxygenase-1 and its receptor, EP4, which localizes to the cilium and activates the cyclic-AMP-mediated signalling cascade, are required for cilium formation and elongation. Importantly, PGE2 signalling increases anterograde but not retrograde velocity of IFT and promotes ciliogenesis in mammalian cells. These findings lead us to propose that Lkt/ABCC4-mediated PGE2 signalling acts through a ciliary G-protein-coupled receptor, EP4, to upregulate cAMP synthesis and increase anterograde IFT, thereby promoting ciliogenesis.

Multi-Compartmentalisation in the MAPK Signalling Pathway Contributes to the Emergence of Oscillatory Behaviour and to Ultrasensitivity

PubMed Central

Shuaib, Aban; Hartwell, Adam; Kiss-Toth, Endre; Holcombe, Mike

2016-01-01

Signal transduction through the Mitogen Activated Protein Kinase (MAPK) pathways is evolutionarily highly conserved. Many cells use these pathways to interpret changes to their environment and respond accordingly. The pathways are central to triggering diverse cellular responses such as survival, apoptosis, differentiation and proliferation. Though the interactions between the different MAPK pathways are complex, nevertheless, they maintain a high level of fidelity and specificity to the original signal. There are numerous theories explaining how fidelity and specificity arise within this complex context; spatio-temporal regulation of the pathways and feedback loops are thought to be very important. This paper presents an agent based computational model addressing multi-compartmentalisation and how this influences the dynamics of MAPK cascade activation. The model suggests that multi-compartmentalisation coupled with periodic MAPK kinase (MAPKK) activation may be critical factors for the emergence of oscillation and ultrasensitivity in the system. Finally, the model also establishes a link between the spatial arrangements of the cascade components and temporal activation mechanisms, and how both contribute to fidelity and specificity of MAPK mediated signalling. PMID:27243235

Succinate receptor GPR91 provides a direct link between high glucose levels and renin release in murine and rabbit kidney

PubMed Central

Toma, Ildikó; Kang, Jung Julie; Sipos, Arnold; Vargas, Sarah; Bansal, Eric; Hanner, Fiona; Meer, Elliott; Peti-Peterdi, János

2008-01-01

Diabetes mellitus is the most common and rapidly growing cause of end-stage renal disease in developed countries. A classic hallmark of early diabetes mellitus includes activation of the renin-angiotensin system (RAS), which may lead to hypertension and renal tissue injury, but the mechanism of RAS activation is elusive. Here we identified a paracrine signaling pathway in the kidney in which high levels of glucose directly triggered the release of the prohypertensive hormone renin. The signaling cascade involved the local accumulation of succinate and activation of the kidney-specific G protein–coupled metabolic receptor, GPR91, in the glomerular endothelium as observed in rat, mouse, and rabbit kidney sections. Elements of signal transduction included endothelial Ca2+, the production of NO and prostaglandin (PGE2), and their paracrine actions on adjacent renin-producing cells. This GPR91 signaling cascade may serve to modulate kidney function and help remove metabolic waste products through renal hyperfiltration, and it could also link metabolic diseases, such as diabetes, or metabolic syndrome with RAS overactivation, systemic hypertension, and organ injury. PMID:18535668

Protein kinase inhibitors in the treatment of inflammatory and autoimmune diseases

PubMed Central

Patterson, H; Nibbs, R; McInnes, I; Siebert, S

2014-01-01

Protein kinases mediate protein phosphorylation, which is a fundamental component of cell signalling, with crucial roles in most signal transduction cascades: from controlling cell growth and proliferation to the initiation and regulation of immunological responses. Aberrant kinase activity is implicated in an increasing number of diseases, with more than 400 human diseases now linked either directly or indirectly to protein kinases. Protein kinases are therefore regarded as highly important drug targets, and are the subject of intensive research activity. The success of small molecule kinase inhibitors in the treatment of cancer, coupled with a greater understanding of inflammatory signalling cascades, has led to kinase inhibitors taking centre stage in the pursuit for new anti-inflammatory agents for the treatment of immune-mediated diseases. Herein we discuss the main classes of kinase inhibitors; namely Janus kinase (JAK), mitogen-activated protein kinase (MAPK) and spleen tyrosine kinase (Syk) inhibitors. We provide a mechanistic insight into how these inhibitors interfere with kinase signalling pathways and discuss the clinical successes and failures in the implementation of kinase-directed therapeutics in the context of inflammatory and autoimmune disorders. PMID:24313320

Identification of MAPK Substrates Using Quantitative Phosphoproteomics.

PubMed

Zhang, Tong; Schneider, Jacqueline D; Zhu, Ning; Chen, Sixue

2017-01-01

Activation of mitogen-activated protein kinases (MAPKs) under diverse biotic and abiotic factors and identification of an array of downstream MAPK target proteins are hot topics in plant signal transduction. Through interactions with a plethora of substrate proteins, MAPK cascades regulate many physiological processes in the course of plant growth, development, and response to environmental factors. Identification and quantification of potential MAPK substrates are essential, but have been technically challenging. With the recent advancement in phosphoproteomics, here we describe a method that couples metal dioxide for phosphopeptide enrichment with tandem mass tags (TMT) mass spectrometry (MS) for large-scale MAPK substrate identification and quantification. We have applied this method to a transient expression system carrying a wild type (WT) and a constitutive active (CA) version of a MAPK. This combination of genetically engineered MAPKs and phosphoproteomics provides a high-throughput, unbiased analysis of MAPK-triggered phosphorylation changes on the proteome scale. Therefore, it is a robust method for identifying potential MAPK substrates and should be applicable in the study of other kinase cascades in plants as well as in other organisms.

Mitogen-activated protein kinase phosphatase-1: a critical phosphatase manipulating mitogen-activated protein kinase signaling in cardiovascular disease (review).

PubMed

Li, Chang-Yi; Yang, Ling-Chao; Guo, Kai; Wang, Yue-Peng; Li, Yi-Gang

2015-04-01

Mitogen-activated protein kinase (MAPK) cascades are important players in the overall representation of cellular signal transduction pathways, and the deregulation of MAPKs is involved in a variety of diseases. The activation of MAPK signals occurs through phosphorylation by MAPK kinases at conserved threonine and tyrosine (Thr-Xaa-Tyr) residues. The mitogen-activated protein kinase phosphatases (MKPs) are a major part of the dual-specificity family of phosphatases and specifically inactivate MAPKs by dephosphorylating both phosphotyrosine and phosphoserine/phosphothreonine residues within the one substrate. MAPKs binding to MKPs can enhance MKP stability and activity, providing an important negative-feedback control mechanism that limits the MAPK cascades. In recent years, accumulating and compelling evidence from studies mainly employing cultured cells and mouse models has suggested that the archetypal MKP family member, MKP-1, plays a pivotal role in cardiovascular disease as a major negative modulator of MAPK signaling pathways. In the present review, we summarize the current knowledge on the pathological properties and the regulation of MKP-1 in cardiovascular disease, which may provide valuable therapeutic options.

A visual water vapor photonic crystal sensor with PVA/SiO2 opal structure

NASA Astrophysics Data System (ADS)

Yang, Haowei; Pan, Lei; Han, Yingping; Ma, Lihua; Li, Yao; Xu, Hongbo; Zhao, Jiupeng

2017-11-01

In study, we proposed a simple yet fast optical sensing motif based on thimbleful of polyvinyl alcohol (PVA) infiltrated photonic crystal (PC), which allows for high efficiency in vapor sensing through changes in their inter-layer space. Linear response to a broad dynamic range of vapor concentration was realized. Ultrafast response time (<1 s) and excellent recyclability were also demonstrated. Selective response to a vapor was exhibited, reflecting well the characteristic sorption properties of PVA, with which colorimetric reporting was readily achieved. These substantial improvements in performance are attributed to the efficacy of signal transduction and the enhanced signal transduction because of thimbleful PVA infiltrated space between adjacent SiO2 nanospheres.

Protein phosphatase 2ACα gene knock-out results in cortical atrophy through activating hippo cascade in neuronal progenitor cells.

PubMed

Liu, Bo; Sun, Li-Hua; Huang, Yan-Fei; Guo, Li-Jun; Luo, Li-Shu

2018-02-01

Protein phosphatase 2ACα (PP2ACα), a vital member of the protein phosphatase family, has been studied primarily as a regulator for the development, growth and protein synthesis of a lot of cell types. Dysfunction of PP2ACα protein results in neurodegenerative disease; however, this finding has not been directly confirmed in the mouse model with PP2ACα gene knock-out. Therefore, in this study presented here, we generated the PP2ACα gene knock-out mouse model by the Cre-loxP targeting gene system, with the purpose to directly observe the regulatory role of PP2ACα gene in the development of mouse's cerebral cortex. We observe that knocking-out PP2ACα gene in the central nervous system (CNS) results in cortical neuronal shrinkage, synaptic plasticity impairments, and learning/memory deficits. Further study reveals that PP2ACα gene knock-out initiates Hippo cascade in cortical neuroprogenitor cells (NPCs), which blocks YAP translocation into the nuclei of NPCs. Notably, p73, directly targeted by Hippo cascade, can bind to the promoter of glutaminase2 (GLS2) that plays a dominant role in the enzymatic regulation of glutamate/glutamine cycle. Finally, we find that PP2ACα gene knock-out inhibits the glutamine synthesis through up-regulating the activity of phosphorylated-p73 in cortical NPCs. Taken together, it concludes that PP2ACα critically supports cortical neuronal growth and cognitive function via regulating the signaling transduction of Hippo-p73 cascade. And PP2ACα indirectly modulates the glutamine synthesis of cortical NPCs through targeting p73 that plays a direct transcriptional regulatory role in the gene expression of GLS2. Copyright © 2017 Elsevier Ltd. All rights reserved.

RO 90-7501 Enhances TLR3 and RLR Agonist Induced Antiviral Response

PubMed Central

Guo, Fang; Mead, Jennifer; Aliya, Nishat; Wang, Lijuan; Cuconati, Andrea; Wei, Lai; Li, Kui; Block, Timothy M.; Guo, Ju-Tao; Chang, Jinhong

2012-01-01

Recognition of virus infection by innate pattern recognition receptors (PRRs), including membrane-associated toll-like receptors (TLR) and cytoplasmic RIG-I-like receptors (RLR), activates cascades of signal transduction pathways leading to production of type I interferons (IFN) and proinflammatory cytokines that orchestrate the elimination of the viruses. Although it has been demonstrated that PRR-mediated innate immunity plays an essential role in defending virus from infection, it also occasionally results in overwhelming production of proinflammatory cytokines that cause severe inflammation, blood vessel leakage and tissue damage. In our efforts to identify small molecules that selectively enhance PRR-mediated antiviral, but not the detrimental inflammatory response, we discovered a compound, RO 90–7501 (‘2’-(4-Aminophenyl)-[2,5′-bi-1H-benzimidazol]-5-amine), that significantly promoted both TLR3 and RLR ligand-induced IFN-β gene expression and antiviral response, most likely via selective activation of p38 mitogen-activated protein kinase (MAPK) pathway. Our results thus imply that pharmacological modulation of PRR signal transduction pathways in favor of the induction of a beneficial antiviral response can be a novel therapeutic strategy. PMID:23056170

Activation of RIG-I-like Receptor Signal Transduction

PubMed Central

Bruns, Annie; Horvath, Curt M.

2011-01-01

Mammalian cells have the ability to recognize virus infection and mount a powerful antiviral response. Pattern recognition receptor proteins detect molecular signatures of virus infection and activate antiviral signaling cascades. The RIG-I-like receptors are cytoplasmic DExD/H box proteins that can specifically recognize virus-derived RNA species as a molecular feature discriminating the pathogen from the host. The RIG-I-like receptor family is composed of three homologous proteins, RIG-I, MDA5, and LGP2. All of these proteins can bind double-stranded RNA species with varying affinities via their conserved DExD/H box RNA helicase domains and C-terminal regulatory domains. The recognition of foreign RNA by the RLRs activates enzymatic functions and initiates signal transduction pathways resulting in the production of antiviral cytokines and the establishment of a broadly effective cellular antiviral state that protects neighboring cells from infection and triggers innate and adaptive immune systems. The propagation of this signal via the interferon antiviral system has been studied extensively, while the precise roles for enzymatic activities of the RNA helicase domain in antiviral responses are only beginning to be elucidated. Here, current models for RLR ligand recognition and signaling are reviewed. PMID:22066529

Activation of the Jnk signaling pathway by a dual-specificity phosphatase, JSP-1.

PubMed

Shen, Y; Luche, R; Wei, B; Gordon, M L; Diltz, C D; Tonks, N K

2001-11-20

The mitogen-activated protein kinases (MAPKs) are integral to the mechanisms by which cells respond to physiological stimuli, such as growth factors, hormones, and cytokines, and to a wide variety of environmental stresses. The MAPKs, which are stimulated by phosphorylation of a TXY motif in their activation loop, are components of signal transduction cascades in which sequential activation of protein kinases culminates in their activation and their subsequent phosphorylation of various effector proteins that mediate the physiological response. MAPKs are also subject to dephosphorylation and inactivation, both by enzymes that recognize the residues of the TXY motif independently and by dual specificity phosphatases, which dephosphroylate both Tyr and Ser/Thr residues. We report the identification and characterization of a novel dual specificity phosphatase. Contrary to expectation, this broadly expressed enzyme did not inactivate MAPKs in transient cotransfection assays but instead displayed the capacity to function as a selective activator of the MAPK Jnk, hence the name, Jnk Stimulatory Phosphatase-1 (JSP-1). This study illustrates a new aspect of the regulation of MAPK-dependent signal transduction and raises the possibility that JSP-1 may offer a different perspective to the study of various inflammatory and proliferative disorders associated with dysfunctional Jnk signaling.

A MAP kinase gene, Clk1, is required for conidiation and pathogenicity in the phytopathogenic fungus Curvularia lunata.

PubMed

Gao, Shi Gang; Zhou, Fei Hong; Liu, Tong; Li, Ying Ying; Chen, Jie

2013-03-01

Mitogen-activated protein kinase (MAPK) cascades are highly conserved signal transduction pathways, which play a wide variety of important roles in extracellular signal transduction. The first MAPK gene of the maize pathogen Curvularia lunata, Clk1, was isolated via a PCR-based approach with a primer pair designed on the basis of conserved regions of known MAPKs. Southern blot analysis showed that the gene existed in the genome as a single copy. The predicted amino acid sequence (352 amino acids) was highly homologous with MAP kinases of other phytopathogenic fungi. Flanking regions of Clk1 were obtained through RACE and genomic walking technology. To understand the role of Clk1 in C. lunata, targeted gene disruption was adopted to construct Clk1 mutants. It was found that mutants lacking functional domain of Clk1 were not able to produce conidia but tended to form a few special chlamydospore-shaped structures. Clk1 mutants grew slower in adverse environments (at 24°C), produced less cell degrading enzymes (CWDEs) than the wild type, and they were almost unable to infect maize leaves via artificial wounds. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Regional expression patterns of taste receptors and gustducin in the mouse tongue.

PubMed

Kim, Mi-Ryung; Kusakabe, Yuko; Miura, Hirohito; Shindo, Yoichiro; Ninomiya, Yuzo; Hino, Akihiro

2003-12-12

In order to understand differences in taste sensitivities of taste bud cells between the anterior and posterior part of tongue, it is important to analyze the regional expression patterns of genes related to taste signal transduction on the tongue. Here we examined the expression pattern of a taste receptor family, the T1r family, and gustducin in circumvallate and fungiform papillae of the mouse tongue using double-labeled in situ hybridization. Each member of the T1r family was expressed in both circumvallate and fungiform papillae with some differences in their expression patterns. The most striking difference between fungiform and circumvallate papillae was observed in their co-expression patterns of T1r2, T1r3, and gustducin. T1r2-positive cells in fungiform papillae co-expressed T1r3 and gustducin, whereas T1r2 and T1r3 double-positive cells in circumvallate papillae merely expressed gustducin. These results suggested that in fungiform papillae, gustducin might play a role in the sweet taste signal transduction cascade mediated by a sweet receptor based on the T1r2 and T1r3 combination, in fungiform papillae.

Interaction of Herbal Compounds with Biological Targets: A Case Study with Berberine

PubMed Central

Chen, Xiao-Wu; Di, Yuan Ming; Zhang, Jian; Zhou, Zhi-Wei; Li, Chun Guang; Zhou, Shu-Feng

2012-01-01

Berberine is one of the main alkaloids found in the Chinese herb Huang lian (Rhizoma Coptidis), which has been reported to have multiple pharmacological activities. This study aimed to analyze the molecular targets of berberine based on literature data followed by a pathway analysis using the PANTHER program. PANTHER analysis of berberine targets showed that the most classes of molecular functions include receptor binding, kinase activity, protein binding, transcription activity, DNA binding, and kinase regulator activity. Based on the biological process classification of in vitro berberine targets, those targets related to signal transduction, intracellular signalling cascade, cell surface receptor-linked signal transduction, cell motion, cell cycle control, immunity system process, and protein metabolic process are most frequently involved. In addition, berberine was found to interact with a mixture of biological pathways, such as Alzheimer's disease-presenilin and -secretase pathways, angiogenesis, apoptosis signalling pathway, FAS signalling pathway, Hungtington disease, inflammation mediated by chemokine and cytokine signalling pathways, interleukin signalling pathway, and p53 pathways. We also explored the possible mechanism of action for the anti-diabetic effect of berberine. Further studies are warranted to elucidate the mechanisms of action of berberine using systems biology approach. PMID:23213296

Carcinogenesis and Reactive Oxygen Species Signaling: Interaction of the NADPH Oxidase NOX1-5 and Superoxide Dismutase 1-3 Signal Transduction Pathways.

PubMed

Parascandolo, Alessia; Laukkanen, Mikko O

2018-04-05

Reduction/oxidation (redox) balance could be defined as an even distribution of reduction and oxidation complementary processes and their reaction end products. There is a consensus that aberrant levels of reactive oxygen species (ROS), commonly observed in cancer, stimulate primary cell immortalization and progression of carcinogenesis. However, the mechanism how different ROS regulate redox balance is not completely understood. Recent Advances: In the current review, we have summarized the main signaling cascades inducing NADPH oxidase NOX1-5 and superoxide dismutase (SOD) 1-3 expression and their connection to cell proliferation, immortalization, transformation, and CD34 + cell differentiation in thyroid, colon, lung, breast, and hematological cancers. Interestingly, many of the signaling pathways activating redox enzymes or mediating the effect of ROS are common, such as pathways initiated from G protein-coupled receptors and tyrosine kinase receptors involving protein kinase A, phospholipase C, calcium, and small GTPase signaling molecules. The clarification of interaction of signal transduction pathways could explain how cells regulate redox balance and may even provide means to inhibit the accumulation of harmful levels of ROS in human pathologies. Antioxid. Redox Signal. 00, 000-000.

Antiviral Defense Mechanisms in Honey Bees

PubMed Central

Brutscher, Laura M.; Daughenbaugh, Katie F.; Flenniken, Michelle L.

2015-01-01

Honey bees are significant pollinators of agricultural crops and other important plant species. High annual losses of honey bee colonies in North America and in some parts of Europe have profound ecological and economic implications. Colony losses have been attributed to multiple factors including RNA viruses, thus understanding bee antiviral defense mechanisms may result in the development of strategies that mitigate colony losses. Honey bee antiviral defense mechanisms include RNA-interference, pathogen-associated molecular pattern (PAMP) triggered signal transduction cascades, and reactive oxygen species generation. However, the relative importance of these and other pathways is largely uncharacterized. Herein we review the current understanding of honey bee antiviral defense mechanisms and suggest important avenues for future investigation. PMID:26273564

Reversible cysteine oxidation in hydrogen peroxide sensing and signal transduction.

PubMed

García-Santamarina, Sarela; Boronat, Susanna; Hidalgo, Elena

2014-04-29

Activation of redox cascades through hydrogen peroxide-mediated reversible cysteine oxidation is a major mechanism for intracellular signaling. Understanding why some cysteine residues are specifically oxidized, in competition with other proximal cysteine residues and in the presence of strong redox buffers, is therefore crucial for understanding redox signaling. In this review, we explore the recent advances in thiol-redox chemistry linked to signaling. We describe the last findings in the field of redox sensors, those that are naturally present in different model organisms as well as those that have been engineered to quantify intracellular hydrogen peroxide concentrations. Finally, we provide a summary of the newest approaches developed to study reversible cysteine oxidation at the proteomic level.

Empirical Comparison of Visualization Tools for Larger-Scale Network Analysis

DOE PAGES

Pavlopoulos, Georgios A.; Paez-Espino, David; Kyrpides, Nikos C.; ...

2017-07-18

Gene expression, signal transduction, protein/chemical interactions, biomedical literature cooccurrences, and other concepts are often captured in biological network representations where nodes represent a certain bioentity and edges the connections between them. While many tools to manipulate, visualize, and interactively explore such networks already exist, only few of them can scale up and follow today’s indisputable information growth. In this review, we shortly list a catalog of available network visualization tools and, from a user-experience point of view, we identify four candidate tools suitable for larger-scale network analysis, visualization, and exploration. Lastly, we comment on their strengths and their weaknesses andmore » empirically discuss their scalability, user friendliness, and postvisualization capabilities.« less

Empirical Comparison of Visualization Tools for Larger-Scale Network Analysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pavlopoulos, Georgios A.; Paez-Espino, David; Kyrpides, Nikos C.

Gene expression, signal transduction, protein/chemical interactions, biomedical literature cooccurrences, and other concepts are often captured in biological network representations where nodes represent a certain bioentity and edges the connections between them. While many tools to manipulate, visualize, and interactively explore such networks already exist, only few of them can scale up and follow today’s indisputable information growth. In this review, we shortly list a catalog of available network visualization tools and, from a user-experience point of view, we identify four candidate tools suitable for larger-scale network analysis, visualization, and exploration. Lastly, we comment on their strengths and their weaknesses andmore » empirically discuss their scalability, user friendliness, and postvisualization capabilities.« less

Signal transduction profile of chemical sensitisers in dendritic cells: An endpoint to be included in a cell-based in vitro alternative approach to hazard identification?

DOE Office of Scientific and Technical Information (OSTI.GOV)

Neves, Bruno Miguel; Centro de Neurociencias e Biologia Celular, Universidade de Coimbra, Coimbra 3004-517; Goncalo, Margarida

2011-01-15

The development of non-animal testing methods for the assessment of skin sensitisation potential is an urgent challenge within the framework of existing and forthcoming legislation. Efforts have been made to replace current animal tests, but so far no alternative methods have been developed. It is widely recognised that alternatives to animal testing cannot be accomplished with a single approach, but rather will require the integration of results obtained from different in vitro and in silico assays. The argument subjacent to the development of in vitro dendritic cell (DC)-based assays is that sensitiser-induced changes in the DC phenotype can be differentiatedmore » from those induced by irritants. This assumption is derived from the unique capacity of DC to convert environmental signals encountered at the skin into a receptor expression pattern (MHC class II molecules, co-stimulatory molecules, chemokine receptors) and a soluble mediator release profile that will stimulate T lymphocytes. Since signal transduction cascades precede changes in surface marker expression and cytokine/chemokine secretion, these phenotypic modifications are a consequence of a signal transduction profile that is specifically triggered by sensitisers and not by irritants. A limited number of studies have addressed this subject and the present review attempts to summarise and highlight all of the signalling pathways modulated by skin sensitisers and irritants. Furthermore, we conclude this review by focusing on the most promising strategies suitable for inclusion into a cell-based in vitro alternative approach to hazard identification.« less

Is rhodopsin isomerization correlated to astronauts' phosphene perceptions in space?

NASA Astrophysics Data System (ADS)

Narici, L.; Altea-Biophys Team

Anomalous Phosphene Perception APP phoenomenon may just be a first example of how microgravity and particle radiation may modify the normal behaviour of the Central Nervous System CNS and also is an evidence that space environment may indeed influence the correct functioning of the visual system The ALTEA program is going to provide i an assesment of the CNS functional hazard due to microgravity and particle radiation during long space human permanence ii a definition for the needed shielding optimized for reducing these risks and iii a survey of ISS radiation environment aimed at the validation of spacecrafts computer models As known Rhodopsin is at the start of the photo-transduction cascade and its involvement in the phosphene perception would suggest a possible physiological pathway The bleaching of few molecules in the retina is sufficient to start the process of vision A very preliminary measurements on rhodopsin irradiation has been conducted in April 2003 Irradiation of 17 vials containing a solution of suine rhodopsine has been performed with 12 C ions at 200 MeV n - total dose has been varied from 10 9 to 10 13 ions over each vial New and more complete data from most recent measurements are now available Preparation and purification of bovine rhodopsin and regenerations of bleached molecules was carried out using reproducible procedures The samples was irradiated with controlled 12 C ion beams and with different amount of light radiation in order 1 to understand if the molecules have been

Cone arrestin confers cone vision of high temporal resolution in zebrafish larvae.

PubMed

Renninger, Sabine L; Gesemann, Matthias; Neuhauss, Stephan C F

2011-02-01

Vision of high temporal resolution depends on careful regulation of photoresponse kinetics, beginning with the lifetime of activated photopigment. The activity of rhodopsin is quenched by high-affinity binding of arrestin to photoexcited phosphorylated photopigment, which effectively terminates the visual transduction cascade. This regulation mechanism is well established for rod photoreceptors, yet its role for cone vision is still controversial. In this study we therefore analyzed arrestin function in the cone-dominated vision of larval zebrafish. For both rod (arrS ) and cone (arr3 ) arrestin we isolated two paralogs, each expressed in the respective subset of photoreceptors. Labeling with paralog-specific antibodies revealed subfunctionalized expression of Arr3a in M- and L-cones, and Arr3b in S- and UV-cones. The inactivation of arr3a by morpholino knockdown technology resulted in a severe delay in photoresponse recovery which, under bright light conditions, was rate-limiting. Comparison to opsin phosphorylation-deficient animals confirmed the role of cone arrestin in late cone response recovery. Arr3a activity partially overlapped with the function of the cone-specific kinase Grk7a involved in initial response recovery. Behavioral measurements further revealed Arr3a deficiency to be sufficient to reduce temporal contrast sensitivity, providing evidence for the importance of arrestin in cone vision of high temporal resolution. © 2011 The Authors. European Journal of Neuroscience © 2011 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.

Universal fieldable assay with unassisted visual detection

NASA Technical Reports Server (NTRS)

Chelyapov, Nicolas (Inventor)

2012-01-01

A universal detection system based on allosteric aptamers, signal amplification cascade, and eye-detectable phrase transition. A broadly applicable homogeneous detection system is provided. It utilizes components of the blood coagulation cascade in the presence of polystyrene microspheres (MS) as a signal amplifier. Russell's viper venom factor X activator (RVV-X) triggers the cascade, which results in an eye-visible phase transition--precipitation of MS bound to clotted fibrin. An allosteric RNA aptamer, RNA132, with affinity for RVV-X and human vascular endothelial growth factor (VEGF.sub.165) was created. RNA132 inhibits enzymatic activity of RVV-X. The effector molecule, VEGF.sub.165, reverses the inhibitory activity of RNA132 on RVV-X and restores its enzymatic activity, thus triggering the cascade and enabling the phase transition. Similar results were obtained for another allosteric aptamer modulated by a protein tyrosine phosphatase. The assay is instrumentation-free for both processing and readout.

Glucose oxidase-initiated cascade catalysis for sensitive impedimetric aptasensor based on metal-organic frameworks functionalized with Pt nanoparticles and hemin/G-quadruplex as mimicking peroxidases.

PubMed

Zhou, Xingxing; Guo, Shijing; Gao, Jiaxi; Zhao, Jianmin; Xue, Shuyan; Xu, Wenju

2017-12-15

Based on cascade catalysis amplification driven by glucose oxidase (GOx), a sensitive electrochemical impedimetric aptasensor for protein (carcinoembryonic antigen, CEA as tested model) was proposed by using Cu-based metal-organic frameworks functionalized with Pt nanoparticles, aptamer, hemin and GOx (Pt@CuMOFs-hGq-GOx). CEA aptamer loaded onto Pt@CuMOFs was bound with hemin to form hemin@G-quadruplex (hGq) with mimicking peroxidase activity. Through sandwich-type reaction of target CEA and CEA aptamers (Apt1 and Apt2), the obtained Pt@CuMOFs-hGq-GOx as signal transduction probes (STPs) was captured to the modified electrode interface. When 3,3-diaminobenzidine (DAB) and glucose were introduced, the cascade reaction was initiated by GOx to catalyze the oxidation of glucose, in situ generating H 2 O 2 . Simultaneously, the decomposition of the generated H 2 O 2 was greatly promoted by Pt@CuMOFs and hGq as synergistic peroxide catalysts, accompanying with the significant oxidation process of DAB and the formation of nonconductive insoluble precipitates (IPs). As a result, the electron transfer in the resultant sensing interface was effectively hindered and the electrochemical impedimetric signal (EIS) was efficiently amplified. Thus, the high sensitivity of the proposed CEA aptasensor was successfully improved with 0.023pgmL -1 , which may be promising and potential in assaying certain clinical disease related to CEA. Copyright © 2017 Elsevier B.V. All rights reserved.

A label-free colorimetric isothermal cascade amplification for the detection of disease-related nucleic acids based on double-hairpin molecular beacon.

PubMed

Wu, Dong; Xu, Huo; Shi, Haimei; Li, Weihong; Sun, Mengze; Wu, Zai-Sheng

2017-03-08

K-Ras mutations at codon 12 play an important role in an early step of carcinogenesis. Here, a label-free colorimetric isothermal cascade amplification for ultrasensitive and specific detection of K-Ras point mutation is developed based on a double-hairpin molecular beacon (DHMB). The biosensor consists of DHMB probe and a primer-incorporated polymerization template (PPT) designed partly complementary to DHMB. In the presence of polymerase, target DNA is designed to trigger strand displacement amplification (SDA) via promote the hybridization of PPT with DHMB and subsequently initiates cascade amplification process with the help of the nicking endonuclease. During the hybridization and enzymatic reaction, G-quadruplex/hemin DNAzymes are generated, catalyzing the oxidation of ABTS 2- by H 2 O 2 in the presence of hemin. Utilizing the proposed facile colorimetric scheme, the target DNA can be quantified down to 4 pM with the dynamic response range of 5 orders of magnitude, indicating the substantially improved detection capability. Even more strikingly, point mutation in K-ras gene can be readily observed by the naked eye without the need for the labeling or expensive equipment. Given the high-performance for K-Ras analysis, the enhanced signal transduction capability associated with double-hairpin structure of DHMB provides a novel rout to screen biomarkers, and the descripted colorimetric biosensor seems to hold great promise for diagnostic applications of genetic diseases. Copyright © 2016 Elsevier B.V. All rights reserved.

Mechanisms for type-II vitellogenesis-inhibiting hormone suppression of vitellogenin transcription in shrimp hepatopancreas: Crosstalk of GC/cGMP pathway with different MAPK-dependent cascades.

PubMed

Chen, Ting; Ren, Chunhua; Jiang, Xiao; Zhang, Lvping; Li, Hongmei; Huang, Wen; Hu, Chaoqun

2018-01-01

Vitellogenesis is the process of yolk formation via accumulating vitellin (Vn) with nutrients in the oocytes. Expression of vitellogenin (Vg), the precursor of Vn, is one of the indicators for the start of vitellogenesis. In Pacific white shrimp (Litopenaeus vannamei), the type-II vitellogenesis-inhibiting hormone (VIH-2) effectively suppresses hepatopancreatic Vg mRNA expression. In this study, we demonstrate the increasing transcript levels of hepatopancreatic Vg during L. vannamei ovarian development, suggesting that the hepatopancreas-derived Vg/Vn may also contribute to vitellogenesis in this species. Using a combination of in vivo injections and in vitro primary cell cultures, we provide evidences that the inhibition of VIH-2 on hepatopancreatic Vg gene expression is mediated through a functional coupling of the GC/cGMP pathway with different MAPK-dependent cascades in female shrimp. In VIH-2 signaling, the NO-independent GC/cGMP/PKG cascades were upstream of the MAPKs. Activations of the MAPK signal by VIH-2 include the phosphorylation of JNK and the mRNA/protein expression of P38MAPK. Additionally, the cAMP/PKA pathway is another positive intracellular signal for hepatopancreatic Vg mRNA expression but is independent of its VIH-2 regulation. Our findings establish a model for the signal transduction mechanism of Vg regulation by VIH and shed light on the biological functions and signaling of the CHH family in crustaceans.

Mechanisms for type-II vitellogenesis-inhibiting hormone suppression of vitellogenin transcription in shrimp hepatopancreas: Crosstalk of GC/cGMP pathway with different MAPK-dependent cascades

PubMed Central

Ren, Chunhua; Jiang, Xiao; Zhang, Lvping; Li, Hongmei; Huang, Wen; Hu, Chaoqun

2018-01-01

Vitellogenesis is the process of yolk formation via accumulating vitellin (Vn) with nutrients in the oocytes. Expression of vitellogenin (Vg), the precursor of Vn, is one of the indicators for the start of vitellogenesis. In Pacific white shrimp (Litopenaeus vannamei), the type-II vitellogenesis-inhibiting hormone (VIH-2) effectively suppresses hepatopancreatic Vg mRNA expression. In this study, we demonstrate the increasing transcript levels of hepatopancreatic Vg during L. vannamei ovarian development, suggesting that the hepatopancreas-derived Vg/Vn may also contribute to vitellogenesis in this species. Using a combination of in vivo injections and in vitro primary cell cultures, we provide evidences that the inhibition of VIH-2 on hepatopancreatic Vg gene expression is mediated through a functional coupling of the GC/cGMP pathway with different MAPK-dependent cascades in female shrimp. In VIH-2 signaling, the NO-independent GC/cGMP/PKG cascades were upstream of the MAPKs. Activations of the MAPK signal by VIH-2 include the phosphorylation of JNK and the mRNA/protein expression of P38MAPK. Additionally, the cAMP/PKA pathway is another positive intracellular signal for hepatopancreatic Vg mRNA expression but is independent of its VIH-2 regulation. Our findings establish a model for the signal transduction mechanism of Vg regulation by VIH and shed light on the biological functions and signaling of the CHH family in crustaceans. PMID:29590153

Nitric oxide in B6 mouse and nitric oxide-sensitive soluble guanylate cyclase in cat modulate acetylcholine release in pontine reticular formation.

PubMed

Lydic, Ralph; Garza-Grande, Ricardo; Struthers, Richard; Baghdoyan, Helen A

2006-05-01

ACh regulates arousal, and the present study was designed to provide insight into the neurochemical mechanisms modulating ACh release in the pontine reticular formation. Nitric oxide (NO)-releasing beads microinjected into the pontine reticular formation of C57BL/6J (B6) mice significantly (P < 0.0001) increased ACh release. Microdialysis delivery of the NO donor N-ethyl-2-(1-ethyl-2-hydroxy-2-nitrosohydrazino)-ethanamine (NOC-12) to the mouse pontine reticular formation also caused a concentration-dependent increase in ACh release (P < 0.001). These are the first neurochemical data showing that ACh release in the pontine reticular formation of the B6 mouse is modulated by NO. The signal transduction cascade through which NO modulates ACh release in the pontine reticular formation has not previously been characterized. Therefore, an additional series of studies quantified the effects of a soluble guanylate cyclase (sGC) inhibitor, 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ), on ACh release in the cat medial pontine reticular formation. During naturally occurring states of sleep and wakefulness, but not anesthesia, ODQ caused a significant (P < 0.001) decrease in ACh release. These results show for the first time that NO modulates ACh in the medial pontine reticular formation of the cat via an NO-sensitive sGC signal transduction cascade. Isoflurane and halothane anesthesia have been shown to decrease ACh release in the medial pontine reticular formation. The finding that ODQ did not alter ACh release during isoflurane or halothane anesthesia demonstrates that these anesthetics disrupt the NO-sensitive sGC-cGMP pathway. Considered together, results from the mouse and cat indicate that NO modulates ACh release in arousal-promoting regions of the pontine reticular formation via an NO-sensitive sGC-cGMP pathway.

Cuscuta reflexa invasion induces Ca release in its host.

PubMed

Albert, M; van der Krol, S; Kaldenhoff, R

2010-05-01

Cuscuta reflexa induces a variety of reaction in its hosts. Some of these are visual reactions, and it is clear that these morphological changes are preceded by events at the molecular level, where signal transduction is one of the early processes. Calcium (Ca(2+)) release is the major second messenger during signal transduction, and we therefore studied Ca(2+) spiking in tomato during infection with C. reflexa. Bioluminescence in aequorin-expressing tomato was monitored for 48 h after the onset of Cuscuta infestation. Signals at the attachment sites were observed from 30 to 48 h. Treatment of aequorin-expressing tomato leaf disks with Cuscuta plant extracts suggested that the substance that induced Ca(2+) release from the host was closely linked to parasite haustoria.

Flow visualization study of the horseshoe vortex in a turbine stator cascade

NASA Technical Reports Server (NTRS)

Gaugler, R. E.; Russell, L. M.

1982-01-01

Flow visualization techniques were used to show the behavior of the horseshoe vortex in a large scale turbine stator cascade. Oil drops on the end wall surface flowed in response to local shear stresses, indicating the limiting flow streamlines at the surface. Smoke injected into the flow and photographed showed time averaged flow behavior. Neutrally bouyant helium filled soap bubbles followed the flow and showed up on photographs as streaks, indicating the paths followed by individual fluid particles. Preliminary attempts to control the vortex were made by injecting air through control jets drilled in the end wall near the vane leading edge. Seventeen different hole locations were tested, one at a time, and the effect of the control jets on the path follwed by smoke in the boundary layer was recorded photographically.

Secondary Vortex Structures in Vortex Generator Induced Flow

NASA Astrophysics Data System (ADS)

Velte, Clara; Okulov, Valery; Hansen, Martin

2010-11-01

Passive rectangular vane actuators can induce a longitudinal vortex that redistributes the momentum in the boundary layer to control the flow. Recent experiments [1] as well as previous studies [2] have shown that a secondary vortex of opposite sign is generated along with the primary one, supposedly from local separation of the boundary layer due to the primary vortex. 2D flow visualizations of a vortex in the vicinity of a boundary support this hypothesis [3]. These secondary vortices are studied for various configurations -- single generator, counter- and co-rotating cascades. The objective is to study their removal through cancelation in cascades using Stereoscopic Particle Image Velocimetry and flow visualization.[4pt] [1] Velte, Hansen and Okulov, J. Fluid Mech. 619, 2009.[0pt] [2] Zhang, Int. J. Heat Fluid Flow 21 2000.[0pt] [3] Harris, Miller and Williamson, APS abstract 2009.

IDC1, a pezizomycotina-specific gene that belongs to the PaMpk1 MAP kinase transduction cascade of the filamentous fungus Podospora anserina.

PubMed

Jamet-Vierny, Corinne; Debuchy, Robert; Prigent, Magali; Silar, Philippe

2007-12-01

Components involved in the activation of the MAPK cascades in filamentous fungi are not well known. Here, we provide evidence that IDC1, a pezizomycotina-specific gene is involved along with the PaNox1 NADPH oxidase in the nuclear localization of the PaMpk1 MAP kinase, a prerequisite for MAPK activity. Mutants of IDC1 display the same phenotypes as mutants in PaNox1 and PaMpk1, i.e., lack of pigment and of aerial hyphae, female sterility, impairment in hyphal interference and inability to develop Crippled Growth cell degeneration. As observed for the PaNox1 mutant, IDC1 mutants are hypostatic to PaMpk1 mutants. IDC1 seems to play a key role in sexual reproduction. Indeed, fertility is diminished in strains with lower level of IDC1. In strains over-expressing IDC1, protoperithecia reach a later stage of development towards perithecia without fertilization; however, upon fertilization maturation of fertile perithecia is diminished and delayed. In addition, heterokaryon construction shows that IDC1 is necessary together with PaNox1 in the perithecial envelope but not in the dikaryon resulting from fertilization.

Introducing fluorescence resonance energy transfer-based biosensors for the analysis of cAMP-PKA signalling in the fungal pathogen Candida glabrata.

PubMed

Demuyser, Liesbeth; Van Genechten, Wouter; Mizuno, Hideaki; Colombo, Sonia; Van Dijck, Patrick

2018-05-29

The cyclic adenosine monophosphate-protein kinase A (cAMP-PKA) pathway is central to signal transduction in many organisms. In pathogenic fungi such as Candida albicans, this signalling cascade has proven to be involved in several processes, such as virulence, indicating its potential importance in antifungal drug discovery. Candida glabrata is an upcoming pathogen of the same species, yet information regarding the role of cAMP-PKA signalling in virulence is largely lacking. To enable efficient monitoring of cAMP-PKA activity in this pathogen, we here present the usage of two FRET-based biosensors. Both variations in the activity of PKA and the quantity of cAMP can be detected in a time-resolved manner, as we exemplify by glucose-induced activation of the pathway. We also present information on how to adequately process and analyse the data in a mathematically correct and physiologically relevant manner. These sensors will be of great benefit for scientists interested in linking the cAMP-PKA signalling cascade to downstream processes, such as virulence, possibly in a host environment. © 2018 John Wiley & Sons Ltd.

Plant cell surface receptor-mediated signaling - a common theme amid diversity.

PubMed

He, Yunxia; Zhou, Jinggeng; Shan, Libo; Meng, Xiangzong

2018-01-29

Sessile plants employ a diverse array of plasma membrane-bound receptors to perceive endogenous and exogenous signals for regulation of plant growth, development and immunity. These cell surface receptors include receptor-like kinases (RLKs) and receptor-like proteins (RLPs) that harbor different extracellular domains for perception of distinct ligands. Several RLK and RLP signaling pathways converge at the somatic embryogenesis receptor kinases (SERKs), which function as shared co-receptors. A repertoire of receptor-like cytoplasmic kinases (RLCKs) associate with the receptor complexes to relay intracellular signaling. Downstream of the receptor complexes, mitogen-activated protein kinase (MAPK) cascades are among the key signaling modules at which the signals converge, and these cascades regulate diverse cellular and physiological responses through phosphorylation of different downstream substrates. In this Review, we summarize the emerging common theme that underlies cell surface receptor-mediated signaling pathways in Arabidopsis thaliana : the dynamic association of RLKs and RLPs with specific co-receptors and RLCKs for signal transduction. We further discuss how signaling specificities are maintained through modules at which signals converge, with a focus on SERK-mediated receptor signaling. © 2018. Published by The Company of Biologists Ltd.

Organ-specific effects of brassinosteroids on stomatal production coordinate with the action of Too Many Mouths.

PubMed

Wang, Ming; Yang, Kezhen; Le, Jie

2015-03-01

In Arabidopsis, stomatal development initiates after protodermal cells acquire stomatal lineage cell fate. Stomata or their precursors communicate with their neighbor epidermal cells to ensure the "one cell spacing" rule. The signals from EPF/EPFL peptide ligands received by Too Many Mouths (TMM) and ERECTA-family receptors are supposed to be transduced by YODA MAPK cascade. A basic helix-loop-helix transcription factor SPEECHLESS (SPCH) is another key regulator of stomatal cell fate determination and asymmetric entry divisions, and SPCH activity is regulated by YODA MAPK cascade. Brassinosteroid (BR) signaling, one of the most well characterized signal transduction pathways in plants, contributes to the control of stomatal production. But opposite organ-specific effects of BR on stomatal production were reported. Here we confirm that stomatal production in hypocotyls is controlled by BR levels. YODA and CYCD4 are not essential for BR stomata-promoting function. Furthermore, we found that BR could confer tmm hypocotyls clustered stomatal phenotype, indicating that the BR organ-specific effects on stomatal production might coordinate with the TMM organ-specific actions. © 2014 Institute of Botany, Chinese Academy of Sciences.

Graviperception and gravitaxis in flagellates

NASA Astrophysics Data System (ADS)

Häder, D.-P.; Richter, P.; Ntefidou, M.; Lebert, M.

Unicellular flagellates perceive and react to the gravitational vector of the Earth. Previous hypotheses have suggested that the orientation is brought about by a passive physical mechanism such as buoyancy or hydrodynamic alignment. Recent results of experiments on parabolic rocket flights have revealed that in the photosynthetic Euglena only 10 % of the orientation can be explained by passive orientation while the remainder relies on an active physiological sensor and an internal sensory transduction chain. The cellular contents is heavier than the surrounding medium and consequently presses onto the lower membrane where it activates mechano-sensitive ion channels located at the front end under the trailing flagellum. These channels allow a gated influx of calcium (visualized by confocal microscopy) which depolarizes the internal electrical potential and eventually causes a course correction by the flagellar beating. Further elements in the transduction chain are cyclic AMP and related enzymes. Recent experiments during parabolic aircraft flights and on sounding rockets have confirmed this hypothesis and provided detailed insight into the biochemical sensory transduction chain. Currently the molecular mechanisms of graviperception are being studied.

Unphosphorylated STATs go nuclear.

PubMed

Brown, Stephen; Zeidler, Martin P

2008-10-01

The JAK/STAT signal transduction pathway has traditionally been viewed as a cytokine-stimulated activator of gene expression consisting of a straightforward receptor/JAK kinase/STAT transcription factor cascade. Recent studies in Drosophila, have, however consistently identified a range of chromatin-remodelling factors as regulators of in vivo JAK/STAT signalling. Now, the detailed analysis of one of these, heterochromatin protein 1 (HP1), has provided an insight into an unexpected non-canonical in vivo role for STAT. In this model, unphosphorylated STATs associate with and maintain the stability of transcriptionally repressed heterochromatin--an effect countered by the recruitment of STAT to the canonical pathway. We examine the background of this new model and its implications for JAK/STAT pathway requirements in stem cell maintenance and cancer.

Neutrophil apoptosis in the context of tuberculosis infection.

PubMed

Alemán, Mercedes

2015-07-01

Polymorphonuclear neutrophils comprise two-thirds of peripheral blood leukocytes and are key components of innate immunity as a first line of defense against bacterial and fungal pathogens. Their microbicidal mechanisms are essential for bacterial killing, the enhancement of inflammatory reactions and also comprise signaling molecules which have been implicated in signal transduction cascades. In tuberculosis, the number of neutrophils increases in the affected lung. In addition, they become activated and apoptotic due the bacterial burden. As apoptosis is promoted by reactive oxygen species (ROS) during phagocytosis, the advantages and benefits to the host as well as the strategies displayed by the pathogen to avoid or retard apoptosis are discussed in this review. Copyright © 2015 Elsevier Ltd. All rights reserved.

Cloning and characterization of a novel human STAR domain containing cDNA KHDRBS2.

PubMed

Wang, Liu; Xu, Jian; Zeng, Li; Ye, Xin; Wu, Qihan; Dai, Jianfeng; Ji, Chaoneng; Gu, Shaohua; Zhao, Chunhua; Xie, Yi; Mao, Yumin

2002-12-01

KHDRBS2, KH domain containing, RNA binding, signal transduction associated 2, is an RNA-binding protein that is tyrosine phosphorylated by Src during mitosis. It contains a KH domain,which is embedded in a larger conserved domain called the STAR domain. This protein has a 99% sequence identity with rat SLM-1 (the Sam68-like mammalian protein 1) and 98% sequence identity with mouse SLM-1 in its STAR domain. KHDRBS2 has the characteristic Sam68 SH2 and SH3 domain binding sites. RT-PCR analysis showed its transcript is ubiquitously expressed. The characterization of KHDRBS2 indicates it may link tyrosine kinase signaling cascades with some aspect of RNA metabolism.

Somatic mutations of GUCY2F, EPHA3, and NTRK3 in human cancers.

PubMed

Wood, Laura D; Calhoun, Eric S; Silliman, Natalie; Ptak, Janine; Szabo, Steve; Powell, Steve M; Riggins, Gregory J; Wang, Tian-Li; Yan, Hai; Gazdar, Adi; Kern, Scott E; Pennacchio, Len; Kinzler, Kenneth W; Vogelstein, Bert; Velculescu, Victor E

2006-10-01

Tyrosine kinases are major regulators of signal transduction cascades involved in cellular proliferation and have important roles in tumorigenesis. We have recently analyzed the tyrosine kinase gene family for alterations in human colorectal cancers and identified somatic mutations in seven members of this gene family. In this study we have used high-throughput sequencing approaches to further evaluate this subset of genes for genetic alterations in other human tumors. We identified somatic mutations in GUCY2F, EPHA3, and NTRK3 in breast, lung, and pancreatic cancers. Our results implicate these tyrosine kinase genes in the pathogenesis of other tumor types and suggest that they may be useful targets for diagnostic and therapeutic intervention in selected patients.

Towards a visual modeling approach to designing microelectromechanical system transducers

NASA Astrophysics Data System (ADS)

Dewey, Allen; Srinivasan, Vijay; Icoz, Evrim

1999-12-01

In this paper, we address initial design capture and system conceptualization of microelectromechanical system transducers based on visual modeling and design. Visual modeling frames the task of generating hardware description language (analog and digital) component models in a manner similar to the task of generating software programming language applications. A structured topological design strategy is employed, whereby microelectromechanical foundry cell libraries are utilized to facilitate the design process of exploring candidate cells (topologies), varying key aspects of the transduction for each topology, and determining which topology best satisfies design requirements. Coupled-energy microelectromechanical system characterizations at a circuit level of abstraction are presented that are based on branch constitutive relations and an overall system of simultaneous differential and algebraic equations. The resulting design methodology is called visual integrated-microelectromechanical VHDL-AMS interactive design (VHDL-AMS is visual hardware design language for analog and mixed signal).

Synchronized Schlieren method for vortex shedding in cascade during acoustic resonance

NASA Astrophysics Data System (ADS)

Nagashima, T.; Tanida, Y.

1986-10-01

An evaluation is made of synchronized schlieren optical system methods for the simultaneous visualization of both the acoustic wave and vortex shedding phenomena encountered during acoustic resonance excited by vortex shedding from the trailing edges of cascade blades. Attention is given to the case of parallel flat plate blades in throughflow velocities of up to 100 m/s. The acoustic wavefront is found to appear in the trailing edge region and travel upstream when a pair of vortices of opposite sign are fully developed at the trailing edge.

Signal transduction, plasma membrane calcium movements, and pigment translocation in freshwater shrimp chromatophores.

PubMed

Milograna, Sarah Ribeiro; Bell, Fernanda Tinti; McNamara, John Campbell

2010-11-01

Crustacean color change results from the differential translocation of chromatophore pigments, regulated by neurosecretory peptides like red pigment concentrating hormone (RPCH) that, in the red ovarian chromatophores of the freshwater shrimp Macrobrachium olfersi, triggers pigment aggregation via increased cytosolic cGMP and Ca(2+) of both smooth endoplasmatic reticulum (SER) and extracellular origin. However, Ca(2+) movements during RPCH signaling and the mechanisms that regulate intracellular [Ca(2+)] are enigmatic. We investigate Ca(2+) transporters in the chromatophore plasma membrane and Ca(2+) movements that occur during RPCH signal transduction. Inhibition of the plasma membrane Ca(2+)-ATPase by La(3+) and indirect inhibition of the Na(+)/Ca(2+) exchanger by ouabain induce pigment aggregation, revealing a role for both in Ca(2+) extrusion. Ca(2+) channel blockade by La(3+) or Cd(2+) strongly inhibits slow-phase RPCH-triggered aggregation during which pigments disperse spontaneously. L-type Ca(2+) channel blockade by gabapentin markedly reduces rapid-phase translocation velocity; N- or P/Q-type blockade by ω-conotoxin MVIIC strongly inhibits RPCH-triggered aggregation and reduces velocity, effects revealing RPCH-signaled influx of extracellular Ca(2+). Plasma membrane depolarization, induced by increasing external K(+) from 5 to 50 mM, produces Ca(2+)-dependent pigment aggregation, whereas removal of K(+) from the perfusate causes pigment hyperdispersion, disclosing a clear correlation between membrane depolarization and pigment aggregation; K(+) channel blockade by Ba(2+) also partially inhibits RPCH action. We suggest that, during RPCH signal transduction, Ca(2+) released from the SER, together with K(+) channel closure, causes chromatophore membrane depolarization, leading to the opening of predominantly N- and/or P/Q-type voltage-gated Ca(2+) channels, and a Ca(2+)/cGMP cascade, resulting in pigment aggregation.

Phosphorylation-mediated negative regulation of RIG-I antiviral activity.

PubMed

Gack, Michaela U; Nistal-Villán, Estanislao; Inn, Kyung-Soo; García-Sastre, Adolfo; Jung, Jae U

2010-04-01

Recognition of invading viruses by the host is elicited by cellular sensors which trigger signaling cascades that lead to type I interferon (IFN) gene expression. Retinoic acid-inducible gene I (RIG-I) has emerged as a key receptor for the detection of viral RNA in the cytosol, inducing IFN-mediated innate immune responses to limit viral replication through its interaction with MAVS (also called IPS-1, CARDIF, or VISA). Upon the recognition of viral RNA, the Lys-172 residue of RIG-I undergoes ubiquitination induced by tripartite motif protein 25 (TRIM25), an essential protein for antiviral signal transduction. Here we demonstrate that phosphorylation represents another regulatory mechanism for RIG-I-mediated antiviral activity. Using protein purification and mass spectrometry analysis, we identified three phosphorylation sites in the amino-terminal caspase recruitment domains (CARDs) of RIG-I. One of these residues, Thr-170, is located in close proximity to Lys-172, and we speculated that its phosphorylation may affect Lys-172 ubiquitination and functional activation of RIG-I. Indeed, a RIG-I mutant carrying a phosphomimetic Glu residue in place of Thr-170 loses TRIM25 binding, Lys-172 ubiquitination, MAVS binding, and downstream signaling ability. This suggests that phosphorylation of RIG-I at Thr-170 inhibits RIG-I-mediated antiviral signal transduction. Immunoblot analysis with a phospho-specific antibody showed that the phosphorylation of the RIG-I Thr-170 residue is present under normal conditions but rapidly declines upon viral infection. Our results indicate that Thr-170 phosphorylation and TRIM25-mediated Lys-172 ubiquitination of RIG-I functionally antagonize each other. While Thr-170 phosphorylation keeps RIG-I latent, Lys-172 ubiquitination enables RIG-I to form a stable complex with MAVS, thereby inducing IFN signal transduction.

Improving the Transduction of Bone Marrow–Derived Cells with an Integrase-Defective Lentiviral Vector

PubMed Central

Pay, S. Louise; Qi, Xiaoping; Willard, Jeffrey F.; Godoy, Juliana; Sankhavaram, Kavya; Horton, Ranier; Mitter, Sayak K.; Quigley, Judith L.; Chang, Lung-Ji; Grant, Maria B.; Boulton, Michael E.

2018-01-01

In lentiviral vector (LV) applications where transient transgene expression is sufficient, integrase-defective lentiviral vectors (IDLVs) are beneficial for reducing the potential for off-target effects associated with insertional mutagenesis. It was previously demonstrated that human RPE65 mRNA expression from an integrating lentiviral vector (ILV) induces endogenous Rpe65 and Cralbp mRNA expression in murine bone marrow–derived cells (BMDCs), initiating programming of the cells to retinal pigment epithelium (RPE)-like cells. These cells regenerate RPE in retinal degeneration models when injected systemically. As transient expression of RPE65 is sufficient to activate endogenous RPE-associated genes for programming BMDCs, use of an ILV is an unnecessary risk. In this study, an IDLV expressing RPE65 (IDLV3-RPE65) was generated. Transduction with IDLV3-RPE65 is less efficient than the integrating vector (ILV3-RPE65). Therefore, IDLV3-RPE65 transduction was enhanced with a combination of preloading 20 × -concentrated viral supernatant on RetroNectin at a multiplicity of infection of 50 and transduction of BMDCs by low-speed centrifugation. RPE65 mRNA levels increased from ∼12-fold to ∼25-fold (p < 0.05) after modification of the IDLV3-RPE65 transduction protocol, achieving expression similar to the ∼27-fold (p < 0.05) increase observed with ILV3-RPE65. Additionally, the study shows that the same preparation of RetroNectin can be used to coat up to three wells with no reduction in transduction. Critically, IDLV3-RPE65 transduction initiates endogenous Rpe65 mRNA expression in murine BMDCs and Cralbp/CRALBP mRNA in both murine and human BMDCs, similar to expression observed in ILV3-RPE65-transduced cells. Systemic administration of ILV3-RPE65 or IDLV3-RPE65 programmed BMDCs in a mouse model of retinal degeneration is sufficient to retain visual function and reduce retinal degeneration compared to mice receiving no treatment or naïve BMDC. It is concluded that IDLV3-RPE65 is appropriate for programming BMDCs to RPE-like cells. PMID:29160102

Activation of the Jnk signaling pathway by a dual-specificity phosphatase, JSP-1

PubMed Central

Shen, Yu; Luche, Ralf; Wei, Bo; Gordon, Marcia L.; Diltz, Curtis D.; Tonks, Nicholas K.

2001-01-01

The mitogen-activated protein kinases (MAPKs) are integral to the mechanisms by which cells respond to physiological stimuli, such as growth factors, hormones, and cytokines, and to a wide variety of environmental stresses. The MAPKs, which are stimulated by phosphorylation of a TXY motif in their activation loop, are components of signal transduction cascades in which sequential activation of protein kinases culminates in their activation and their subsequent phosphorylation of various effector proteins that mediate the physiological response. MAPKs are also subject to dephosphorylation and inactivation, both by enzymes that recognize the residues of the TXY motif independently and by dual specificity phosphatases, which dephosphroylate both Tyr and Ser/Thr residues. We report the identification and characterization of a novel dual specificity phosphatase. Contrary to expectation, this broadly expressed enzyme did not inactivate MAPKs in transient cotransfection assays but instead displayed the capacity to function as a selective activator of the MAPK Jnk, hence the name, Jnk Stimulatory Phosphatase-1 (JSP-1). This study illustrates a new aspect of the regulation of MAPK-dependent signal transduction and raises the possibility that JSP-1 may offer a different perspective to the study of various inflammatory and proliferative disorders associated with dysfunctional Jnk signaling. PMID:11717427

Molecular Cloning and Characterization of G Alpha Proteins from the Western Tarnished Plant Bug, Lygus hesperus

PubMed Central

Hull, J. Joe; Wang, Meixian

2014-01-01

The Gα subunits of heterotrimeric G proteins play critical roles in the activation of diverse signal transduction cascades. However, the role of these genes in chemosensation remains to be fully elucidated. To initiate a comprehensive survey of signal transduction genes, we used homology-based cloning methods and transcriptome data mining to identity Gα subunits in the western tarnished plant bug (Lygus hesperus Knight). Among the nine sequences identified were single variants of the Gαi, Gαo, Gαs, and Gα12 subfamilies and five alternative splice variants of the Gαq subfamily. Sequence alignment and phylogenetic analyses of the putative L. hesperus Gα subunits support initial classifications and are consistent with established evolutionary relationships. End-point PCR-based profiling of the transcripts indicated head specific expression for LhGαq4, and largely ubiquitous expression, albeit at varying levels, for the other LhGα transcripts. All subfamilies were amplified from L. hesperus chemosensory tissues, suggesting potential roles in olfaction and/or gustation. Immunohistochemical staining of cultured insect cells transiently expressing recombinant His-tagged LhGαi, LhGαs, and LhGαq1 revealed plasma membrane targeting, suggesting the respective sequences encode functional G protein subunits. PMID:26463065

Mechanotransduction molecules in the plant gravisensory response: amyloplast/statolith membranes contain a beta 1 integrin-like protein

NASA Technical Reports Server (NTRS)

Lynch, T. M.; Lintilhac, P. M.; Domozych, D.

1998-01-01

It has been hypothesized that the sedimentation of amyloplasts within root cap cells is the primary event in the plant gravisensory-signal transduction cascade. Statolith sedimentation, with its ability to generate weighty mechanical signals, is a legitimate means for organisms to discriminate the direction of the gravity vector. However, it has been demonstrated that starchless mutants with reduced statolith densities maintain some ability to sense gravity, calling into question the statolith sedimentation hypothesis. Here we report on the presence of a beta 1 integrin-like protein localized inside amyloplasts of tobacco NT-1 suspension culture, callus cells, and whole-root caps. Two different antibodies to the beta 1 integrin, one to the cytoplasmic domain and one to the extracellular domain, localize in the vicinity of the starch grains within amyloplasts of NT-1. Biochemical data reveals a 110-kDa protein immunoprecipitated from membrane fractions of NT-1 suspension culture indicating size homology to known beta 1 integrin in animals. This study provides the first direct evidence for the possibility of integrin-mediated signal transduction in the perception of gravity by higher plants. An integrin-mediated pathway, initiated by starch grain sedimentation within the amyloplast, may provide the signal amplification necessary to explain the gravitropic response in starch-depleted cultivars.

The molecular circuitry of brassinosteroid signaling.

PubMed

Belkhadir, Youssef; Jaillais, Yvon

2015-04-01

Because they are tethered in space, plants have to make the most of their local growth environment. In order to grow in an ever-changing environment, plants constantly remodel their shapes. This adaptive attribute requires the orchestration of complex environmental signals at the cellular and organismal levels. A battery of small molecules, classically known as phytohormones, allows plants to change their body plan by using highly integrated signaling networks and transcriptional cascades. Amongst these hormones, brassinosteroids (BRs), the polyhydroxylated steroid of plants, influence plant responsiveness to the local environment and exquisitely promote, or interfere with, many aspects of plant development. The molecular circuits that wire steroid signals at the cell surface to the promoters of thousands of genes in the nucleus have been defined in the past decade. This review recapitulates how the transduction of BR signals impacts the temporally unfolding programs of plant growth. First, we summarize the paradigmatic BR signaling pathway acting primarily in cellular expansion. Secondly, we describe the current wiring diagram and the temporal dynamics of the BR signal transduction network. And finally we provide an overview of how key players in BR signaling act as molecular gates to transduce BR signals onto other signaling pathways. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kirfel, Jutta; Pantelis, Dimitrios; Kabba, Mustapha

Four and one half LIM domain protein FHL2 participates in many cellular processes involved in tissue repair such as regulation of gene expression, cytoarchitecture, cell adhesion, migration and signal transduction. The repair process after wounding is initiated by the release of peptides and bioactive lipids. These molecules induce synthesis and deposition of a provisional extracellular matrix. We showed previously that sphingosine-1-phosphate (S1P) triggers a signal transduction cascade mediating nuclear translocation of FHL2 in response to activation of the RhoA GTPase. Our present study shows that FHL2 is an important signal transducer influencing the outcome of intestinal anastomotic healing. Early woundmore » healing is accompanied by reconstitution and remodelling of the extracellular matrix and collagen is primarily responsible for wound strength. Our results show that impaired intestinal wound healing in Fhl2-deficient mice is due to disturbed collagen III metabolism. Impaired collagen III synthesis reduced the mechanical stability of the anastomoses and led to lower bursting pressure in Fhl2-deficient mice after surgery. Our data confirm that FHL2 is an important factor regulating collagen expression in the early phase of wound healing, and thereby is critically involved in the physiologic process of anastomosis healing after bowel surgery and thus may represent a new therapeutic target.« less

Two rice GRAS family genes responsive to N -acetylchitooligosaccharide elicitor are induced by phytoactive gibberellins: evidence for cross-talk between elicitor and gibberellin signaling in rice cells.

PubMed

Day, R Bradley; Tanabe, Shigeru; Koshioka, Masaji; Mitsui, Toshiaki; Itoh, Hironori; Ueguchi-Tanaka, Miyako; Matsuoka, Makoto; Kaku, Hanae; Shibuya, Naoto; Minami, Eiichi

2004-01-01

In this study, we present data showing that two members of the GRAS family of genes from rice, CIGR1 and CIGR2 (chitin-inducible gibberellin-responsive), inducible by the potent elicitor N -acetylchitooligosaccharide (GN), are rapidly induced by exogenous gibberellins. The pattern of mRNA accumulation was dependent on the dose and biological activity of the gibberellins, suggesting that the induction of the genes by gibberellin is mediated by a biological receptor capable of specific recognition and signal transduction upon perception of the phytoactive compounds. Further pharmacological analysis revealed that the CIGR1 and CIGR2 mRNA accumulation by treatment with gibberellin is dependent upon protein phosphorylation/dephosphorylation events. In rice calli derived from slender rice 1, a constitutive gibberellin-responsive mutant, or d1, a mutant deficient in the alpha -subunit of the heterotrimeric G-protein, CIGR1 and CIGR2 were induced by a GN elicitor, yet not by gibberellin. Neither gibberellin nor GN showed related activities in defense or development, respectively. These results strongly suggested that the signal transduction cascade from gibberellin is independent of that from GN, and further implied that CIGR1 and CIGR2 have dual, distinct roles in defense and development.

The transcription factor FoxB mediates temporal loss of cellular competence for notochord induction in ascidian embryos.

PubMed

Hashimoto, Hidehiko; Enomoto, Takashi; Enomoto, Atsushi; Kumano, Gaku; Nishida, Hiroki

2011-06-01

In embryos of the ascidian Halocynthia roretzi, the competence of isolated presumptive notochord blastomeres to respond to fibroblast growth factor (FGF) for induction of the primary notochord decays by 1 hour after cleavage from the 32- to 64-cell stage. This study analyzes the molecular mechanisms responsible for this loss of competence and provides evidence for a novel mechanism. A forkhead family transcription factor, FoxB, plays a role in competence decay by preventing the induction of notochord-specific Brachyury (Bra) gene expression by the FGF/MAPK signaling pathway. Unlike the mechanisms reported previously in other animals, no component in the FGF signal transduction cascade appeared to be lost or inactivated at the time of competence loss. Knockdown of FoxB functions allowed the isolated cells to retain their competence for a longer period, and to respond to FGF with expression of Bra beyond the stage at which competence was normally lost. FoxB acts as a transcription repressor by directly binding to the cis-regulatory element of the Bra gene. Our results suggest that FoxB prevents ectopic induction of the notochord fate within the cells that assume a default nerve cord fate, after the stage when notochord induction has been completed. The merit of this system is that embryos can use the same FGF signaling cascade again for another purpose in the same cell lineage at later stages by keeping the signaling cascade itself available. Temporally and spatially regulated FoxB expression in nerve cord cells was promoted by the ZicN transcription factor and absence of FGF/MAPK signaling.

The crustacean eye: dark/light adaptation, polarization sensitivity, flicker fusion frequency, and photoreceptor damage.

PubMed

Meyer-Rochow, V B

2001-12-01

Compound eyes, nauplius eyes, frontal organs, intracerebral ocelli, and caudal photoreceptors are the main light and darkness detectors in crustaceans, but they need not be present all at once in an individual and in some crustaceans no photoreceptors whatsoever are known. Compound eye designs reflect on their functions and have evolved to allow the eye to operate optimally under a variety of environmental conditions. Dark-light-adaptational changes manifest themselves in pigment granule translocations, cell movements, and optical adjustments which fine-tune an eye's performance to rapid and unpredictable fluctuations in ambient light intensities as well as to the slower and predictable light level changes associated with day and night oscillations. Recycling of photoreceptive membrane and light-induced membrane collapse are superficially similar events that involve the transduction cascade, intracellular calcium, and membrane fatty acid composition, but which differ in aetiology and longterm consequence. Responses to intermittant illumination and linearly polarized light evoke in the eye of many crustaceans characteristic responses that appear to be attuned to each species' special needs. How the visual responses are processed more centrally and to what extent a crustacean makes behavioural use of e-vector discrimination and flickering lights are questions, however, that still have not been satisfactorily answered for the vast majority of all crustacean species. The degree of light-induced photoreceptor damage depends on a large number of variables, but once manifest, it tends to be progressive and irreversible. Concomittant temperature stress aggravates the situation and there is evidence that free radicals and lipid hydroperoxides are involved.

Direct interaction of the inhibitory gamma-subunit of Rod cGMP phosphodiesterase (PDE6) with the PDE6 GAFa domains.

PubMed

Muradov, Khakim G; Granovsky, Alexey E; Schey, Kevin L; Artemyev, Nikolai O

2002-03-26

Retinal rod and cone cGMP phosphodiesterases (PDE6 family) function as the effector enzyme in the vertebrate visual transduction cascade. The activity of PDE6 catalytic subunits is controlled by the Pgamma-subunits. In addition to the inhibition of cGMP hydrolysis at the catalytic sites, Pgamma is known to stimulate a noncatalytic binding of cGMP to the regulatory GAFa-GAFb domains of PDE6. The latter role of Pgamma has been attributed to its polycationic region. To elucidate the structural basis for the regulation of cGMP binding to the GAF domains of PDE6, a photoexcitable peptide probe corresponding to the polycationic region of Pgamma, Pgamma-21-45, was specifically cross-linked to rod PDE6alphabeta. The site of Pgamma-21-45 cross-linking was localized to Met138Gly139 within the PDE6alpha GAFa domain using mass spectrometric analysis. Chimeras between PDE5 and cone PDE6alpha', containing GAFa and/or GAFb domains of PDE6alpha' have been generated to probe a potential role of the GAFb domains in binding to Pgamma. Analysis of the inhibition of the PDE5/PDE6alpha' chimeras by Pgamma supported the role of PDE6 GAFa but not GAFb domains in the interaction with Pgamma. Our results suggest that a direct binding of the polycationic region of Pgamma to the GAFa domains of PDE6 may lead to a stabilization of the noncatalytic cGMP-binding sites.

Beam-modulation methods in quantitative and flow visualization holographic interferometry

NASA Technical Reports Server (NTRS)

Decker, A.

1986-01-01

This report discusses heterodyne holographic interferometry and time-average holography with a frequency shifted reference beam. Both methods will be used for the measurement and visualization of internal transonic flows, where the target facility is a flutter cascade. The background and experimental requirements for both methods are reviewed. Measurements using heterodyne holographic interferometry are presented. The performance of the laser required for time-average holography of time-varying transonic flows is discussed.

Beam-modulation methods in quantitative and flow-visualization holographic interferometry

NASA Technical Reports Server (NTRS)

Decker, Arthur J.

1986-01-01

Heterodyne holographic interferometry and time-average holography with a frequency shifted reference beam are discussed. Both methods will be used for the measurement and visualization of internal transonic flows where the target facility is a flutter cascade. The background and experimental requirements for both methods are reviewed. Measurements using heterodyne holographic interferometry are presented. The performance of the laser required for time-average holography of time-varying transonic flows is discussed.

Short-wavelength sensitive opsin (SWS1) as a new marker for vertebrate phylogenetics

PubMed Central

van Hazel, Ilke; Santini, Francesco; Müller, Johannes; Chang, Belinda SW

2006-01-01

Background Vertebrate SWS1 visual pigments mediate visual transduction in response to light at short wavelengths. Due to their importance in vision, SWS1 genes have been isolated from a surprisingly wide range of vertebrates, including lampreys, teleosts, amphibians, reptiles, birds, and mammals. The SWS1 genes exhibit many of the characteristics of genes typically targeted for phylogenetic analyses. This study investigates both the utility of SWS1 as a marker for inferring vertebrate phylogenetic relationships, and the characteristics of the gene that contribute to its phylogenetic utility. Results Phylogenetic analyses of vertebrate SWS1 genes produced topologies that were remarkably congruent with generally accepted hypotheses of vertebrate evolution at both higher and lower taxonomic levels. The few exceptions were generally associated with areas of poor taxonomic sampling, or relationships that have been difficult to resolve using other molecular markers. The SWS1 data set was characterized by a substantial amount of among-site rate variation, and a relatively unskewed substitution rate matrix, even when the data were partitioned into different codon sites and individual taxonomic groups. Although there were nucleotide biases in some groups at third positions, these biases were not convergent across different taxonomic groups. Conclusion Our results suggest that SWS1 may be a good marker for vertebrate phylogenetics due to the variable yet consistent patterns of sequence evolution exhibited across fairly wide taxonomic groups. This may result from constraints imposed by the functional role of SWS1 pigments in visual transduction. PMID:17107620

RDF SKETCH MAPS - KNOWLEDGE COMPLEXITY REDUCTION FOR PRECISION MEDICINE ANALYTICS.

PubMed

Thanintorn, Nattapon; Wang, Juexin; Ersoy, Ilker; Al-Taie, Zainab; Jiang, Yuexu; Wang, Duolin; Verma, Megha; Joshi, Trupti; Hammer, Richard; Xu, Dong; Shin, Dmitriy

2016-01-01

Realization of precision medicine ideas requires significant research effort to be able to spot subtle differences in complex diseases at the molecular level to develop personalized therapies. It is especially important in many cases of highly heterogeneous cancers. Precision diagnostics and therapeutics of such diseases demands interrogation of vast amounts of biological knowledge coupled with novel analytic methodologies. For instance, pathway-based approaches can shed light on the way tumorigenesis takes place in individual patient cases and pinpoint to novel drug targets. However, comprehensive analysis of hundreds of pathways and thousands of genes creates a combinatorial explosion, that is challenging for medical practitioners to handle at the point of care. Here we extend our previous work on mapping clinical omics data to curated Resource Description Framework (RDF) knowledge bases to derive influence diagrams of interrelationships of biomarker proteins, diseases and signal transduction pathways for personalized theranostics. We present RDF Sketch Maps - a computational method to reduce knowledge complexity for precision medicine analytics. The method of RDF Sketch Maps is inspired by the way a sketch artist conveys only important visual information and discards other unnecessary details. In our case, we compute and retain only so-called RDF Edges - places with highly important diagnostic and therapeutic information. To do this we utilize 35 maps of human signal transduction pathways by transforming 300 KEGG maps into highly processable RDF knowledge base. We have demonstrated potential clinical utility of RDF Sketch Maps in hematopoietic cancers, including analysis of pathways associated with Hairy Cell Leukemia (HCL) and Chronic Myeloid Leukemia (CML) where we achieved up to 20-fold reduction in the number of biological entities to be analyzed, while retaining most likely important entities. In experiments with pathways associated with HCL a generated RDF Sketch Map of the top 30% paths retained important information about signaling cascades leading to activation of proto-oncogene BRAF, which is usually associated with a different cancer, melanoma. Recent reports of successful treatments of HCL patients by the BRAF-targeted drug vemurafenib support the validity of the RDF Sketch Maps findings. We therefore believe that RDF Sketch Maps will be invaluable for hypothesis generation for precision diagnostics and therapeutics as well as drug repurposing studies.

Identification of Tyrosine Phosphorylated Proteins by SH2 Domain Affinity Purification and Mass Spectrometry.

PubMed

Buhs, Sophia; Gerull, Helwe; Nollau, Peter

2017-01-01

Phosphotyrosine signaling plays a major role in the control of many important biological functions such as cell proliferation and apoptosis. Deciphering of phosphotyrosine-dependent signaling is therefore of great interest paving the way for the understanding of physiological and pathological processes of signal transduction. On the basis of the specific binding of SH2 domains to phosphotyrosine residues, we here present an experimental workflow for affinity purification and subsequent identification of tyrosine phosphorylated proteins by mass spectrometry. In combination with SH2 profiling, a broadly applicable platform for the characterization of phosphotyrosine profiles in cell extracts, our pull down strategy enables researchers by now to identify proteins in signaling cascades which are differentially phosphorylated and selectively recognized by distinct SH2 domains.

Measuring phospholipase D activity in insulin-secreting pancreatic beta-cells and insulin-responsive muscle cells and adipocytes.

PubMed

Cazzolli, Rosanna; Huang, Ping; Teng, Shuzhi; Hughes, William E

2009-01-01

Phospholipase D (PLD) is an enzyme producing phosphatidic acid and choline through hydrolysis of phosphatidylcholine. The enzyme has been identified as a member of a variety of signal transduction cascades and as a key regulator of numerous intracellular vesicle trafficking processes. A role for PLD in regulating glucose homeostasis is emerging as the enzyme has recently been identified in events regulating exocytosis of insulin from pancreatic beta-cells and also in insulin-stimulated glucose uptake through controlling GLUT4 vesicle exocytosis in muscle and adipose tissue. We present methodologies for assessing cellular PLD activity in secretagogue-stimulated insulin-secreting pancreatic beta-cells and also insulin-stimulated adipocyte and muscle cells, two of the principal insulin-responsive cell types controlling blood glucose levels.

Phospholipase C-mediated hydrolysis of phosphatidylcholine is a target of transforming growth factor beta 1 inhibitory signals.

PubMed Central

Diaz-Meco, M T; Dominguez, I; Sanz, L; Municio, M M; Berra, E; Cornet, M E; Garcia de Herreros, A; Johansen, T; Moscat, J

1992-01-01

Cell growth and tumor transformation can be restrained in certain cell systems by the action of transforming growth factor beta (TGF-beta). It has been established that the mechanism whereby TGF-beta 1 inhibits cell growth does not interfere with the triggering of early mitogenic signal transduction mechanisms. Phospholipase C-catalyzed hydrolysis of phosphatidylcholine (PC) is a relatively late step in the cascade activated by growth factors. Therefore, conceivably activation of phospholipase C-catalyzed hydrolysis of PC could be the target of TGF-beta 1 action. In the study reported here, we demonstrate that TGF-beta 1 inhibits the coupling of ras p21 to the activation of PC hydrolysis, which appears to be critical for the antiproliferative effects of TGF-beta 1. Images PMID:1309592

Frontier of Epilepsy Research - mTOR signaling pathway

PubMed Central

2011-01-01

Studies of epilepsy have mainly focused on the membrane proteins that control neuronal excitability. Recently, attention has been shifting to intracellular proteins and their interactions, signaling cascades and feedback regulation as they relate to epilepsy. The mTOR (mammalian target of rapamycin) signal transduction pathway, especially, has been suggested to play an important role in this regard. These pathways are involved in major physiological processes as well as in numerous pathological conditions. Here, involvement of the mTOR pathway in epilepsy will be reviewed by presenting; an overview of the pathway, a brief description of key signaling molecules, a summary of independent reports and possible implications of abnormalities of those molecules in epilepsy, a discussion of the lack of experimental data, and questions raised for the understanding its epileptogenic mechanism. PMID:21467839

No significant association between the genetic polymorphisms in the GSK-3 beta gene and schizophrenia in the Chinese population.

PubMed

Meng, Junwei; Shi, Yongyong; Zhao, Xinzhi; Zhou, Jian; Zheng, Yonglan; Tang, Ruqi; Ma, Gang; Zhu, Xuming; He, Zangdong; Wang, Zhe; Xu, Yifeng; Feng, Guoyin; He, Lin

2008-04-01

The GSK-3 beta gene encodes a protein kinase which is abundant in the brain, and its product is involved in signal transduction cascades of neuronal cell development, energy metabolism and body pattern formation. Previous studies have suggested that GSK-3 beta might act as a potential candidate locus for schizophrenia susceptibility. We genotyped six SNPs within the gene and conducted a case-control study involving 329 schizophrenic patients and 288 healthy subjects in the Chinese population. We examined allele and genotype frequencies and haplotype distributions in the subtype of paranoid schizophrenic patients as well as schizophrenic subjects in general. Our results fail to replicate the association of the GSK-3 beta gene with susceptibility to schizophrenia in the Chinese population.

Localization of phosphatidylinositol signaling components in rat taste cells: Role in bitter taste transduction

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hwang, P.M.; Verma, A.; Bredt, D.S.

1990-10-01

To assess the role of phosphatidylinositol turnover in taste transduction we have visualized, in rat tongue, ATP-dependent endoplasmic reticular accumulation of {sup 45}Ca{sup 2+}, inositol 1,4,5-trisphosphate receptor binding sites, and phosphatidylinositol turnover monitored by autoradiography of ({sup 3}H)cytidine diphosphate diacylglycerol formed from ({sup 3}H)cytidine. Accumulated {sup 45}Ca{sup 2+}, inositol 1,4,5-trisphosphate receptors, and phosphatidylinositol turnover are selectively localized to apical areas of the taste buds of circumvallate papillae, which are associated with bitter taste. Further evidence for a role of phosphatidylinositol turnover in bitter taste is our observation of a rapid, selective increase in mass levels of inositol 1,4,5-trisphosphate elicited bymore » low concentrations of denatonium, a potently bitter tastant.« less

Novel optical methodologies in studying mechanical signal transduction in mammalian cells

NASA Technical Reports Server (NTRS)

Stamatas, G. N.; McIntire, L. V.

1999-01-01

For the last 3 decades evidence has been accumulating that some types of mammalian cells respond to their mechanically active environment by altering their morphology, growth rate, and metabolism. The study of such responses is very important in understanding, physiological and pathological conditions ranging from bone formation to atherosclerosis. Obtaining this knowledge has been the goal for an active research area in bioengineering termed cell mechanotransduction. The advancement of optical methodologies used in cell biology research has given the tools to elucidate cellular mechanisms that would otherwise be impossible to visualize. Combined with molecular biology techniques, they give engineers invaluable tools in understanding the chemical pathways involved in mechanotransduction. Herein we briefly review the current knowledge on mechanical signal transduction in mammalian cells, focusing on the application of novel optical techniques in the ongoing research.

Neural mechanisms of oculomotor abnormalities in the infantile strabismus syndrome.

PubMed

Walton, Mark M G; Pallus, Adam; Fleuriet, Jérome; Mustari, Michael J; Tarczy-Hornoch, Kristina

2017-07-01

Infantile strabismus is characterized by numerous visual and oculomotor abnormalities. Recently nonhuman primate models of infantile strabismus have been established, with characteristics that closely match those observed in human patients. This has made it possible to study the neural basis for visual and oculomotor symptoms in infantile strabismus. In this review, we consider the available evidence for neural abnormalities in structures related to oculomotor pathways ranging from visual cortex to oculomotor nuclei. These studies provide compelling evidence that a disturbance of binocular vision during a sensitive period early in life, whatever the cause, results in a cascade of abnormalities through numerous brain areas involved in visual functions and eye movements. Copyright © 2017 the American Physiological Society.

Oxidative and Molecular Responses in Capsicum annuum L. after Hydrogen Peroxide, Salicylic Acid and Chitosan Foliar Applications

PubMed Central

Mejía-Teniente, Laura; de Dalia Durán-Flores, Flor; Chapa-Oliver, Angela María; Torres-Pacheco, Irineo; Cruz-Hernández, Andrés; González-Chavira, Mario M.; Ocampo-Velázquez, Rosalía V.; Guevara-González, Ramón G.

2013-01-01

Hydrogen peroxide (H2O2) is an important ROS molecule (Reactive oxygen species) that serves as a signal of oxidative stress and activation of signaling cascades as a result of the early response of the plant to biotic stress. This response can also be generated with the application of elicitors, stable molecules that induce the activation of transduction cascades and hormonal pathways, which trigger induced resistance to environmental stress. In this work, we evaluated the endogenous H2O2 production caused by salicylic acid (SA), chitosan (QN), and H2O2 elicitors in Capsicum annuum L. Hydrogen peroxide production after elicitation, catalase (CAT) and phenylalanine ammonia lyase (PAL) activities, as well as gene expression analysis of cat1, pal, and pathogenesis-related protein 1 (pr1) were determined. Our results displayed that 6.7 and 10 mM SA concentrations, and, 14 and 18 mM H2O2 concentrations, induced an endogenous H2O2 and gene expression. QN treatments induced the same responses in lesser proportion than the other two elicitors. Endogenous H2O2 production monitored during several days, showed results that could be an indicator for determining application opportunity uses in agriculture for maintaining plant alert systems against a stress. PMID:23676352

A Molecular Smart Surface for Spatio-Temporal Studies of Cell Mobility

PubMed Central

Lee, Eun-ju; Luo, Wei; Chan, Eugene W. L.; Yousaf, Muhammad N.

2015-01-01

Active migration in both healthy and malignant cells requires the integration of information derived from soluble signaling molecules with positional information gained from interactions with the extracellular matrix and with other cells. How a cell responds and moves involves complex signaling cascades that guide the directional functions of the cytoskeleton as well as the synthesis and release of proteases that facilitate movement through tissues. The biochemical events of the signaling cascades occur in a spatially and temporally coordinated manner then dynamically shape the cytoskeleton in specific subcellular regions. Therefore, cell migration and invasion involve a precise but constantly changing subcellular nano-architecture. A multidisciplinary effort that combines new surface chemistry and cell biological tools is required to understand the reorganization of cytoskeleton triggered by complex signaling during migration. Here we generate a class of model substrates that modulate the dynamic environment for a variety of cell adhesion and migration experiments. In particular, we use these dynamic substrates to probe in real-time how the interplay between the population of cells, the initial pattern geometry, ligand density, ligand affinity and integrin composition affects cell migration and growth. Whole genome microarray analysis indicates that several classes of genes ranging from signal transduction to cytoskeletal reorganization are differentially regulated depending on the nature of the surface conditions. PMID:26030281

Distinct responses of human granulosa lutein cells after hCG or LH stimulation in a spheroidal cell culture system.

PubMed

Becker, Julia; Walz, Andrea; Daube, Stefanie; Keck, Christoph; Pietrowski, Detlef

2007-10-01

The growth and development of the corpus luteum (CL) is regulated by gonadotropic hormones. It is formed by granulosa cells (GCs), theca cells, and endothelial cells, and is the primary source of circulating progesterone. During early pregnancy only human chorionic gonadotropin (hCG) but not luteinizing hormone (LH) extends the life span of the CL, although hCG and LH interact with the same receptor and have similar actions on the CL. In this study a recently by our group established spheroidal GC culture assay served as a model of CL development on which we compared the actions of the gonadotropic hormones LH and hCG. To find out which signal pathways take part in the hormonal regulation of GC we stimulated GC-spheroids with modulators of protein kinases A and C dependent signaling cascades and determined their impact on sprout forming activity in GC. Our results indicate that PKA-dependent signaling pathways play a major role in mediating the hormonal-induced signaling cascades leading to sprouting in GC. Furthermore, this study strongly indicates that the different effects of hCG and LH in the maintenance of the CL may be reasoned in different signal transduction pathways triggered by hCG or LH. (c) 2007 Wiley-Liss, Inc.

Unsteady pressure measurements on a biconvex airfoil in a transonic oscillating cascade

NASA Technical Reports Server (NTRS)

Shaw, L. M.; Boldman, D. R.; Buggele, A. E.; Buffum, D. H.

1985-01-01

Flush-mounted dynamic pressure transducers were installed on the center airfoil of a transonic oscillating cascade to measure the unsteady aerodynamic response as nine airfroils were simultaneously driven to provide 1.2 deg of pitching motion about the midchord. Initial tests were performed at an incidence and angle of 0 deg and A Mach number of 0.65 in order to obtain results in a shock-free compressible flowfield. Subsequent tests were performed at an incidence angle of 7 deg and Mach number of 0.8 in order to observe the surface pressures with an oscillating shock near the leading edge of the airfoil. Results are presented for interblade phase angles of 90 and -90 deg and at blade oscillatory frequencies of 200 and 500 Hz (semi-chord reduced frequencies up to about 0.5 at a Mach number of 0.8). Results from the zero-incidence cascade are compared with a classical unsteady flat-plate analysis. Flow visualization results depicting the shock motion on the airfoils in the high-incidence cascade are discussed. The airfoil pressure data are tabulated.

Improved medical image fusion based on cascaded PCA and shift invariant wavelet transforms.

PubMed

Reena Benjamin, J; Jayasree, T

2018-02-01

In the medical field, radiologists need more informative and high-quality medical images to diagnose diseases. Image fusion plays a vital role in the field of biomedical image analysis. It aims to integrate the complementary information from multimodal images, producing a new composite image which is expected to be more informative for visual perception than any of the individual input images. The main objective of this paper is to improve the information, to preserve the edges and to enhance the quality of the fused image using cascaded principal component analysis (PCA) and shift invariant wavelet transforms. A novel image fusion technique based on cascaded PCA and shift invariant wavelet transforms is proposed in this paper. PCA in spatial domain extracts relevant information from the large dataset based on eigenvalue decomposition, and the wavelet transform operating in the complex domain with shift invariant properties brings out more directional and phase details of the image. The significance of maximum fusion rule applied in dual-tree complex wavelet transform domain enhances the average information and morphological details. The input images of the human brain of two different modalities (MRI and CT) are collected from whole brain atlas data distributed by Harvard University. Both MRI and CT images are fused using cascaded PCA and shift invariant wavelet transform method. The proposed method is evaluated based on three main key factors, namely structure preservation, edge preservation, contrast preservation. The experimental results and comparison with other existing fusion methods show the superior performance of the proposed image fusion framework in terms of visual and quantitative evaluations. In this paper, a complex wavelet-based image fusion has been discussed. The experimental results demonstrate that the proposed method enhances the directional features as well as fine edge details. Also, it reduces the redundant details, artifacts, distortions.

Stressor-induced NMDAR dysfunction as a unifying hypothesis for the aetiology, pathogenesis and comorbidity of clinical depression.

PubMed

Marsden, W N

2011-10-01

Typically the monoamine system has been the central focus of neurobiological research into depression and represents the major target of modern antidepressant medications; although the extent to which monoamines such as serotonin play a role in the pathogenesis of depression is still not clear. Recent research advancements have expanded the neurotransmitter-level focus of mood disorders to incorporate intracellular pathways and regional brain circuitry. As such the importance of other systems has emerged including those related to neuroplastic signal transduction and gene transcription cascades within cortico-limbic circuits. Indeed mounting evidence suggests interaction with these pathways is required for the chronic therapeutic effect of current clinical antidepressants. Dysfunction of the glutamatergic system has also emerged as a major pathological feature in depression, and glutamatergic agents have demonstrated rapid and robust antidepressant activity in humans. In particular, the glutamate receptors (AMPAR, NMDAR & mGluR) are intrinsically connected to neuronal efficiency and inefficiency cascades, so their dysfunction may account for alterations to multiple signal transduction pathways in depression. This article presents concepts supporting a NMDA hypothesis of depression, whereby the pathogenesis of depression may arise from stressors inducing excessive NMDAR activity which acts heterogeneously at both cellular and regional levels to disrupt normal neurobiological function and induce the depressive phenotype. In this hypothesis multiple psychological and environmental stressors are united in their capacity to potentiate excessive tonic and phasic NMDAR activation on neurons and glia. Such NMDAR dysfunction may lead to: disruption of glia processes and tripartite signalling; potentiation of extrasynaptic inefficiency/LTD pathways in some regions (e.g. prefrontal cortex & hippocampus); potentiation of synaptic efficiency/LTP pathways in other regions (e.g. amygdala); and regional disruption of cortico-limbic circuits and dopaminergic reward pathways (e.g. nucleus accumbens). This model unites depression with a variety of stressors including glucocorticoids, inflammation, oxidative stress, magnesium deficiency, hyperhomocysteinemia, and bio-energetic dysfunction; and also helps explain comorbidity with other neurological and affective disorders. In particular, a neurometabolic contribution to the aetiology of depressive as well as other neurological and affective disorders is explored. Copyright © 2011 Elsevier Ltd. All rights reserved.

Vane Separation Control in a Linear Cascade with Area Expansion using AC DBD Plasma Actuators

NASA Astrophysics Data System (ADS)

Kleven, Christopher; Corke, Thomas

2013-11-01

Experiments are presented on the use of AC dielectric barrier discharge (DBD) plasma actuators to prevent flow separation on vanes in a linear cascade with area expansion. The inlet Mach number to the cascade ranged from 0.3 to 0.5, and the vane chord Reynolds numbers ranged from 0 . 9 ×106 to 1 . 5 ×106 . Three cascade designs with different amounts of area expansion, providing different degrees of adverse pressure gradients, were examined. Surface flow visualization revealed a 3-D separation bubble with strong recirculation that formed on the suction side of the vanes. The pattern agreed well with CFD simulations. Plasma actuators were placed on the suction sides of the vanes, just upstream of the flow separation location. Quantitative measurements were performed in the wakes of the vanes using a 5-hole Pitot probe. The measurements were used to determine the effect of the plasma actuator separation control on the pressure loss coefficient, and flow turning angle through the cascades. Overall, the plasma actuators separation control increased the velocity magnitude and dynamic pressure in the passage between the vanes, resulted in a more spanwise-uniform flow turning angle in the vane passage, and significantly lowered the loss coefficient compared to the baseline.

The associations between multisensory temporal processing and symptoms of schizophrenia.

PubMed

Stevenson, Ryan A; Park, Sohee; Cochran, Channing; McIntosh, Lindsey G; Noel, Jean-Paul; Barense, Morgan D; Ferber, Susanne; Wallace, Mark T

2017-01-01

Recent neurobiological accounts of schizophrenia have included an emphasis on changes in sensory processing. These sensory and perceptual deficits can have a cascading effect onto higher-level cognitive processes and clinical symptoms. One form of sensory dysfunction that has been consistently observed in schizophrenia is altered temporal processing. In this study, we investigated temporal processing within and across the auditory and visual modalities in individuals with schizophrenia (SCZ) and age-matched healthy controls. Individuals with SCZ showed auditory and visual temporal processing abnormalities, as well as multisensory temporal processing dysfunction that extended beyond that attributable to unisensory processing dysfunction. Most importantly, these multisensory temporal deficits were associated with the severity of hallucinations. This link between atypical multisensory temporal perception and clinical symptomatology suggests that clinical symptoms of schizophrenia may be at least partly a result of cascading effects from (multi)sensory disturbances. These results are discussed in terms of underlying neural bases and the possible implications for remediation. Copyright © 2016 Elsevier B.V. All rights reserved.

Association of p21ras with phosphatidylinositol 3-kinase.

PubMed Central

Sjölander, A; Yamamoto, K; Huber, B E; Lapetina, E G

1991-01-01

In mammalian cells, ras genes code for 21-kDa GTP-binding proteins. Increased expression and mutations in specific amino acids have been closely linked to alterations of normal cell morphology, growth, and differentiation and, in particular, to neoplastic transformation. The signal transduction induced by these p21ras proteins is largely unknown; however, the signaling pathways of several growth factors have been reported to involve phosphatidylinositol (PtdIns) 3-kinase. In the present study of a Ha-ras-transformed epithelial cell line, we demonstrated increased PtdIns 3-kinase activity in anti-phosphotyrosine and anti-receptor (insulin and hybrid insulin-like growth factor I) immunoprecipitates of cells that had been stimulated with insulin or insulin-like growth factor I. The PtdIns 3-kinase activity was also immunoprecipitated in these experiments by the anti-Ras monoclonal antibody Y13-259. The specificity of this association with p21ras was ascertained by the neutralizing effect of the antigen peptide and the absence of PtdIns 3-kinase activity in Y13-259 immunoprecipitates from cells in which the ras gene was turned off. These data indicate that PtdIns 3-kinase activity is an important step in the cascade of reactions in p21ras signal transduction, suggesting that the alterations of the cytoskeleton and growth in ras-transformed cells could be mediated by PtdIns 3-kinase activity. Images PMID:1716764

Specialized sugar sensing in diverse fungi.

PubMed

Brown, Victoria; Sabina, Jeffrey; Johnston, Mark

2009-03-10

S. cerevisiae senses glucose and galactose differently. Glucose is detected through sensors that reside in the cellular plasma membrane. When activated, the sensors initiate a signal-transduction cascade that ultimately inactivates the Rgt1 transcriptional repressor by causing degradation of its corepressors Mth1 and Std1. This results in the expression of many HXT genes encoding glucose transporters. The ensuing flood of glucose into the cell activates Mig1, a transcriptional repressor that mediates "glucose repression" of many genes, including the GAL genes; hence, glucose sensing hinders galactose utilization. Galactose is sensed in the cytoplasm via Gal3. Upon binding galactose (and ATP), Gal3 sequesters the Gal80 protein, thereby emancipating the Gal4 transcriptional activator of the GAL genes. Gal4 also activates expression of MTH1, encoding a corepressor critical for Rgt1 function. Thus, galactose inhibits glucose assimilation by encouraging repression of HXT genes. C. albicans senses glucose similarly to S. cerevisiae but does not sense galactose through Gal3-Gal80-Gal4. Its genome harbors no GAL80 ortholog, and the severely truncated CaGal4 does not regulate CaGAL genes. We present evidence that C. albicans senses galactose with its Hgt4 glucose sensor, a capability that is enabled by transcriptional "rewiring" of its sugar-sensing signal-transduction pathways. We suggest that galactose sensing through Hgt4 is ancestral in fungi.

An Arabidopsis mutation in translation elongation factor 2 causes superinduction of CBF/DREB1 transcription factor genes but blocks the induction of their downstream targets under low temperatures.

PubMed

Guo, Yan; Xiong, Liming; Ishitani, Manabu; Zhu, Jian-Kang

2002-05-28

Low temperature regulates gene expression in bacteria, yeast, and animals as well as in plants. However, the signal transduction cascades mediating the low temperature responses are not well understood in any organism. To identify components in low temperature signaling genetically, we isolated Arabidopsis thaliana mutants in which cold-responsive genes are no longer induced by low temperatures. One of these mutations, los1-1, specifically blocks low temperature-induced transcription of cold-responsive genes. Surprisingly, cold-induced expression of the early response transcriptional activators, C-repeat/dehydration responsive element binding factors (CBF/DREB1s), is enhanced by the los1-1 mutation. The los1-1 mutation also reduces the capacity of plants to develop freezing tolerance but does not impair the vernalization response. Genetic analysis indicated that los1-1 is a recessive mutation in a single nuclear gene. The LOS1 gene encodes a translation elongation factor 2-like protein. Protein labeling studies show that new protein synthesis is blocked in los1-1 mutant plants specifically in the cold. These results reveal a critical role of new protein synthesis in the proper transduction of low temperature signals. Our results also suggest that cold-induced transcription of CBF/DREB1s is feedback inhibited by their gene products or by products of their downstream target genes.

Differential expression profiles of poplar MAP kinase kinases in response to abiotic stresses and plant hormones, and overexpression of PtMKK4 improves the drought tolerance of poplar.

PubMed

Wang, Lei; Su, Hongyan; Han, Liya; Wang, Chuanqi; Sun, Yanlin; Liu, Fenghong

2014-07-15

Mitogen-activated protein kinase (MAPK) cascades are universal signal transduction modules that play essential roles in plant growth, development and stress response. MAPK kinases (MAPKKs), which link MAPKs and MAPKK kinases (MAPKKKs), are integral in mediating various stress responses in plants. However, to date few data about the roles of poplar MAPKKs in stress signal transduction are available. In this study, we performed a systemic analysis of poplar MAPKK gene family expression profiles in response to several abiotic stresses and stress-associated hormones. Furthermore, Populus trichocarpa MAPKK4 (PtMKK4) was chosen for functional characterization. Transgenic analysis showed that overexpression of the PtMKK4 gene remarkably enhanced drought stress tolerance in the transgenic poplar plants. The PtMKK4-overexpressing plants also exhibited much lower levels of H2O2 and higher antioxidant enzyme activity after exposure to drought stress compared to the wide type lines. Besides, some drought marker genes including PtP5CS, PtSUS3, PtLTP3 and PtDREB8 exhibited higher expression levels in the transgenic lines than in the wide type under drought conditions. This study provided valuable information for understanding the putative functions of poplar MAPKKs involved in important signaling pathways under different stress conditions. Copyright © 2014 Elsevier B.V. All rights reserved.

Nasal solitary chemoreceptor cell responses to bitter and trigeminal stimulants in vitro.

PubMed

Gulbransen, Brian D; Clapp, Tod R; Finger, Thomas E; Kinnamon, Sue C

2008-06-01

Nasal trigeminal chemosensitivity in mice and rats is mediated in part by epithelial solitary chemoreceptor (chemosensory) cells (SCCs), but the exact role of these cells in chemoreception is unclear. Histological evidence suggests that SCCs express elements of the bitter taste transduction pathway including T2R (bitter taste) receptors, the G protein alpha-gustducin, PLCbeta2, and TRPM5, leading to speculation that SCCs are the receptor cells that mediate trigeminal nerve responses to bitter taste receptor ligands. To test this hypothesis, we used calcium imaging to determine whether SCCs respond to classic bitter-tasting or trigeminal stimulants. SCCs from the anterior nasal cavity were isolated from transgenic mice in which green fluorescent protein (GFP) expression was driven by either TRPM5 or gustducin. Isolated cells were exposed to a variety of test stimuli to determine which substances caused an increase in intracellular Ca2+ ([Ca2+]i). GFP-positive cells respond with increased [Ca2+]i to the bitter receptor ligand denatonium and this response is blocked by the PLC inhibitor U73122. In addition, GFP+ cells respond to the neuromodulators adenosine 5'-triphosphate and acetylcholine but only very rarely to other bitter-tasting or trigeminal stimuli. Our results demonstrate that TRPM5- and gustducin-expressing nasal SCCs respond to the T2R agonist denatonium via a PLC-coupled transduction cascade typical of T2Rs in the taste system.

HIV-1 Tat binds to SH3 domains: cellular and viral outcome of Tat/Grb2 interaction

PubMed Central

Rom, Slava; Pacifici, Marco; Passiatore, Giovanni; Aprea, Susanna; Waligorska, Agnieszka; Valle, Luis Del; Peruzzi, Francesca

2011-01-01

The Src-homology 3 (SH3) domain is one of the most frequent protein recognition modules (PRMs), being represented in signal transduction pathways and in several pathologies such as cancer and AIDS. Grb2 (growth factor receptor-bound protein 2) is an adaptor protein that contains two SH3 domains and is involved in receptor tyrosine kinase (RTK) signal transduction pathways. The HIV-1 transactivator factor Tat is required for viral replication and it has been shown to bind directly or indirectly to several host proteins, deregulating their functions. In this study, we show interaction between the cellular factor Grb2 and the HIV-1 trans-activating protein Tat. The binding is mediated by the proline-rich sequence of Tat and the SH3 domain of Grb2. As the adaptor protein Grb2 participates in a wide variety of signaling pathways, we characterized at least one of the possible downstream effects of the Tat/Grb2 interaction on the well-known IGF-1R/Raf/MAPK cascade. We show that the binding of Tat to Grb2 impairs activation of the Raf/MAPK pathway, while potentiating the PKA/Raf inhibitory pathway. The Tat/Grb2 interaction affects also viral function by inhibiting the Tat-mediated transactivation of HIV-1 LTR and viral replication in infected primary microglia. PMID:21745501

Elevated ATF4 Expression, in the Absence of Other Signals, Is Sufficient for Transcriptional Induction via CCAAT Enhancer-binding Protein-activating Transcription Factor Response Elements*

PubMed Central

Shan, Jixiu; Örd, Daima; Örd, Tõnis; Kilberg, Michael S.

2009-01-01

Protein limitation in vivo or amino acid deprivation of cells in culture causes a signal transduction cascade consisting of activation of the kinase GCN2 (general control nonderepressible 2), phosphorylation of eukaryotic initiation factor 2, and increased synthesis of activating transcription factor (ATF) 4 by a translational control mechanism. In a self-limiting transcriptional program, ATF4 transiently activates a wide range of downstream target genes involved in transport, cellular metabolism, and other cell functions. Simultaneous activation of other signal transduction pathways by amino acid deprivation led to the question of whether or not the increased abundance of ATF4 alone was sufficient to trigger the transcriptional control mechanisms. Using 293 cells that ectopically express ATF4 in a tetracycline-inducible manner showed that ATF4 target genes were activated in the absence of amino acid deprivation. Ectopic expression of ATF4 alone resulted in effective recruitment of the general transcription machinery, but some reduction in histone modification was observed. These data document that ATF4 alone is sufficient to trigger the amino acid-responsive transcriptional control program. However, the absolute amount of ectopic ATF4 required to achieve the same degree of transcriptional activation observed after amino acid limitation was greater, suggesting that other factors may serve to enhance ATF4 function. PMID:19509279

MEK1 inhibits cardiac PPARα activity by direct interaction and prevents its nuclear localization.

PubMed

el Azzouzi, Hamid; Leptidis, Stefanos; Bourajjaj, Meriem; van Bilsen, Marc; da Costa Martins, Paula A; De Windt, Leon J

2012-01-01

The response of the postnatal heart to growth and stress stimuli includes activation of a network of signal transduction cascades, including the stress activated protein kinases such as p38 mitogen-activated protein kinase (MAPK), c-Jun NH2-terminal kinase (JNK) and the extracellular signal-regulated kinase (ERK1/2) pathways. In response to increased workload, the mitogen-activated protein kinase kinase (MAPKK) MEK1 has been shown to be active. Studies embarking on mitogen-activated protein kinase (MAPK) signaling cascades in the heart have indicated peroxisome-proliferators activated-receptors (PPARs) as downstream effectors that can be regulated by this signaling cascade. Despite the importance of PPARα in controlling cardiac metabolism, little is known about the relationship between MAPK signaling and cardiac PPARα signaling. Using co-immunoprecipitation and immunofluorescence approaches we show a complex formation of PPARα with MEK1 and not with ERK1/2. Binding of PPARα to MEK1 is mediated via a LXXLL motif and results in translocation from the nucleus towards the cytoplasm, hereby disabling the transcriptional activity of PPARα. Mice subjected to voluntary running-wheel exercise showed increased cardiac MEK1 activation and complex formation with PPARα, subsequently resulting in reduced PPARα activity. Inhibition of MEK1, using U0126, blunted this effect. Here we show that activation of the MEK1-ERK1/2 pathway leads to specific inhibition of PPARα transcriptional activity. Furthermore we show that this inhibitory effect is mediated by MEK1, and not by its downstream effector kinase ERK1/2, through a mechanism involving direct binding to PPARα and subsequent stimulation of PPARα export from the nucleus.

Pigment Translocation in Caridean Shrimp Chromatophores: Receptor Type, Signal Transduction, Second Messengers, and Cross Talk Among Multiple Signaling Cascades.

PubMed

Milograna, Sarah Ribeiro; Ribeiro, Márcia Regina; Bell, Fernanda Tinti; McNamara, John Campbell

2016-11-01

Pigment aggregation in shrimp chromatophores is triggered by red pigment concentrating hormone (RPCH), a neurosecretory peptide whose plasma membrane receptor may be a G-protein coupled receptor (GPCR). While RPCH binding activates the Ca 2+ /cGMP signaling cascades, a role for cyclic AMP (cAMP) in pigment aggregation is obscure, as are the steps governing Ca 2+ release from the smooth endoplasmic reticulum (SER). A role for the antagonistic neuropeptide, pigment dispersing homone (α-PDH) is also unclear. In red, ovarian chromatophores from the freshwater shrimp Macrobrachium olfersi, we show that a G-protein antagonist (AntPG) strongly inhibits RPCH-triggered pigment aggregation, suggesting that RPCH binds to a GPCR, activating an inhibitory G-protein. Decreasing cAMP levels may cue pigment aggregation, since cytosolic cAMP titers, when augmented by cholera toxin, forskolin or vinpocentine, completely or partially impair pigment aggregation. Triggering opposing Ca 2+ /cGMP and cAMP cascades by simultaneous perfusion with lipid-soluble cyclic nucleotide analogs induces a "tug-of-war" response, pigments aggregating in some chromatosomes with unpredictable, oscillatory movements in others. Inhibition of cAMP-dependent protein kinase accelerates aggregation and reduces dispersion velocities, suggesting a role in phosphorylation events, possibly regulating SER Ca 2+ release and pigment aggregation. The second messengers IP 3 and cADPR do not stimulate SER Ca 2+ release. α-PDH does not sustain pigment dispersion, suggesting that pigment translocation in caridean chromatophores may be regulated solely by RPCH, since PDH is not required. We propose a working hypothesis to further unravel key steps in the mechanisms of pigment translocation within crustacean chromatophores that have remained obscure for nearly a century. © 2016 Wiley Periodicals, Inc.

The design and instrumentation of the Purdue annular cascade facility with initial data acquisition and analysis

NASA Technical Reports Server (NTRS)

Stauter, R. C.; Fleeter, S.

1982-01-01

Three dimensional aerodynamic data, required to validate and/or indicate necessary refinements to inviscid and viscous analyses of the flow through turbomachine blade rows, are discussed. Instrumentation and capabilities for pressure measurement, probe insertion and traversing, and flow visualization are reviewed. Advanced measurement techniques including Laser Doppler Anemometers, are considered. Data processing is reviewed. Predictions were correlated with the experimental data. A flow visualization technique using helium filled soap bubbles was demonstrated.

Sensitivity to timing and order in human visual cortex

PubMed Central

Singer, Jedediah M.; Madsen, Joseph R.; Anderson, William S.

2014-01-01

Visual recognition takes a small fraction of a second and relies on the cascade of signals along the ventral visual stream. Given the rapid path through multiple processing steps between photoreceptors and higher visual areas, information must progress from stage to stage very quickly. This rapid progression of information suggests that fine temporal details of the neural response may be important to the brain's encoding of visual signals. We investigated how changes in the relative timing of incoming visual stimulation affect the representation of object information by recording intracranial field potentials along the human ventral visual stream while subjects recognized objects whose parts were presented with varying asynchrony. Visual responses along the ventral stream were sensitive to timing differences as small as 17 ms between parts. In particular, there was a strong dependency on the temporal order of stimulus presentation, even at short asynchronies. From these observations we infer that the neural representation of complex information in visual cortex can be modulated by rapid dynamics on scales of tens of milliseconds. PMID:25429116

Visualizing autophosphorylation in histidine kinases.

PubMed

Casino, Patricia; Miguel-Romero, Laura; Marina, Alberto

2014-01-01

Reversible protein phosphorylation is the most widespread regulatory mechanism in signal transduction. Autophosphorylation in a dimeric sensor histidine kinase is the first step in two-component signalling, the predominant signal-transduction device in bacteria. Despite being the most abundant sensor kinases in nature, the molecular bases of the histidine kinase autophosphorylation mechanism are still unknown. Furthermore, it has been demonstrated that autophosphorylation can occur in two directions, cis (intrasubunit) or trans (intersubunit) within the dimeric histidine kinase. Here, we present the crystal structure of the complete catalytic machinery of a chimeric histidine kinase. The structure shows an asymmetric histidine kinase dimer where one subunit is caught performing the autophosphorylation reaction. A structure-guided functional analysis on HK853 and EnvZ, two prototypical cis- and trans-phosphorylating histidine kinases, has allowed us to decipher the catalytic mechanism of histidine kinase autophosphorylation, which seems to be common independently of the reaction directionality.

Nitric oxide inhibits exocytosis of cytolytic granules from lymphokine-activated killer cells

PubMed Central

Ferlito, Marcella; Irani, Kaikobad; Faraday, Nauder; Lowenstein, Charles J.

2006-01-01

NO inhibits cytotoxic T lymphocyte killing of target cells, although the precise mechanism is unknown. We hypothesized that NO decreases exocytosis of cytotoxic granules from activated lymphocytes. We now show that NO inhibits lymphokine-activated killer cell killing of K562 target cells. Exogenous and endogenous NO decreases the release of granzyme B, granzyme A, and perforin: all contents of cytotoxic granules. NO inhibits the signal transduction cascade initiated by cross-linking of the T cell receptor that leads to granule exocytosis. In particular, we found that NO decreases the expression of Ras, a critical signaling component within the exocytic pathway. Ectopic expression of Ras prevents NO inhibition of exocytosis. Our data suggest that Ras mediates NO inhibition of lymphocyte cytotoxicity and emphasize that alterations in the cellular redox state may regulate the exocytic signaling pathway. PMID:16857739

Cell proliferation inhibition in reduced gravity

NASA Technical Reports Server (NTRS)

Moos, P. J.; Fattaey, H. K.; Johnson, T. C.; Spooner, B. S. (Principal Investigator)

1994-01-01

Extended durations of spaceflight have been shown to be deleterious on an organismic level; however, mechanisms underlying cellular sensitivity to the gravitational environment remain to be elucidated. The majority of the gravitational studies to date indicates that cell regulatory pathways may be influenced by their gravitational environment. Still, few cell biology experiments have been performed in space flight and even fewer experiments have been repeated on subsequent flights. With flight opportunities on STS-50, 54, and 57, Sf9 cells were flown in the BioServe Fluids Processing Apparatus and cell proliferation was measured with and without exposure to a cell regulatory sialoglycopeptide (CeReS) inhibitor. Results from these flights indicate that the Sf9 cells grew comparable to ground controls, that the CeReS inhibitor bound to its specific receptor, and that its signal transduction cascade was not gravity sensitive.

High affinity binding of a fungal oligopeptide elicitor to parsley plasma membranes triggers multiple defense responses.

PubMed

Nürnberger, T; Nennstiel, D; Jabs, T; Sacks, W R; Hahlbrock, K; Scheel, D

1994-08-12

An oligopeptide of 13 amino acids (Pep-13) identified within a 42 kDa glycoprotein elicitor from P. mega-sperma was shown to be necessary and sufficient to stimulate a complex defense response in parsley cells comprising H+/Ca2+ influxes, K+/Cl- effluxes, an oxidative burst, defense-related gene activation, and phytoalexin formation. Binding of radiolabeled Pep-13 to parsley microsomes and protoplasts was specific, reversible, and saturable. Identical structural features of Pep-13 were found to be responsible for specific binding and initiation of all plant responses analyzed. The high affinity binding site recognizing the peptide ligand (KD = 2.4 nM) may therefore represent a novel class of receptors in plants, and the rapidly induced ion fluxes may constitute elements of the signal transduction cascade triggering pathogen defense in plants.

Age-related changes in the response of intestinal cells to parathyroid hormone.

PubMed

Russo de Boland, Ana

2004-12-01

The concept of the role(s) of parathyroid hormone (PTH), has expanded from that on acting on the classical target tissues, bone and kidney, to the intestine where its actions are of regulatory and developmental importance: regulation of intracellular calcium through modulation of second messengers and, activation of mitogenic cascades leading to cell proliferation. Several causes have been postulated to modify the hormone response in intestinal cells with ageing, among them, alterations of PTH receptor (PTHR1) binding sites, reduced expression of G proteins and hormone signal transduction changes. The current review summarizes the actual knowledge regarding the molecular and biochemical basis of age-impaired PTH receptor-mediated signaling in intestinal cells. A fundamental understanding of why PTH functions are impaired with age will enhance our understanding of its importance in intestinal cell physiology.

Mechano-sensing and transduction by endothelial surface glycocalyx: composition, structure, and function

PubMed Central

Fu, Bingmei M.; Tarbell, John M.

2014-01-01

The endothelial cells (ECs) lining every blood vessel wall are constantly exposed to the mechanical forces generated by blood flow. The EC responses to these hemodynamic forces play a critical role in the homeostasis of the circulatory system. To ensure proper EC mechano-sensing and transduction, there are a variety of mechano-sensors and transducers that have been identified on the EC surface, intra- and trans-EC membrane and within the EC cytoskeleton. Among them, the most recent candidate is the endothelial surface glycocalyx (ESG), which is a matrix-like thin layer covering the luminal surface of the EC. It consists of various proteoglycans, glycosaminoglycans, and plasma proteins, and is close to other prominent EC mechano-sensors and transducers. The ESG thickness was found to be in the order of 0.1–1 μm by different visualization techniques and in different types of vessels. Detailed analysis on the electron microscopy (EM) images of the microvascular ESG revealed a quasi-periodic substructure with the ESG fiber diameter of 10–12 and 20 nm spacing between adjacent fibers. Atomic force microscopy and optical tweezers were applied to investigate the mechanical properties of the ESG on the cultured EC monolayers and in solutions. Enzymatic degradation of specific ESG glycosaminoglycan components was used to directly elucidate the role of the ESG in EC mechano-sensing and transduction by measuring the shear-induced productions of nitric oxide and prostacyclin, two characteristic responses of the ECs to the flow. The unique location, composition, and structure of the ESG determine its role in EC mechano-sensing and transduction. PMID:23401243

Effects of harvest on carbon and nitrogen dynamics in a Pacific Northwest forest catchment

EPA Science Inventory

We used a new ecohydrological model, Visualizing Ecosystems for Land Management Assessments (VELMA), to analyze the effects of forest harvest on catchment carbon and nitrogen dynamics. We applied the model to a 10 ha headwater catchment in the western Oregon Cascade Range where t...

Neurally and Ocularly Informed Graph-Based Models for Searching 3D Environments

DTIC Science & Technology

2014-06-03

hBCI = hybrid brain–computer interface, TAG = transductive annotation by graph, CV = computer vision, TSP = traveling salesman problem . are navigated...environment that are most likely to contain objects that the subject would like to visit. 2.9. Route planning A traveling salesman problem (TSP) solver...fixations in a visual search task using fixation-related potentials J. Vis. 13 Croes G 1958 A method for solving traveling - salesman problems Oper. Res

The D153del Mutation in GNB3 Gene Causes Tissue Specific Signalling Patterns and an Abnormal Renal Morphology in Rge Chickens

PubMed Central

Tummala, Hemanth; Fleming, Stewart; Hocking, Paul M.; Wehner, Daniel; Naseem, Zahid; Ali, Manir; Inglehearn, Christopher F.; Zhelev, Nikolai; Lester, Douglas H.

2011-01-01

Background The GNB3 gene is expressed in cone but not rod photoreceptors of vertebrates, where it acts as the β transducin subunit in the colour visual transduction process. A naturally occurring mutation ‘D153del’ in the GNB3 gene causes the recessively inherited blinding phenotype retinopathy globe enlarged (rge) disease in chickens. GNB3 is however also expressed in most other vertebrate tissues suggesting that the D153del mutation may exert pathological effects that outlie from eye. Principal Findings Recombinant studies in COS-7 cells that were transfected with normal and mutant recombinant GNB3 constructs and subjected to cycloheximide chase showed that the mutant GNB3d protein had a much shorter half life compared to normal GNB3. GNB3 codes for the Gβ3 protein subunit that, together with different Gγ and Gα subunits, activates and regulates phosphorylation cascades in different tissues. As expected, the relative levels of cGMP and cAMP secondary messengers and their activated kinases such as MAPK, AKT and GRK2 were also found to be altered significantly in a tissue specific manner in rge chickens. Histochemical analysis on kidney tissue sections, from rge homozygous affected chickens, showed the chickens had enlargement of the glomerular capsule, causing glomerulomegaly and tubulointerstitial inflammation whereas other tissues (brain, heart, liver, pancreas) were unaffected. Significance These findings confirm that the D153del mutation in GNB3 gene targets GNB3 protein to early degradation. Lack of GNB3 signalling causes reduced phosphorylation activity of ERK2 and AKT leading to severe pathological phenotypes such as blindness and renal abnormalities in rge chickens. PMID:21887213

Evidence of solitary chemosensory cells in a large mammal: the diffuse chemosensory system in Bos taurus airways

PubMed Central

Tizzano, Marco; Merigo, Flavia; Sbarbati, Andrea

2006-01-01

The diffuse chemosensory system (DCS) of the respiratory apparatus is composed of solitary chemosensory cells (SCCs) that resemble taste cells but are not organized in end organs. The discovery of the DCS may open up new approaches to respiratory diseases. However, available data on mammalian SCCs have so far been collected from rodents, the airways of which display some differences from those of large mammals. Here we investigated the presence of the DCS and of SCCs in cows and bulls (Bos taurus), in which the airway cytology is similar to that in humans, focusing our attention on detection in the airways of molecules involved in the transduction cascade of taste [i.e. α-gustducin and phospholipase C of the β2 subtype (PLCβ2)]. The aim of the research was to extend our understanding of airway chemoreceptors and to compare the organization of the DCS in a large mammal with that in rodents. Using immunocytochemistry for α-gustducin, the taste buds of the tongue and arytenoid were visualized. In the trachea and bronchi, α-gustducin-immunoreactive SCCs were frequently found. Using immunocytochemistry for PLCβ2, the staining pattern was generally similar to those seen for α-gustducin. Immunoblotting confirmed the expression of α-gustducin in the tongue and in all the airway regions tested. The study demonstrated the presence of SCCs in cows and bulls, suggesting that DCSs are present in many mammalian species. The description of areas with a high density of SCCs in bovine bronchi seems to indicate that the view of the DCS as made up of isolated cells totally devoid of ancillary elements is probably an oversimplification. PMID:16928202

Unique structural features of the AIPL1–FKBP domain that support prenyl lipid binding and underlie protein malfunction in blindness

PubMed Central

Yadav, Ravi P.; Gakhar, Lokesh; Yu, Liping

2017-01-01

FKBP-domain proteins (FKBPs) are pivotal modulators of cellular signaling, protein folding, and gene transcription. Aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1) is a distinctive member of the FKBP superfamily in terms of its biochemical properties, and it plays an important biological role as a chaperone of phosphodiesterase 6 (PDE6), an effector enzyme of the visual transduction cascade. Malfunction of mutant AIPL1 proteins triggers a severe form of Leber congenital amaurosis and leads to blindness. The mechanism underlying the chaperone activity of AIPL1 is largely unknown, but involves the binding of isoprenyl groups on PDE6 to the FKBP domain of AIPL1. We solved the crystal structures of the AIPL1–FKBP domain and its pathogenic mutant V71F, both in the apo form and in complex with isoprenyl moieties. These structures reveal a module for lipid binding that is unparalleled within the FKBP superfamily. The prenyl binding is enabled by a unique “loop-out” conformation of the β4-α1 loop and a conformational “flip-out” switch of the key W72 residue. A second major conformation of apo AIPL1–FKBP was identified by NMR studies. This conformation, wherein W72 flips into the ligand-binding pocket and renders the protein incapable of prenyl binding, is supported by molecular dynamics simulations and appears to underlie the pathogenicity of the V71F mutant. Our findings offer critical insights into the mechanisms that underlie AIPL1 function in health and disease, and highlight the structural and functional diversity of the FKBPs. PMID:28739921

Identification of the gamma subunit-interacting residues on photoreceptor cGMP phosphodiesterase, PDE6alpha '.

PubMed

Granovsky, A E; Artemyev, N O

2000-12-29

Photoreceptor cGMP phosphodiesterase (PDE6) is the effector enzyme in the G protein-mediated visual transduction cascade. In the dark, the activity of PDE6 is shut off by the inhibitory gamma subunit (Pgamma). Chimeric proteins between cone PDE6alpha' and cGMP-binding and cGMP-specific PDE (PDE5) have been constructed and expressed in Sf9 cells to study the mechanism of inhibition of PDE6 catalytic activity by Pgamma. Substitution of the segment PDE5-(773-820) by the corresponding PDE6alpha'-(737-784) sequence in the wild-type PDE5 or in a PDE5/PDE6alpha' chimera containing the catalytic domain of PDE5 results in chimeric enzymes capable of inhibitory interaction with Pgamma. The catalytic properties of the chimeric PDEs remained similar to those of PDE5. Ala-scanning mutational analysis of the Pgamma-binding region, PDE6alpha'-(750-760), revealed PDE6alpha' residues essential for the interaction. The M758A mutation markedly impaired and the Q752A mutation moderately impaired the inhibition of chimeric PDE by Pgamma. The analysis of the catalytic properties of mutant PDEs and a model of the PDE6 catalytic domain suggest that residues Met(758) and Gln(752) directly bind Pgamma. A model of the PDE6 catalytic site shows that PDE6alpha'-(750-760) forms a loop at the entrance to the cGMP-binding pocket. Binding of Pgamma to Met(758) would effectively block access of cGMP to the catalytic cavity, providing a structural basis for the mechanism of PDE6 inhibition.

Anagrelide represses GATA-1 and FOG-1 expression without interfering with thrombopoietin receptor signal transduction.

PubMed

Ahluwalia, M; Donovan, H; Singh, N; Butcher, L; Erusalimsky, J D

2010-10-01

 Anagrelide is a selective inhibitor of megakaryocytopoiesis used to treat thrombocytosis in patients with chronic myeloproliferative disorders. The effectiveness of anagrelide in lowering platelet counts is firmly established, but its primary mechanism of action remains elusive.  Here, we have evaluated whether anagrelide interferes with the major signal transduction cascades stimulated by thrombopoietin in the hematopoietic cell line UT-7/mpl and in cultured CD34(+) -derived human hematopoietic cells. In addition, we have used quantitative mRNA expression analysis to assess whether the drug affects the levels of known transcription factors that control megakaryocytopoiesis.  In UT-7/mpl cells, anagrelide (1μm) did not interfere with MPL-mediated signaling as monitored by its lack of effect on JAK2 phosphorylation. Similarly, the drug did not affect the phosphorylation of STAT3, ERK1/2 or AKT in either UT-7/mpl cells or primary hematopoietic cells. In contrast, during thrombopoietin-induced megakaryocytic differentiation of normal hematopoietic cultures, anagrelide (0.3μm) reduced the rise in the mRNA levels of the transcription factors GATA-1 and FOG-1 as well as those of the downstream genes encoding FLI-1, NF-E2, glycoprotein IIb and MPL. However, the drug showed no effect on GATA-2 or RUNX-1 mRNA expression. Furthermore, anagrelide did not diminish the rise in GATA-1 and FOG-1 expression during erythropoietin-stimulated erythroid differentiation. Cilostamide, an exclusive and equipotent phosphodiesterase III (PDEIII) inhibitor, did not alter the expression of these genes.  Anagrelide suppresses megakaryocytopoiesis by reducing the expression levels of GATA-1 and FOG-1 via a PDEIII-independent mechanism that is differentiation context-specific and does not involve inhibition of MPL-mediated early signal transduction events. © 2010 International Society on Thrombosis and Haemostasis.

Stimulation of spinal dorsal horn β2-adrenergic receptor ameliorates neuropathic mechanical hypersensitivity through a reduction of phosphorylation of microglial p38 MAP kinase and astrocytic c-jun N-terminal kinase.

PubMed

Zhang, Fang Fang; Morioka, Norimitsu; Abe, Hiromi; Fujii, Shiori; Miyauchi, Kazuki; Nakamura, Yoki; Hisaoka-Nakashima, Kazue; Nakata, Yoshihiro

2016-12-01

The noradrenaline-adrenergic system has a crucial role in controlling nociceptive transduction at the spinal level. While α-adrenergic receptors are known to regulate nociceptive neurotransmitter release at the spinal presynaptic level, it is not entirely clear whether β-adrenergic receptors are involved in controlling pain transduction at the spinal level as well. The current study elucidated a role of β-adrenergic receptors in neuropathic pain in mice following a partial sciatic nerve ligation (PSNL). In addition, the cellular and intracellular signaling cascade induced by β-adrenergic receptors in neuropathic mice was elaborated. Intrathecal injection of isoproterenol (1 nmol), a nonselective β-adrenergic receptor agonist, briefly ameliorated hind paw mechanical hypersensitivity of PSNL mice. Isoproterenol's antinociceptive effect was mediated through β2-adrenergic receptors since pretreatment with ICI118551, a selective β2-adrenergic receptor antagonist, but not with CGP20712A, a selective β1-adrenergic receptor antagonist, significantly attenuated isoproterenol's effect. Furthermore, intrathecal treatment with a selective β2-adrenergic receptor agonist, terbutaline, but not a selective β1-adrenergic receptor agonist, dobutamine, also significantly ameliorated neuropathic pain. Fourteen days after PSNL, increased phosphorylation of both p38 Mitogen-activated protein kinase (MAPK) in microglia and c-jun N-terminal kinase (JNK) in astrocytes of ipsilateral spinal dorsal horn were observed. Phosphorylation of both microglial p38 MAPK and astrocytic JNK were downregulated by stimulation of the β2-adrenergic receptor. Together, these results suggest that spinal β2-adrenergic receptor have an inhibitory role in neuropathic nociceptive transduction at the spinal level through a downregulation of glial activity, perhaps through modulation of MAP kinases phosphorylation. Thus, targeting of β2-adrenergic receptors could be an effective therapeutic strategy in treating neuropathic pain. Copyright © 2016 Elsevier Ltd. All rights reserved.

Evolution of the vertebrate phototransduction cascade activation steps.

PubMed

Lamb, Trevor D; Hunt, David M

2017-11-01

We examine the molecular phylogeny of the proteins underlying the activation steps of vertebrate phototransduction, for both agnathan and jawed vertebrate taxa. We expand the number of taxa analysed and we update the alignment and tree building methodology from a previous analysis. For each of the four primary components (the G-protein transducin alpha subunit, Gα T , the cyclic GMP phosphodiesterase, PDE6, and the alpha and beta subunits of the cGMP-gated ion channel, CNGC), the phylogenies appear consistent with expansion from an ancestral proto-vertebrate cascade during two rounds of whole-genome duplication followed by divergence of the agnathan and jawed vertebrate lineages. In each case, we consider possible scenarios for the underlying gene duplications and losses, and we apply relevant constraints to the tree construction. From tests of the topology of the resulting trees, we obtain a scenario for the expansion of each component during 2R that accurately fits the observations. Similar analysis of the visual opsins indicates that the only expansion to have occurred during 2R was the formation of Rh1 and Rh2. Finally, we propose a hypothetical scenario for the conversion of an ancestral chordate cascade into the proto-vertebrate phototransduction cascade, prior to whole-genome duplication. Together, our models provide a plausible account for the origin and expansion of the vertebrate phototransduction cascade. Copyright © 2017 Elsevier Inc. All rights reserved.

Molecular Mechanisms at the Basis of Plasticity in the Developing Visual Cortex: Epigenetic Processes and Gene Programs

PubMed Central

Maya-Vetencourt, José Fernando; Pizzorusso, Tommaso

2013-01-01

Neuronal circuitries in the mammalian visual system change as a function of experience. Sensory experience modifies neuronal networks connectivity via the activation of different physiological processes such as excitatory/inhibitory synaptic transmission, neurotrophins, and signaling of extracellular matrix molecules. Long-lasting phenomena of plasticity occur when intracellular signal transduction pathways promote epigenetic alterations of chromatin structure that regulate the induction of transcription factors that in turn drive the expression of downstream targets, the products of which then work via the activation of structural and functional mechanisms that modify synaptic connectivity. Here, we review recent findings in the field of visual cortical plasticity while focusing on how physiological mechanisms associated with experience promote structural changes that determine functional modifications of neural circuitries in V1. We revise the role of microRNAs as molecular transducers of environmental stimuli and the role of immediate early genes that control gene expression programs underlying plasticity in the developing visual cortex. PMID:25157210

Why pleiotropic interventions are needed for Alzheimer's disease.

PubMed

Frautschy, Sally A; Cole, Greg M

2010-06-01

Alzheimer's disease (AD) involves a complex pathological cascade thought to be initially triggered by the accumulation of beta-amyloid (Abeta) peptide aggregates or aberrant amyloid precursor protein (APP) processing. Much is known of the factors initiating the disease process decades prior to the onset of cognitive deficits, but an unclear understanding of events immediately preceding and precipitating cognitive decline is a major factor limiting the rapid development of adequate prevention and treatment strategies. Multiple pathways are known to contribute to cognitive deficits by disruption of neuronal signal transduction pathways involved in memory. These pathways are altered by aberrant signaling, inflammation, oxidative damage, tau pathology, neuron loss, and synapse loss. We need to develop stage-specific interventions that not only block causal events in pathogenesis (aberrant tau phosphorylation, Abeta production and accumulation, and oxidative damage), but also address damage from these pathways that will not be reversed by targeting prodromal pathways. This approach would not only focus on blocking early events in pathogenesis, but also adequately correct for loss of synapses, substrates for neuroprotective pathways (e.g., docosahexaenoic acid), defects in energy metabolism, and adverse consequences of inappropriate compensatory responses (aberrant sprouting). Monotherapy targeting early single steps in this complicated cascade may explain disappointments in trials with agents inhibiting production, clearance, or aggregation of the initiating Abeta peptide or its aggregates. Both plaque and tangle pathogenesis have already reached AD levels in the more vulnerable brain regions during the "prodromal" period prior to conversion to "mild cognitive impairment (MCI)." Furthermore, many of the pathological events are no longer proceeding in series, but are going on in parallel. By the MCI stage, we stand a greater chance of success by considering pleiotropic drugs or cocktails that can independently limit the parallel steps of the AD cascade at all stages, but that do not completely inhibit the constitutive normal functions of these pathways. Based on this hypothesis, efforts in our laboratories have focused on the pleiotropic activities of omega-3 fatty acids and the anti-inflammatory, antioxidant, and anti-amyloid activity of curcumin in multiple models that cover many steps of the AD pathogenic cascade (Cole and Frautschy, Alzheimers Dement 2:284-286, 2006).

Comparison of visualized turbine endwall secondary flows and measured heat transfer patterns

NASA Technical Reports Server (NTRS)

Gaugler, R. E.; Russell, L. M.

1983-01-01

Various flow visualization techniques were used to define the secondary flows near the endwall in a large heat transfer data. A comparison of the visualized flow patterns and the measured Stanton number distribution was made for cases where the inlet Reynolds number and exit Mach number were matched. Flows were visualized by using neutrally buoyant helium-filled soap bubbles, by using smoke from oil soaked cigars, and by a few techniques using permanent marker pen ink dots and synthetic wintergreen oil. Details of the horseshoe vortex and secondary flows can be directly compared with heat transfer distribution. Near the cascade entrance there is an obvious correlation between the two sets of data, but well into the passage the effect of secondary flow is not as obvious.

Promoter-level expression clustering identifies time development of transcriptional regulatory cascades initiated by ErbB receptors in breast cancer cells.

PubMed

Mina, Marco; Magi, Shigeyuki; Jurman, Giuseppe; Itoh, Masayoshi; Kawaji, Hideya; Lassmann, Timo; Arner, Erik; Forrest, Alistair R R; Carninci, Piero; Hayashizaki, Yoshihide; Daub, Carsten O; Okada-Hatakeyama, Mariko; Furlanello, Cesare

2015-07-16

The analysis of CAGE (Cap Analysis of Gene Expression) time-course has been proposed by the FANTOM5 Consortium to extend the understanding of the sequence of events facilitating cell state transition at the level of promoter regulation. To identify the most prominent transcriptional regulations induced by growth factors in human breast cancer, we apply here the Complexity Invariant Dynamic Time Warping motif EnRichment (CIDER) analysis approach to the CAGE time-course datasets of MCF-7 cells stimulated by epidermal growth factor (EGF) or heregulin (HRG). We identify a multi-level cascade of regulations rooted by the Serum Response Factor (SRF) transcription factor, connecting the MAPK-mediated transduction of the HRG stimulus to the negative regulation of the MAPK pathway by the members of the DUSP family phosphatases. The finding confirms the known primary role of FOS and FOSL1, members of AP-1 family, in shaping gene expression in response to HRG induction. Moreover, we identify a new potential regulation of DUSP5 and RARA (known to antagonize the transcriptional regulation induced by the estrogen receptors) by the activity of the AP-1 complex, specific to HRG response. The results indicate that a divergence in AP-1 regulation determines cellular changes of breast cancer cells stimulated by ErbB receptors.

Effects of nitric oxide on expressions of nitrosocysteine and calcium-activated potassium channels in the supraoptic nuclei and neural lobe of dehydrated rats

PubMed Central

Kadekaro, Massako; Su, Guangxiao; Chu, Rong; Lei, Yongzhong; Li, Junfa; Fang, Li

2007-01-01

Nitric oxide (NO) is an important gas mediator in the signal transduction cascade regulating osmotic function in the hypothalamo-neurohypophysial system. We previously found that increased nitric oxide synthase (NOS) activity in the supraoptic nuclei (SON) and neural lobe following osmotic stimulation and NO could regulate the expression of Ca2+-activated K+ channel (BK channels) protein in the magnocellular system during dehydration. The aim of the current study is to examine the role of NO in the regulation of nitrosocysteine and BK channel protein in the magnocellular system in dehydrated animals. Using Western blot analysis and quantitative immunofluorescent staining study, we found that water deprivation in rats significantly enhanced the expression of nitrosocysteine protein in SON and neural lobes. Immunohistochemistry study indicated that dehydration significantly increased the profiles of SON neurons co-expressing nitrosocysteine with BK-channel protein. Intracerebroventricular administration of L-NAME (an inhibitor of NO synthase) significantly reduced the neuronal profiles of nitrosocysteine, as well as their co-expression with BK-channel in SON of dehydrated rats. However, treatment of sodium nitroprusside (a donor of NO) increased this co-expression. Our results indicate that NO signaling cascade may control the expression of BK channels through the regulation of nitrosocysteine in SON and neural lobe of rats during osmotic regulation. PMID:17098363

Modulation of transcription factors by curcumin.

PubMed

Shishodia, Shishir; Singh, Tulika; Chaturvedi, Madan M

2007-01-01

Curcumin is the active ingredient of turmeric that has been consumed as a dietary spice for ages. Turmeric is widely used in traditional Indian medicine to cure biliary disorders, anorexia, cough, diabetic wounds, hepatic disorders, rheumatism, and sinusitis. Extensive investigation over the last five decades has indicated that curcumin reduces blood cholesterol, prevents low-density lipoprotein oxidation, inhibits platelet aggregation, suppresses thrombosis and myocardial infarction, suppresses symptoms associated with type II diabetes, rheumatoid arthritis, multiple sclerosis, and Alzheimer's disease, inhibits HIV replication, enhances wound healing, protects from liver injury, increases bile secretion, protects from cataract formation, and protects from pulmonary toxicity and fibrosis. Evidence indicates that the divergent effects of curcumin are dependent on its pleiotropic molecular effects. These include the regulation of signal transduction pathways and direct modulation of several enzymatic activities. Most of these signaling cascades lead to the activation of transcription factors. Curcumin has been found to modulate the activity of several key transcription factors and, in turn, the cellular expression profiles. Curcumin has been shown to elicit vital cellular responses such as cell cycle arrest, apoptosis, and differentiation by activating a cascade of molecular events. In this chapter, we briefly review the effects of curcumin on transcription factors NF-KB, AP-1, Egr-1, STATs, PPAR-gamma, beta-catenin, nrf2, EpRE, p53, CBP, and androgen receptor (AR) and AR-related cofactors giving major emphasis to the molecular mechanisms of its action.

miR-302b inhibits tumorigenesis by targeting EphA2 via Wnt/ β-catenin/EMT signaling cascade in gastric cancer.

PubMed

Huang, Jin; He, Yijing; Mcleod, Howard L; Xie, Yanchun; Xiao, Desheng; Hu, Huabin; Chen, Pan; Shen, Liangfang; Zeng, Shan; Yin, Xianli; Ge, Jie; Li, Li; Tang, Lanhua; Ma, Jian; Chen, Zihua

2017-12-22

EphA2 is a crucial oncogene in gastric cancer (GC) development and metastasis, this study aims to identify microRNAs that target it and serve as key regulators of gastric carcinogenesis. We identified several potential microRNAs targeting EphA2 by bioinformatics websites and then analyzed the role of miR-302b in modulating EphA2 in vitro and in vivo of GC, and it's mechanism. Our analysis identified miR-302b, a novel regulator of EphA2, as one of the most significantly downregulated microRNA (miRNA) in GC tissues. Overexpression of miR-302b impaired GC cell migratory and invasive properties robustly and suppressed cell proliferation by arresting cells at G0-G1 phase in vitro. miR-302b exhibited anti-tumor activity by reversing EphA2 regulation, which relayed a signaling transduction cascade that attenuated the functions of N-cadherin, β-catenin, and Snail (markers of Wnt/β-catenin and epithelial-mesenchymal transition, EMT). This modulation of EphA2 also had distinct effects on cell proliferation and migration in GC in vivo. miR-302b serves as a critical suppressor of GC cell tumorigenesis and metastasis by targeting the EphA2/Wnt/β-catenin/EMT pathway.

Activation of the Saccharomyces cerevisiae filamentation/invasion pathway by osmotic stress in high-osmolarity glycogen pathway mutants

NASA Technical Reports Server (NTRS)

Davenport, K. D.; Williams, K. E.; Ullmann, B. D.; Gustin, M. C.; McIntire, L. V. (Principal Investigator)

1999-01-01

Mitogen-activated protein kinase (MAPK) cascades are frequently used signal transduction mechanisms in eukaryotes. Of the five MAPK cascades in Saccharomyces cerevisiae, the high-osmolarity glycerol response (HOG) pathway functions to sense and respond to hypertonic stress. We utilized a partial loss-of-function mutant in the HOG pathway, pbs2-3, in a high-copy suppressor screen to identify proteins that modulate growth on high-osmolarity media. Three high-copy suppressors of pbs2-3 osmosensitivity were identified: MSG5, CAK1, and TRX1. Msg5p is a dual-specificity phosphatase that was previously demonstrated to dephosphorylate MAPKs in yeast. Deletions of the putative MAPK targets of Msg5p revealed that kss1delta could suppress the osmosensitivity of pbs2-3. Kss1p is phosphorylated in response to hyperosmotic shock in a pbs2-3 strain, but not in a wild-type strain nor in a pbs2-3 strain overexpressing MSG5. Both TEC1 and FRE::lacZ expressions are activated in strains lacking a functional HOG pathway during osmotic stress in a filamentation/invasion-pathway-dependent manner. Additionally, the cellular projections formed by a pbs2-3 mutant on high osmolarity are absent in strains lacking KSS1 or STE7. These data suggest that the loss of filamentation/invasion pathway repression contributes to the HOG mutant phenotype.

Activation of the Saccharomyces cerevisiae filamentation/invasion pathway by osmotic stress in high-osmolarity glycogen pathway mutants.

PubMed Central

Davenport, K D; Williams, K E; Ullmann, B D; Gustin, M C

1999-01-01

Mitogen-activated protein kinase (MAPK) cascades are frequently used signal transduction mechanisms in eukaryotes. Of the five MAPK cascades in Saccharomyces cerevisiae, the high-osmolarity glycerol response (HOG) pathway functions to sense and respond to hypertonic stress. We utilized a partial loss-of-function mutant in the HOG pathway, pbs2-3, in a high-copy suppressor screen to identify proteins that modulate growth on high-osmolarity media. Three high-copy suppressors of pbs2-3 osmosensitivity were identified: MSG5, CAK1, and TRX1. Msg5p is a dual-specificity phosphatase that was previously demonstrated to dephosphorylate MAPKs in yeast. Deletions of the putative MAPK targets of Msg5p revealed that kss1delta could suppress the osmosensitivity of pbs2-3. Kss1p is phosphorylated in response to hyperosmotic shock in a pbs2-3 strain, but not in a wild-type strain nor in a pbs2-3 strain overexpressing MSG5. Both TEC1 and FRE::lacZ expressions are activated in strains lacking a functional HOG pathway during osmotic stress in a filamentation/invasion-pathway-dependent manner. Additionally, the cellular projections formed by a pbs2-3 mutant on high osmolarity are absent in strains lacking KSS1 or STE7. These data suggest that the loss of filamentation/invasion pathway repression contributes to the HOG mutant phenotype. PMID:10545444

Real-Time Single Molecule Visualization of SH2 Domain Membrane Recruitment in Growth Factor Stimulated Cells.

PubMed

Oh, Dongmyung

2017-01-01

In the last decade, single molecule tracking (SMT) techniques have emerged as a versatile tool for molecular cell biology research. This approach allows researchers to monitor the real-time behavior of individual molecules in living cells with nanometer and millisecond resolution. As a result, it is possible to visualize biological processes as they occur at a molecular level in real time. Here we describe a method for the real-time visualization of SH2 domain membrane recruitment from the cytoplasm to epidermal growth factor (EGF) induced phosphotyrosine sites on the EGF receptor. Further, we describe methods that utilize SMT data to define SH2 domain membrane dynamics parameters such as binding (τ), dissociation (k d ), and diffusion (D) rates. Together these methods may allow us to gain greater understanding of signal transduction dynamics and the molecular basis of disease-related aberrant pathways.

Development of DNAzyme-based PCR signal cascade amplification for visual detection of Listeria monocytogenes in food.

PubMed

Liu, Zhanmin; Yao, Chenhui; Yang, Cuiyun; Wang, Yanming; Wan, Sibao; Huang, Junyi

2018-05-16

Listeria monocytogenes is an important foodborne pathogen, and it can cause severe diseases. Rapid detection of L. monocytogenes is crucial to control this pathogen. A simple and robust strategy based on the cascade of PCR and G-quadruplex DNAzyme catalyzed reaction was used to detect L. monocytogenes. In the presence of hemin and the aptamer formed during PCR, the catalytic horseradish peroxidase-mimicking G-quadruplex DNAzymes allow the colorimetric responses of target DNA from L. monocytogenes. This assay can detect genomic DNA of L. monocytogenes specifically with as low as 50 pg/reaction with the naked eye. Through 20 pork samples assay, visual detection assay had the same results as conventional detection methods, and had a good performance. This is a powerful demonstration of the ability of G-quadruplex DNAzyme to be used for PCR-based assay with significant advantages of high sensitivity, low cost and simple manipulation over existing approaches and offers the opportunity for application in pathogen detection. Copyright © 2018 Elsevier Inc. All rights reserved.

Sensitivity to timing and order in human visual cortex.

PubMed

Singer, Jedediah M; Madsen, Joseph R; Anderson, William S; Kreiman, Gabriel

2015-03-01

Visual recognition takes a small fraction of a second and relies on the cascade of signals along the ventral visual stream. Given the rapid path through multiple processing steps between photoreceptors and higher visual areas, information must progress from stage to stage very quickly. This rapid progression of information suggests that fine temporal details of the neural response may be important to the brain's encoding of visual signals. We investigated how changes in the relative timing of incoming visual stimulation affect the representation of object information by recording intracranial field potentials along the human ventral visual stream while subjects recognized objects whose parts were presented with varying asynchrony. Visual responses along the ventral stream were sensitive to timing differences as small as 17 ms between parts. In particular, there was a strong dependency on the temporal order of stimulus presentation, even at short asynchronies. From these observations we infer that the neural representation of complex information in visual cortex can be modulated by rapid dynamics on scales of tens of milliseconds. Copyright © 2015 the American Physiological Society.

Mechanism of colour discrimination by a bacterial sensory rhodopsin

NASA Technical Reports Server (NTRS)

Spudich, J. L.; Bogomolni, R. A.

1984-01-01

A photosensitive protein resembling the visual pigments of invertebrates enables phototactic archaebacteria to distinguish color. This protein exists in two spectrally-distinct forms, one of which is a transient photoproduct of the other and each of which undergoes photochemical reactions controlling the cell's swimming behaviour. Activation of a single pigment molecule in the cell is sufficient to signal the flagellar motor. This signal-transduction mechanism makes evident a color-sensing capability inherent in the retinal/protein chromophore.

Bicarbonate Modulates Photoreceptor Guanylate Cyclase (ROS-GC) Catalytic Activity*

PubMed Central

Duda, Teresa; Wen, Xiao-Hong; Isayama, Tomoki; Sharma, Rameshwar K.; Makino, Clint L.

2015-01-01

By generating the second messenger cGMP in retinal rods and cones, ROS-GC plays a central role in visual transduction. Guanylate cyclase-activating proteins (GCAPs) link cGMP synthesis to the light-induced fall in [Ca2+]i to help set absolute sensitivity and assure prompt recovery of the response to light. The present report discloses a surprising feature of this system: ROS-GC is a sensor of bicarbonate. Recombinant ROS-GCs synthesized cGMP from GTP at faster rates in the presence of bicarbonate with an ED50 of 27 mm for ROS-GC1 and 39 mm for ROS-GC2. The effect required neither Ca2+ nor use of the GCAPs domains; however, stimulation of ROS-GC1 was more powerful in the presence of GCAP1 or GCAP2 at low [Ca2+]. When applied to retinal photoreceptors, bicarbonate enhanced the circulating current, decreased sensitivity to flashes, and accelerated flash response kinetics. Bicarbonate was effective when applied either to the outer or inner segment of red-sensitive cones. In contrast, bicarbonate exerted an effect when applied to the inner segment of rods but had little efficacy when applied to the outer segment. The findings define a new regulatory mechanism of the ROS-GC system that affects visual transduction and is likely to affect the course of retinal diseases caused by cGMP toxicity. PMID:25767116

Non-canonical Phototransduction Mediates Synchronization of the Drosophila melanogaster Circadian Clock and Retinal Light Responses.

PubMed

Ogueta, Maite; Hardie, Roger C; Stanewsky, Ralf

2018-06-04

The daily light-dark cycles represent a key signal for synchronizing circadian clocks. Both insects and mammals possess dedicated "circadian" photoreceptors but also utilize the visual system for clock resetting. In Drosophila, circadian clock resetting is achieved by the blue-light photoreceptor cryptochrome (CRY), which is expressed within subsets of the brain clock neurons. In addition, rhodopsin-expressing photoreceptor cells contribute to light synchronization. Light resets the molecular clock by CRY-dependent degradation of the clock protein Timeless (TIM), although in specific subsets of key circadian pacemaker neurons, including the small ventral lateral neurons (s-LNvs), TIM and Period (PER) oscillations can be synchronized by light independent of CRY and canonical visual Rhodopsin phototransduction. Here, we show that at least three of the seven Drosophila rhodopsins can utilize an alternative transduction mechanism involving the same α-subunit of the heterotrimeric G protein operating in canonical visual phototransduction (Gq). Surprisingly, in mutants lacking the canonical phospholipase C-β (PLC-β) encoded by the no receptor potential A (norpA) gene, we uncovered a novel transduction pathway using a different PLC-β encoded by the Plc21C gene. This novel pathway is important for behavioral clock resetting to semi-natural light-dark cycles and mediates light-dependent molecular synchronization within the s-LNv clock neurons. The same pathway appears to be responsible for norpA-independent light responses in the compound eye. We show that Rhodopsin 5 (Rh5) and Rh6, present in the R8 subset of retinal photoreceptor cells, drive both the long-term circadian and rapid light responses in the eye. Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.

Transduction between worlds: using virtual and mixed reality for earth and planetary science

NASA Astrophysics Data System (ADS)

Hedley, N.; Lochhead, I.; Aagesen, S.; Lonergan, C. D.; Benoy, N.

2017-12-01

Virtual reality (VR) and augmented reality (AR) have the potential to transform the way we visualize multidimensional geospatial datasets in support of geoscience research, exploration and analysis. The beauty of virtual environments is that they can be built at any scale, users can view them at many levels of abstraction, move through them in unconventional ways, and experience spatial phenomena as if they had superpowers. Similarly, augmented reality allows you to bring the power of virtual 3D data visualizations into everyday spaces. Spliced together, these interface technologies hold incredible potential to support 21st-century geoscience. In my ongoing research, my team and I have made significant advances to connect data and virtual simulations with real geographic spaces, using virtual environments, geospatial augmented reality and mixed reality. These research efforts have yielded new capabilities to connect users with spatial data and phenomena. These innovations include: geospatial x-ray vision; flexible mixed reality; augmented 3D GIS; situated augmented reality 3D simulations of tsunamis and other phenomena interacting with real geomorphology; augmented visual analytics; and immersive GIS. These new modalities redefine the ways in which we can connect digital spaces of spatial analysis, simulation and geovisualization, with geographic spaces of data collection, fieldwork, interpretation and communication. In a way, we are talking about transduction between real and virtual worlds. Taking a mixed reality approach to this, we can link real and virtual worlds. This paper presents a selection of our 3D geovisual interface projects in terrestrial, coastal, underwater and other environments. Using rigorous applied geoscience data, analyses and simulations, our research aims to transform the novelty of virtual and augmented reality interface technologies into game-changing mixed reality geoscience.

Real-time monitoring of pH-dependent intracellular trafficking of ovarian cancer G protein-coupled receptor 1 in living leukocytes.

PubMed

Tan, Modong; Yamaguchi, Satoshi; Nakamura, Motonao; Nagamune, Teruyuki

2018-04-11

G-protein coupled receptors (GPCRs) are involved in many diseases and important biological phenomena; elucidating the mechanisms underlying regulation of their signal transduction potentially provides both novel targets for drug discovery and insight into living systems. A proton-sensing GPCR, ovarian cancer G protein-coupled receptor 1 (OGR1), has been reported to be related to acidosis and diseases that cause tissue acidification, but the mechanism of proton-induced activation of OGR1-mediated signal transduction in acidic conditions remains unclear. Here, pH-dependent intracellular trafficking of OGR1 was visualized in living leukocytes by a real-time fluorescence microscopic method based on sortase A-mediated pulse labeling of OGR1. OGR1 labeled on the cell surface with a small fluorescent dye was clearly observed to remain in the plasma membrane during incubation in mildly acidic medium (pH 6.6) and to be internalized to the intracellular compartments on changing the medium to slightly basic pH (7.7). Quantitative single-cell image analysis showed that most of the internalized OGR1s were then recycled to the plasma membrane for signal transduction if the extracellular pH was returned to the mildly acidic state. However, in a minor population of cells (40%), the internalized OGR1s were retained in endosomes or transported to lysosomes and degraded, leading to low efficiency of their recycling to the plasma membrane. Thus, the present live-cell monitoring strongly suggests that the signal transduction activity of OGR1 is regulated by pH-dependent internalization and recycling to the plasma membrane. Copyright © 2018 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Nfatc1 Is a Functional Transcriptional Factor Mediating Nell-1-Induced Runx3 Upregulation in Chondrocytes.

PubMed

Li, Chenshuang; Zheng, Zhong; Zhang, Xinli; Asatrian, Greg; Chen, Eric; Song, Richard; Culiat, Cymbeline; Ting, Kang; Soo, Chia

2018-01-06

Neural EGFL like 1 (Nell-1) is essential for chondrogenic differentiation, maturation, and regeneration. Our previous studies have demonstrated that Nell-1's pro-chondrogenic activities are predominantly reliant upon runt-related transcription factor 3 (Runx3)-mediated Indian hedgehog (Ihh) signaling. Here, we identify the nuclear factor of activated T-cells 1 (Nfatc1) as the key transcriptional factor mediating the Nell-1 → Runx3 signal transduction in chondrocytes. Using chromatin immunoprecipitation assay, we were able to determine that Nfatc1 binds to the -833--810 region of the Runx3 -promoter in response to Nell-1 treatment. By revealing the Nell-1 → Nfatc1 → Runx3 → Ihh cascade, we demonstrate the involvement of Nfatc1, a nuclear factor of activated T-cells, in chondrogenesis, while providing innovative insights into developing a novel therapeutic strategy for cartilage regeneration and other chondrogenesis-related conditions.

[MAP kinases--molecular transistors in animals and plants].

PubMed

Petersen, Morten; Brodersen, Peter; Mundy, John

2002-06-10

The survival of multicellular organisms depends on the ability of their cells to communicate with each other and to respond to environmental changes. A goal of modern biology is to uncover the processes by which these cellular signals are transduced. Recent studies have shown that MAP-kinases (MAPKs) are important constituents of such signal transduction pathways. MAPKs function as modules in phosphorelay cascades to activate or repress the activity of downstream target proteins. For example, recent research with knockout mice has shown that mammalian MAPKs are involved in the control of neuronal apoptosis and the activation of immune responses. These mammalian MAPKs exert their control by both promoting and inhibiting specific processes. Surprisingly, plants also use MAPKs to control their immune responses, and plant MAPKs also seem to play dual roles as positive and negative regulators. Such mechanistic similarities provide the basis for fruitful conceptual exchange between molecular research on animals and plants.

The canonical Wnt signaling pathway in autism.

PubMed

Zhang, Yinghua; Yuan, Xiangshan; Wang, Zhongping; Li, Ruixi

2014-01-01

Mounting attention is being focused on the canonical Wnt signaling pathway which has been implicated in the pathogenesis of autism in some our and other recent studies. The canonical Wnt pathway is involved in cell proliferation, differentiation and migration, especially during nervous system development. Given its various functions, dysfunction of the canonical Wnt pathway may exert adverse effects on neurodevelopment and therefore leads to the pathogenesis of autism. Here, we review human and animal studies that implicate the canonical Wnt signal transduction pathway in the pathogenesis of autism. We also describe the crosstalk between the canonical Wnt pathway and the Notch signaling pathway in several types of autism spectrum disorders, including Asperger syndrome and Fragile X. Further research on the crosstalk between the canonical Wnt signaling pathway and other signaling cascades in autism may be an efficient avenue to understand the etiology of autism and ultimately lead to alternative medications for autism-like phenotypes.

Protein tyrosine phosphatase 1B as a target for the treatment of impaired glucose tolerance and type II diabetes.

PubMed

Liu, Gang; Trevillyan, James M

2002-11-01

Protein tyrosine phosphatase 1B (PTP1B) is a negative regulator of the insulin signal transduction cascade, initiated when insulin binds to the insulin receptor. PTP1B-deficient mice are more sensitive to insulin, and have improved glycemic control and resistance to diet-induced obesity than wild-type control mice. Diabetic mice treated with PTP1B antisense oligonucleotides intraperitoneally have lower PTP1B protein levels in liver and fat, reduced plasma insulin, blood glucose and hemoglobin A1c (HbA1c) levels. These studies validate PTP1B as a promising drug discovery target for the treatment of insulin resistance, diabetes and obesity. Herein we review the recent advances in the structure-based design of potent and selective small molecule inhibitors of PTP1B, and discuss th e challenge of developing compounds with improved cell permeability and bioavailability.

Crystallization and preliminary X-ray crystallographic analysis of the extracellular domain of LePRK2 from Lycopersicon esculentum.

PubMed

Xu, Anbi; Huang, Laiqiang

2014-02-01

The tomato (Lycopersicon esculentum) pollen-specific receptor kinase 2 (LePRK2) is a member of the large receptor-like kinase (RLK) family and is expressed specifically in mature pollen and pollen tubes in L. esculentum. Like other RLKs, LePRK2 contains a characteristic N-terminal leucine-rich repeat (LRR) extracellular domain, the primary function of which is in protein-protein interactions. The LePRK2 LRR is likely to bind candidate ligands from the external environment, leading to a signal transduction cascade required for successful pollination. LePRK2-LRR was purified using an insect-cell secretion expression system and was crystallized using the vapour-diffusion method. The crystals diffracted to a resolution of 2.50 Å and belonged to space group I4(1)22, with unit-cell parameters a = b = 93.94, c = 134.44 Å and one molecule per asymmetric unit.

The current state of GPCR-based drug discovery to treat metabolic disease.

PubMed

Sloop, Kyle W; Emmerson, Paul J; Statnick, Michael A; Willard, Francis S

2018-02-02

One approach of modern drug discovery is to identify agents that enhance or diminish signal transduction cascades in various cell types and tissues by modulating the activity of GPCRs. This strategy has resulted in the development of new medicines to treat many conditions, including cardiovascular disease, psychiatric disorders, HIV/AIDS, certain forms of cancer and Type 2 diabetes mellitus (T2DM). These successes justify further pursuit of GPCRs as disease targets and provide key learning that should help guide identifying future therapeutic agents. This report reviews the current landscape of GPCR drug discovery with emphasis on efforts aimed at developing new molecules for treating T2DM and obesity. We analyse historical efforts to generate GPCR-based drugs to treat metabolic disease in terms of causal factors leading to success and failure in this endeavour. © 2018 The British Pharmacological Society.

Nickel impact on human health: An intrinsic disorder perspective.

PubMed

Zambelli, Barbara; Uversky, Vladimir N; Ciurli, Stefano

2016-12-01

The interplay of the presence of nickel and protein disorder in processes affecting human health is the focus of the present review. Many systems involving nickel as either a cofactor or as a toxic contaminant are characterized by large disorder. The role of nickel in the biochemistry of bacterial enzymes is discussed here, covering both the beneficial effects of nickel in the human microbiota as well as the role of nickel-depending bacteria in human pathogenesis. In addition, the hazardous health effects caused by nickel exposure to humans, namely nickel-induced carcinogenesis and allergy, are triggered by non-specific interactions of nickel with macromolecules and formation of reactive compounds that mediate cellular damage. Cellular response to nickel is also related to signal transduction cascades. This review thus highlights the most promising systems for future studies aimed at decreasing the adverse effects of nickel on human health. Copyright © 2016 Elsevier B.V. All rights reserved.

How a mycoparasite employs g-protein signaling: using the example of trichoderma.

PubMed

Omann, Markus; Zeilinger, Susanne

2010-01-01

Mycoparasitic Trichoderma spp. act as potent biocontrol agents against a number of plant pathogenic fungi, whereupon the mycoparasitic attack includes host recognition followed by infection structure formation and secretion of lytic enzymes and antifungal metabolites leading to the host's death. Host-derived signals are suggested to be recognized by receptors located on the mycoparasite's cell surface eliciting an internal signal transduction cascade which results in the transcription of mycoparasitism-relevant genes. Heterotrimeric G proteins of fungi transmit signals originating from G-protein-coupled receptors mainly to the cAMP and the MAP kinase pathways resulting in regulation of downstream effectors. Components of the G-protein signaling machinery such as Gα subunits and G-protein-coupled receptors were recently shown to play crucial roles in Trichoderma mycoparasitism as they govern processes such as the production of extracellular cell wall lytic enzymes, the secretion of antifungal metabolites, and the formation of infection structures.

Root gravitropism

NASA Technical Reports Server (NTRS)

Masson, P. H.

1995-01-01

When a plant root is reoriented within the gravity field, it responds by initiating a curvature which eventually results in vertical growth. Gravity sensing occurs primarily in the root tip. It may involve amyloplast sedimentation in the columella cells of the root cap, or the detection of forces exerted by the mass of the protoplast on opposite sides of its cell wall. Gravisensing activates a signal transduction cascade which results in the asymmetric redistribution of auxin and apoplastic Ca2+ across the root tip, with accumulation at the bottom side. The resulting lateral asymmetry in Ca2+ and auxin concentration is probably transmitted to the elongation zone where differential cellular elongation occurs until the tip resumes vertical growth. The Cholodny-Went theory proposes that gravity-induced auxin redistribution across a gravistimulated plant organ is responsible for the gravitropic response. However, recent data indicate that the gravity-induced reorientation is more complex, involving both auxin gradient-dependent and auxin gradient-independent events.

Mid-Ir Cavity Ring-Down Spectrometer for Biological Trace Nitric Oxide Detection

NASA Astrophysics Data System (ADS)

Kan, Vincent; Ragab, Ahemd; Stsiapura, Vitali; Lehmann, Kevin K.; Gaston, Benjamin M.

2011-06-01

S-nitrosothiols have received much attention in biochemistry and medicine as donors of nitrosonium ion (NO^+) and nitric oxide (NO) - physiologically active molecules involved in vasodilation and signal transduction. Determination of S-nitrosothiols content in cells and tissues is of great importance for fundamental research and medical applications. We will report on our ongoing development of a instrument to measure trace levels of nitric oxide gas (NO), released from S-nitrosothiols after exposure to UV light (340 nm) or reaction with L-Cysteine+CuCl mixture. The instrument uses the method of cavity ring-down spectroscopy, probing rotationally resolved lines in the vibrational fundamental transition near 5.2 μm. The laser source is a continuous-wave, room temperature external cavity quantum cascade laser. An acousto-optic modulator is used to abruptly turn off the optical power incident on the cavity when the laser and cavity pass through resonance.

The protein kinase Pelle mediates feedback regulation in the Drosophila Toll signaling pathway.

PubMed

Towb, P; Bergmann, A; Wasserman, S A

2001-12-01

Dorsoventral polarity in the Drosophila embryo is established through a signal transduction cascade triggered in ventral and ventrolateral regions. Activation of a transmembrane receptor, Toll, leads to localized recruitment of the adaptor protein Tube and protein kinase Pelle. Signaling through these components directs degradation of the IkappaB-like inhibitor Cactus and nuclear translocation of the Rel protein Dorsal. Here we show through confocal immunofluorescence microscopy that Pelle functions to downregulate the signal-dependent relocalization of Tube. Inactivation of the Pelle kinase domain, or elimination of the Tube-Pelle interaction, dramatically increases Tube recruitment to the ventral plasma membrane in regions of active signaling. We also characterize a large collection of pelle alleles, identifying the molecular lesions in these alleles and their effects on Pelle autophosphorylation, Tube phosphorylation and Tube relocalization. Our results point to a mechanism operating to modulate the domain or duration of signaling downstream from Tube and Pelle.

Micro-Plasticity of Genomes As Illustrated by the Evolution of Glutathione Transferases in 12 Drosophila Species

PubMed Central

Saisawang, Chonticha; Ketterman, Albert J.

2014-01-01

Glutathione transferases (GST) are an ancient superfamily comprising a large number of paralogous proteins in a single organism. This multiplicity of GSTs has allowed the copies to diverge for neofunctionalization with proposed roles ranging from detoxication and oxidative stress response to involvement in signal transduction cascades. We performed a comparative genomic analysis using FlyBase annotations and Drosophila melanogaster GST sequences as templates to further annotate the GST orthologs in the 12 Drosophila sequenced genomes. We found that GST genes in the Drosophila subgenera have undergone repeated local duplications followed by transposition, inversion, and micro-rearrangements of these copies. The colinearity and orientations of the orthologous GST genes appear to be unique in many of the species which suggests that genomic rearrangement events have occurred multiple times during speciation. The high micro-plasticity of the genomes appears to have a functional contribution utilized for evolution of this gene family. PMID:25310450

Mapping the local protein interactome of the NuA3 histone acetyltransferase

PubMed Central

Smart, Sherri K; Mackintosh, Samuel G; Edmondson, Ricky D; Taverna, Sean D; Tackett, Alan J

2009-01-01

Protein–protein interactions modulate cellular functions ranging from the activity of enzymes to signal transduction cascades. A technology termed transient isotopic differentiation of interactions as random or targeted (transient I-DIRT) is described for the identification of stable and transient protein–protein interactions in vivo. The procedure combines mild in vivo chemical cross-linking and non-stringent affinity purification to isolate low abundance chromatin-associated protein complexes. Using isotopic labeling and mass spectrometric readout, purified proteins are categorized with respect to the protein ‘bait’ as stable, transient, or contaminant. Here we characterize the local interactome of the chromatin-associated NuA3 histone lysine-acetyltransferase protein complex. We describe transient associations with the yFACT nucleosome assembly complex, RSC chromatin remodeling complex and a nucleosome assembly protein. These novel, physical associations with yFACT, RSC, and Nap1 provide insight into the mechanism of NuA3-associated transcription and chromatin regulation. PMID:19621382

How a Mycoparasite Employs G-Protein Signaling: Using the Example of Trichoderma

PubMed Central

Omann, Markus; Zeilinger, Susanne

2010-01-01

Mycoparasitic Trichoderma spp. act as potent biocontrol agents against a number of plant pathogenic fungi, whereupon the mycoparasitic attack includes host recognition followed by infection structure formation and secretion of lytic enzymes and antifungal metabolites leading to the host's death. Host-derived signals are suggested to be recognized by receptors located on the mycoparasite's cell surface eliciting an internal signal transduction cascade which results in the transcription of mycoparasitism-relevant genes. Heterotrimeric G proteins of fungi transmit signals originating from G-protein-coupled receptors mainly to the cAMP and the MAP kinase pathways resulting in regulation of downstream effectors. Components of the G-protein signaling machinery such as Gα subunits and G-protein-coupled receptors were recently shown to play crucial roles in Trichoderma mycoparasitism as they govern processes such as the production of extracellular cell wall lytic enzymes, the secretion of antifungal metabolites, and the formation of infection structures. PMID:21637351

The power and promise of "rewiring" the mitogen-activated protein kinase network in prostate cancer therapeutics.

PubMed

Papatsoris, Athanasios G; Karamouzis, Michalis V; Papavassiliou, Athanasios G

2007-03-01

Prostate cancer is the most frequently diagnosed cancer among men and the second leading cause of male cancer deaths. Initially, tumor growth is androgen dependent and thus responsive to pharmacologic androgen deprivation, but there is a high rate of treatment failure because the disease evolves in an androgen-independent state. Growing evidence suggests that the Ras/mitogen-activated protein kinase (MAPK) signaling cascade represents a pivotal molecular circuitry participating directly or indirectly in prostate cancer evolution. The crucial role of the protein elements comprising this complex signal transduction network makes them potential targets for pharmacologic interference. Here, we will delineate the current knowledge regarding the involvement of the Ras/MAPK pathway in prostate carcinogenesis, spotlight ongoing research concerning the development of novel targeted agents such as the Ras/MAPK inhibitors in prostate cancer, and discuss the future perspectives of their therapeutic efficacy.

Age-related changes in the response of intestinal cells to 1α,25(OH)2-vitamin D3.

PubMed

Gonzalez Pardo, Verónica; Russo de Boland, Ana

2013-01-01

The hormonally active form of vitamin D(3), 1α,25(OH)(2)-vitamin D(3), acts in intestine, its major target tissue, where its actions are of regulatory and developmental importance: regulation of intracellular calcium through modulation of second messengers and activation of mitogenic cascades leading to cell proliferation. Several causes have been postulated to modify the hormone response in intestinal cells with ageing, among them, alterations of vitamin D receptor (VDR) levels and binding sites, reduced expression of G-proteins and hormone signal transduction changes. The current review summarizes the actual knowledge regarding the molecular and biochemical basis of age-impaired 1α,25(OH)(2)-vitamin D(3) receptor-mediated signaling in intestinal cells. A fundamental understanding why the hormone functions are impaired with age will enhance our knowledge of its importance in intestinal cell physiology. Copyright © 2012 Elsevier B.V. All rights reserved.

Mechanisms of acetylcholine- and bradykinin-induced preconditioning.

PubMed

Critz, Stuart D; Cohen, Michael V; Downey, James M

2005-01-01

Acetylcholine (ACh) and bradykinin (BK) are potent pharmacological agents which mimic ischemic preconditioning (IPC) enabling hearts to resist infarction during a subsequent period of ischemia. The cardioprotective pathways activated by BK but not ACh may also protect when activated at reperfusion. ACh and BK stimulate Gi/o-linked receptors and ultimately mediate protection by opening mitochondrial ATP-sensitive potassium channels with the generation of reactive oxygen species that act as second messengers to activate protein kinase C (PKC). There appear to be key differences, however, in the pathways prior to potassium channel opening for these two receptors. This review aims to summarize what is currently known about pharmacological preconditioning by ACh and BK with an emphasis on differences that are seen in the signal transduction cascades. Understanding the cellular basis of protection by ACh and BK is a critical step towards developing pharmacological agents that will prevent infarction during ischemia resulting from coronary occlusion or heart attack.

Ca(2+) signaling mechanisms in bovine adrenal chromaffin cells.

PubMed

Weiss, Jamie L

2012-01-01

Calcium (Ca(2+)) is a crucial intracellular messenger in physiological aspects of cell signaling. Adrenal chromaffin cells are the secretory cells from the adrenal gland medulla that secrete catecholamines, which include epinephrine and norepinephrine important in the 'fight or flight' response. Bovine adrenal chromaffin cells have long been used as an important model for secretion -(exocytosis) not only due to their importance in the short-term stress response, but also as a neuroendocrine model of neurotransmtter release, as they have all the same exocytotic proteins as neurons but are easier to prepare, culture and use in functional assays. The components of the Ca(2+) signal transduction cascade and it role in secretion has been extensively characterized in bovine adrenal chromaffin cells. The Ca(2+) sources, signaling molecules and how this relates to the short-term stress response are reviewed in this book chapter in an endeavor to generally -overview these mechanisms in a concise and uncomplicated manner.

Discovery of a Dual PRMT5-PRMT7 Inhibitor.

PubMed

Smil, David; Eram, Mohammad S; Li, Fengling; Kennedy, Steven; Szewczyk, Magdalena M; Brown, Peter J; Barsyte-Lovejoy, Dalia; Arrowsmith, Cheryl H; Vedadi, Masoud; Schapira, Matthieu

2015-04-09

The protein arginine methyltransferases PRMT7 and PRMT5, respectively, monomethylate and symmetrically dimethylate arginine side-chains of proteins involved in diverse cellular mechanisms, including chromatin-mediated control of gene transcription, splicing, and the RAS to ERK transduction cascade. It is believed that PRMT5 and PRMT7 act in conjunction to methylate their substrates, and genetic deletions support the notion that these enzymes derepress cell proliferation and migration in cancer. Using available structures of PRMT5, we designed DS-437, a PRMT5 inhibitor with an IC50 value of 6 μM against both PRMT5 and PRMT7 that is inactive against 29 other human protein-, DNA-, and RNA-methyltransferases and inhibits symmetrical dimethylation of PRMT5 substrates in cells. This compound behaves as a cofactor competitor and represents a valid scaffold to interrogate the potential of the PRMT5-PRMT7 axis as a target for therapy.

Discovery of a Dual PRMT5–PRMT7 Inhibitor

PubMed Central

2015-01-01

The protein arginine methyltransferases PRMT7 and PRMT5, respectively, monomethylate and symmetrically dimethylate arginine side-chains of proteins involved in diverse cellular mechanisms, including chromatin-mediated control of gene transcription, splicing, and the RAS to ERK transduction cascade. It is believed that PRMT5 and PRMT7 act in conjunction to methylate their substrates, and genetic deletions support the notion that these enzymes derepress cell proliferation and migration in cancer. Using available structures of PRMT5, we designed DS-437, a PRMT5 inhibitor with an IC50 value of 6 μM against both PRMT5 and PRMT7 that is inactive against 29 other human protein-, DNA-, and RNA-methyltransferases and inhibits symmetrical dimethylation of PRMT5 substrates in cells. This compound behaves as a cofactor competitor and represents a valid scaffold to interrogate the potential of the PRMT5–PRMT7 axis as a target for therapy. PMID:25893041

Palytoxins and cytoskeleton: An overview.

PubMed

Louzao, M Carmen; Ares, Isabel R; Cagide, Eva; Espiña, Begoña; Vilariño, Natalia; Alfonso, Amparo; Vieytes, Mercedes R; Botana, Luis M

2011-03-01

Cytoskeleton is a dynamic structure essential for a wide variety of normal cellular processes, including the maintenance of cell shape and morphology, volume regulation, membrane dynamics and signal transduction. Cytoskeleton is organized into microtubules, actin meshwork and intermediate filaments. Actin has been identified as a major target for destruction during apoptosis and is also important under pathological conditions such as cancers. Several natural compounds actively modulate actin organization by specific signaling cascades being useful tools to study cytoskeleton dynamics. Palytoxin is a large bioactive compound, first isolated from zoanthids, with a complex structure and different analogs such as ostreocin-D or ovatoxin-a. This toxin has been identified as a potent tumor promoter and cytotoxic molecule, which leads to actin filament distortion and triggers cell death or apoptosis. In this review we report the findings on the involvement of palytoxin and analogues modulating the actin cytoskeleton within different cellular models. Copyright © 2010 Elsevier Ltd. All rights reserved.

Healthy clocks, healthy body, healthy mind.

PubMed

Reddy, Akhilesh B; O'Neill, John S

2010-01-01

Circadian rhythms permeate mammalian biology. They are manifested in the temporal organisation of behavioural, physiological, cellular and neuronal processes. Whereas it has been shown recently that these approximately 24-hour cycles are intrinsic to the cell and persist in vitro, internal synchrony in mammals is largely governed by the hypothalamic suprachiasmatic nuclei that facilitate anticipation of, and adaptation to, the solar cycle. Our timekeeping mechanism is deeply embedded in cell function and is modelled as a network of transcriptional and/or post-translational feedback loops. Concurrent with this, we are beginning to understand how this ancient timekeeper interacts with myriad cell systems, including signal transduction cascades and the cell cycle, and thus impacts on disease. An exemplary area where this knowledge is rapidly expanding and contributing to novel therapies is cancer, where the Period genes have been identified as tumour suppressors. In more complex disorders, where aetiology remains controversial, interactions with the clockwork are only now starting to be appreciated.

Custos controls β-catenin to regulate head development during vertebrate embryogenesis.

PubMed

Komiya, Yuko; Mandrekar, Noopur; Sato, Akira; Dawid, Igor B; Habas, Raymond

2014-09-09

Precise control of the canonical Wnt pathway is crucial in embryogenesis and all stages of life, and dysregulation of this pathway is implicated in many human diseases including cancers and birth defect disorders. A key aspect of canonical Wnt signaling is the cytoplasmic to nuclear translocation of β-catenin, a process that remains incompletely understood. Here we report the identification of a previously undescribed component of the canonical Wnt signaling pathway termed Custos, originally isolated as a Dishevelled-interacting protein. Custos contains casein kinase phosphorylation sites and nuclear localization sequences. In Xenopus, custos mRNA is expressed maternally and then widely throughout embryogenesis. Depletion or overexpression of Custos produced defective anterior head structures by inhibiting the formation of the Spemann-Mangold organizer. In addition, Custos expression blocked secondary axis induction by positive signaling components of the canonical Wnt pathway and inhibited β-catenin/TCF-dependent transcription. Custos binds to β-catenin in a Wnt responsive manner without affecting its stability, but rather modulates the cytoplasmic to nuclear translocation of β-catenin. This effect on nuclear import appears to be the mechanism by which Custos inhibits canonical Wnt signaling. The function of Custos is conserved as loss-of-function and gain-of-function studies in zebrafish also demonstrate a role for Custos in anterior head development. Our studies suggest a role for Custos in fine-tuning canonical Wnt signal transduction during embryogenesis, adding an additional layer of regulatory control in the Wnt-β-catenin signal transduction cascade.

An Arabidopsis mutation in translation elongation factor 2 causes superinduction of CBF/DREB1 transcription factor genes but blocks the induction of their downstream targets under low temperatures

PubMed Central

Guo, Yan; Xiong, Liming; Ishitani, Manabu; Zhu, Jian-Kang

2002-01-01

Low temperature regulates gene expression in bacteria, yeast, and animals as well as in plants. However, the signal transduction cascades mediating the low temperature responses are not well understood in any organism. To identify components in low temperature signaling genetically, we isolated Arabidopsis thaliana mutants in which cold-responsive genes are no longer induced by low temperatures. One of these mutations, los1–1, specifically blocks low temperature-induced transcription of cold-responsive genes. Surprisingly, cold-induced expression of the early response transcriptional activators, C-repeat/dehydration responsive element binding factors (CBF/DREB1s), is enhanced by the los1–1 mutation. The los1–1 mutation also reduces the capacity of plants to develop freezing tolerance but does not impair the vernalization response. Genetic analysis indicated that los1–1 is a recessive mutation in a single nuclear gene. The LOS1 gene encodes a translation elongation factor 2-like protein. Protein labeling studies show that new protein synthesis is blocked in los1–1 mutant plants specifically in the cold. These results reveal a critical role of new protein synthesis in the proper transduction of low temperature signals. Our results also suggest that cold-induced transcription of CBF/DREB1s is feedback inhibited by their gene products or by products of their downstream target genes. PMID:12032361

Nasal solitary chemoreceptor cell responses to bitter and trigeminal stimulants in vitro

PubMed Central

Gulbransen, Brian D; Clapp, Tod R; Kinnamon, Sue C; Finger, Thomas E

2009-01-01

Nasal trigeminal chemosensitivity in mice and rats is mediated in part by epithelial solitary chemoreceptor (chemosensory) cells (SCCs), but the exact role of these cells in chemoreception is unclear (Finger et al. 2003). Histological evidence suggests that SCCs express elements of the bitter taste transduction pathway including T2R (bitter taste) receptors, the G protein α-gustducin, PLCβ2, and TRPM5, leading to speculation that SCCs are the receptor cells that mediate trigeminal nerve responses to bitter taste receptor ligands. To test this hypothesis, we used calcium imaging to determine whether SCCs respond to classic bitter-tasting or trigeminal stimulants. SCCs from the anterior nasal cavity were isolated from transgenic mice in which green fluorescent protein (GFP) expression was driven by either TRPM5 or gustducin. Isolated cells were exposed to a variety of test stimuli to determine which substances caused an increase in intracellular Ca2+ ([Ca2+]i). GFP positive cells respond with increased [Ca2+]i to the bitter receptor ligand denatonium, and this response is blocked by the PLC inhibitor U73122. In addition GFP+ cells respond to the PLC activator 3M3FBS, the neuromodulators ATP and ACh, but only very rarely to other bitter-tasting or trigeminal stimuli. Our results demonstrate that TRPM5- and gustducin-expressing nasal SCCs respond to the T2R agonist, denatonium via a PLC-coupled transduction cascade typical of T2Rs in the taste system. PMID:18417634

Low-dose γ-radiation-induced oxidative stress response in mouse brain and gut: regulation by NFκB-MnSOD cross-signaling.

PubMed

Veeraraghavan, Jamunarani; Natarajan, Mohan; Herman, Terence S; Aravindan, Natarajan

2011-01-10

Radiation-induced amplification of reactive oxygen species (ROS) may be a sensing mechanism for activation of signaling cascades that influence cell fate. However, the regulated intrinsic mechanisms and targets of low-dose ionizing radiation (LDIR) are still unclear. Accordingly, we investigated the effects of LDIR on NFκB signal transduction and manganese superoxide dismutase (SOD2) activity in mice brain and gut. LDIR resulted in both dose-dependent and persistent NFκB activation in gut and brain. QPCR displayed a dose- and tissue-dependent differential modulation of 88 signaling molecules. With stringent criteria, a total of 15 (2cGy), 43 (10cGy) and 19 (50cGy) genes were found to be commonly upregulated between brain and gut. SOD2 immunostaining showed a LDIR-dose dependent increase. Consistent with the NFκB results, we observed a persistent increase in SOD2 activity after LDIR. Moreover, muting of LDIR-induced NFκB attenuated SOD2 transactivation and cellular localization. These results imply that exposure of healthy tissues to LDIR results in induced NFκB and SOD2 activity and transcriptional activation of NFκB-signal transduction/target molecules. More importantly, the results suggest that NFκB initiates a feedback response through transcriptional activation of SOD2 that may play a key role in the LDIR-induced oxidative stress response and may control the switch that directs cell fate. 2010 Elsevier B.V. All rights reserved.

Postnatal Experience Modulates Functional Properties of Mouse Olfactory Sensory Neurons

PubMed Central

He, Jiwei; Tian, Huikai; Lee, Anderson C.; Ma, Minghong

2012-01-01

Early experience considerably modulates the organization and function of all sensory systems. In the mammalian olfactory system, deprivation of the sensory inputs via neonatal, unilateral naris closure has been shown to induce structural, molecular, and functional changes from the olfactory epithelium to the olfactory bulb and cortex. However, it remains unknown how early experience shapes functional properties of individual olfactory sensory neurons (OSNs), the primary odor detectors in the nose. To address this question, we examined odorant response properties of mouse OSNs in both the closed and open nostril after four weeks of unilateral naris closure with age-matched untreated animals as control. Using patch-clamp technique on genetically-tagged OSNs with defined odorant receptors (ORs), we found that sensory deprivation increased the sensitivity of MOR23 neurons in the closed side while overexposure caused the opposite effect in the open side. We next analyzed the response properties including rise time, decay time, and adaptation induced by repeated stimulation in MOR23 and M71 neurons. Even though these two types of neurons showed distinct properties in dynamic range and response kinetics, sensory deprivation significantly slowed down the decay phase of odorant-induced transduction events in both types. Using western blotting and antibody staining, we confirmed upregulation of several signaling proteins in the closed side as compared with the open side. This study suggests that early experience modulates functional properties of OSNs, probably via modifying the signal transduction cascade. PMID:22703547

Spatial frequency supports the emergence of categorical representations in visual cortex during natural scene perception.

PubMed

Dima, Diana C; Perry, Gavin; Singh, Krish D

2018-06-11

In navigating our environment, we rapidly process and extract meaning from visual cues. However, the relationship between visual features and categorical representations in natural scene perception is still not well understood. Here, we used natural scene stimuli from different categories and filtered at different spatial frequencies to address this question in a passive viewing paradigm. Using representational similarity analysis (RSA) and cross-decoding of magnetoencephalography (MEG) data, we show that categorical representations emerge in human visual cortex at ∼180 ms and are linked to spatial frequency processing. Furthermore, dorsal and ventral stream areas reveal temporally and spatially overlapping representations of low and high-level layer activations extracted from a feedforward neural network. Our results suggest that neural patterns from extrastriate visual cortex switch from low-level to categorical representations within 200 ms, highlighting the rapid cascade of processing stages essential in human visual perception. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Ultra-Rapid serial visual presentation reveals dynamics of feedforward and feedback processes in the ventral visual pathway.

PubMed

Mohsenzadeh, Yalda; Qin, Sheng; Cichy, Radoslaw M; Pantazis, Dimitrios

2018-06-21

Human visual recognition activates a dense network of overlapping feedforward and recurrent neuronal processes, making it hard to disentangle processing in the feedforward from the feedback direction. Here, we used ultra-rapid serial visual presentation to suppress sustained activity that blurs the boundaries of processing steps, enabling us to resolve two distinct stages of processing with MEG multivariate pattern classification. The first processing stage was the rapid activation cascade of the bottom-up sweep, which terminated early as visual stimuli were presented at progressively faster rates. The second stage was the emergence of categorical information with peak latency that shifted later in time with progressively faster stimulus presentations, indexing time-consuming recurrent processing. Using MEG-fMRI fusion with representational similarity, we localized recurrent signals in early visual cortex. Together, our findings segregated an initial bottom-up sweep from subsequent feedback processing, and revealed the neural signature of increased recurrent processing demands for challenging viewing conditions. © 2018, Mohsenzadeh et al.

Comparison of visualized turbine endwall secondary flows and measured heat transfer patterns

NASA Technical Reports Server (NTRS)

Gaugler, R. E.; Russell, L. M.

1984-01-01

Various flow visualization techniques were used to define the seondary flows near the endwall in a large heat transfer data. A comparison of the visualized flow patterns and the measured Stanton number distribution was made for cases where the inlet Reynolds number and exit Mach number were matched. Flows were visualized by using neutrally buoyant helium-filled soap bubbles, by using smoke from oil soaked cigars, and by a few techniques using permanent marker pen ink dots and synthetic wintergreen oil. Details of the horseshoe vortex and secondary flows can be directly compared with heat transfer distribution. Near the cascade entrance there is an obvious correlation between the two sets of data, but well into the passage the effect of secondary flow is not as obvious. Previously announced in STAR as N83-14435

Use of Intraductal Adenovins Transduction to Assess the Mammary Tumorigenic Potential of a Constitutively Active Prolactin Receptor

DTIC Science & Technology

2000-10-01

interfere with the function of the mammary cells in which they are expressed. Transgenic technology has been used to evaluate the effects of an activated... wheat germ agglutinin; pfu, plaque-forming units; Cy3, a red fluorescent used for visualization of cell structures in the presence of GFP; DAPI, a...tumorigenesis in mice. The second objective has been achieved in part using transgenic mouse technology. We have begun exploration of the third objective. BODY

Logic Modeling in Quantitative Systems Pharmacology

PubMed Central

Traynard, Pauline; Tobalina, Luis; Eduati, Federica; Calzone, Laurence

2017-01-01

Here we present logic modeling as an approach to understand deregulation of signal transduction in disease and to characterize a drug's mode of action. We discuss how to build a logic model from the literature and experimental data and how to analyze the resulting model to obtain insights of relevance for systems pharmacology. Our workflow uses the free tools OmniPath (network reconstruction from the literature), CellNOpt (model fit to experimental data), MaBoSS (model analysis), and Cytoscape (visualization). PMID:28681552

The visual display of regulatory information and networks.

PubMed

Pirson, I; Fortemaison, N; Jacobs, C; Dremier, S; Dumont, J E; Maenhaut, C

2000-10-01

Cell regulation and signal transduction are becoming increasingly complex, with reports of new cross-signalling, feedback, and feedforward regulations between pathways and between the multiple isozymes discovered at each step of these pathways. However, this information, which requires pages of text for its description, can be summarized in very simple schemes, although there is no consensus on the drawing of such schemes. This article presents a simple set of rules that allows a lot of information to be inserted in easily understandable displays.

Evidence for a Limited-Cascading Account of Written Word Naming

ERIC Educational Resources Information Center

Bonin, Patrick; Roux, Sebastien; Barry, Christopher; Canell, Laura

2012-01-01

We address the issue of how information flows within the written word production system by examining written object-naming latencies. We report 4 experiments in which we manipulate variables assumed to have their primary impact at the level of object recognition (e.g., quality of visual presentation of pictured objects), at the level of semantic…

Comparison of 7.5-minute and 1-degree digital elevation models

NASA Technical Reports Server (NTRS)

Isaacson, Dennis L.; Ripple, William J.

1995-01-01

We compared two digital elevation models (DEM's) for the Echo Mountain SE quadrangle in the Cascade Mountains of Oregon. Comparisons were made between 7.5-minute (1:24,000-scale) and 1-degree (1:250,000-scale) images using the variables of elevation, slope aspect, and slope gradient. Both visual and statistical differences are presented.

Comparison of 7.5-minute and 1-degree digital elevation models

NASA Technical Reports Server (NTRS)

Isaacson, Dennis L.; Ripple, William J.

1990-01-01

Two digital elevation models are compared for the Echo Mountain SE quadrangle in the Cascade Mountains of Oregon. Comparisons were made between 7.5-minute (1:24,000-scale) and 1-degree (1:250,000-scale) images using the variables of elevation, slope aspect, and slope gradient. Both visual and statistical differences are presented.

The Materosion project, a sediment cascade modeling for torrential sediment transfers: final results and perspectives

NASA Astrophysics Data System (ADS)

Rudaz, Benjamin; Loye, Alexandre; Mazotti, Benoit; Bardou, Eric; Jaboyedoff, Michel

2013-04-01

The Materosion project, conducted between the swiss canton of Valais (CREALP) and University of Lausanne (CRET) aims at forecasting sediment transfer in alpine torrents using the sediment cascade concept. The study site is the high Anniviers valley, around the village of Zinal (Valais). The torrents are divided in homogeneous reaches, to and from which sediments are transported by debris flows and bedload transport events. The model runs simulations of 100 years, with a 1-month time step, each with a given a random meteorological event ranging from no activity up to high magnitude debris flows. These events are calibrated using local rain data and observed corresponding debris flow frequencies. The model is applied to ten torrent systems with variable geological context, watershed geometries and sediment supplies. Given the high number of possible event scenarios, 10'000 simulations per torrent are performed, giving a statistical distribution of cumulated volumes and an event size distribution. A way to visualize the complex results data is proposed, and a back-analysis of the internal sediment cascade dynamic is performed. The back-analysis shows that the results' distribution stabilize after ~5'000 simulations. The model results, especially the range of debris flow volumes are crucial to maintain mitigation measures such as retention dams, and give clues for future sediment cascade modeling.

Restoration of visual response in aged dystrophic RCS rats using AAV-mediated channelopsin-2 gene transfer.

PubMed

Tomita, Hiroshi; Sugano, Eriko; Yawo, Hiromu; Ishizuka, Toru; Isago, Hitomi; Narikawa, Satoko; Kügler, Sebastian; Tamai, Makoto

2007-08-01

To investigate whether the channelopsin-2 (Chop2) gene would restore visual responses in 10-month-old dystrophic Royal College of Surgeons (aged RCS; rdy/rdy) rats, the authors transferred the Chop2 gene into the retinal cells of aged RCS rats using the adenoassociated virus (AAV) vector. The N-terminal fragment (residues 1-315) of Chop2 was fused to a fluorescent protein, Venus, in frame at the end of the Chop2 coding fragment. The viral vector construct (AAV-Chop2V) for the expression of the Chop2V in the retina was made by subcloning into an adenoassociated virus vector, including the CAG promoter. To evaluate the expression profile of Chop2V in the retina, the rats were killed and the eyes were removed and fixed with 4% paraformaldehyde in 0.1 M phosphate-buffered saline. Retinal wholemount specimens and cryosections were made. Under anesthetized conditions, electrodes for the recording of visually evoked potentials (VEPs) were implanted onto the visual cortex in aged-RCS (rdy/rdy) rats. AAV-Chop2V vectors were then injected into the vitreous cavity of the left eyes. As a control, AAV-Venus vectors were applied to the right eyes. VEPs were evoked by the flash of a blue, white, or red light-emitting diode (LED) and were recorded from the visual cortex of the rats at various time points after the AAV vector injection. Chop2V fluorescence was predominantly observed in retinal ganglion cells (RGCs). Some fluorescence was observed in the inner nuclear layer and the inner plexiform layer neurites. A tendency of recovery was observed in the VEPs of aged RCS (rdy/rdy) rats after the AAV-Chop2V injection but not after the AAV-Venus injection. The visual response of AAV-Chop2V-injected aged RCS (rdy/rdy) rats was less sensitive to the blue LED flash than that of nondystrophic RCS (+/+) rats. The AAV-Chop2V-injected aged RCS (rdy/rdy) rats were insensitive to the red LED flash, which evoked a robust VEP in the RCS (+/+) rats. The visual response of aged RCS (rdy/rdy) rats was partially restored by transduction of the Chop2 gene through AAV into the inner retinal neurons, mainly RGCs. These results suggest that the transduction of Chop2 would provide a new strategy to treat some retinitis pigmentosa (RP) symptoms independent of their etiology.

Protein sorting, targeting and trafficking in photoreceptor cells

PubMed Central

Pearring, Jillian N.; Salinas, Raquel Y.; Baker, Sheila A.; Arshavsky, Vadim Y.

2013-01-01

Vision is the most fundamental of our senses initiated when photons are absorbed by the rod and cone photoreceptor neurons of the retina. At the distal end of each photoreceptor resides a light-sensing organelle, called the outer segment, which is a modified primary cilium highly enriched with proteins involved in visual signal transduction. At the proximal end, each photoreceptor has a synaptic terminal, which connects this cell to the downstream neurons for further processing of the visual information. Understanding the mechanisms involved in creating and maintaining functional compartmentalization of photoreceptor cells remains among the most fascinating topics in ocular cell biology. This review will discuss how photoreceptor compartmentalization is supported by protein sorting, targeting and trafficking, with an emphasis on the best-studied cases of outer segment-resident proteins. PMID:23562855

Bicarbonate Modulates Photoreceptor Guanylate Cyclase (ROS-GC) Catalytic Activity.

PubMed

Duda, Teresa; Wen, Xiao-Hong; Isayama, Tomoki; Sharma, Rameshwar K; Makino, Clint L

2015-04-24

By generating the second messenger cGMP in retinal rods and cones, ROS-GC plays a central role in visual transduction. Guanylate cyclase-activating proteins (GCAPs) link cGMP synthesis to the light-induced fall in [Ca(2+)]i to help set absolute sensitivity and assure prompt recovery of the response to light. The present report discloses a surprising feature of this system: ROS-GC is a sensor of bicarbonate. Recombinant ROS-GCs synthesized cGMP from GTP at faster rates in the presence of bicarbonate with an ED50 of 27 mM for ROS-GC1 and 39 mM for ROS-GC2. The effect required neither Ca(2+) nor use of the GCAPs domains; however, stimulation of ROS-GC1 was more powerful in the presence of GCAP1 or GCAP2 at low [Ca(2+)]. When applied to retinal photoreceptors, bicarbonate enhanced the circulating current, decreased sensitivity to flashes, and accelerated flash response kinetics. Bicarbonate was effective when applied either to the outer or inner segment of red-sensitive cones. In contrast, bicarbonate exerted an effect when applied to the inner segment of rods but had little efficacy when applied to the outer segment. The findings define a new regulatory mechanism of the ROS-GC system that affects visual transduction and is likely to affect the course of retinal diseases caused by cGMP toxicity. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Accelerating object detection via a visual-feature-directed search cascade: algorithm and field programmable gate array implementation

NASA Astrophysics Data System (ADS)

Kyrkou, Christos; Theocharides, Theocharis

2016-07-01

Object detection is a major step in several computer vision applications and a requirement for most smart camera systems. Recent advances in hardware acceleration for real-time object detection feature extensive use of reconfigurable hardware [field programmable gate arrays (FPGAs)], and relevant research has produced quite fascinating results, in both the accuracy of the detection algorithms as well as the performance in terms of frames per second (fps) for use in embedded smart camera systems. Detecting objects in images, however, is a daunting task and often involves hardware-inefficient steps, both in terms of the datapath design and in terms of input/output and memory access patterns. We present how a visual-feature-directed search cascade composed of motion detection, depth computation, and edge detection, can have a significant impact in reducing the data that needs to be examined by the classification engine for the presence of an object of interest. Experimental results on a Spartan 6 FPGA platform for face detection indicate data search reduction of up to 95%, which results in the system being able to process up to 50 1024×768 pixels images per second with a significantly reduced number of false positives.

Model identification of signal transduction networks from data using a state regulator problem.

PubMed

Gadkar, K G; Varner, J; Doyle, F J

2005-03-01

Advances in molecular biology provide an opportunity to develop detailed models of biological processes that can be used to obtain an integrated understanding of the system. However, development of useful models from the available knowledge of the system and experimental observations still remains a daunting task. In this work, a model identification strategy for complex biological networks is proposed. The approach includes a state regulator problem (SRP) that provides estimates of all the component concentrations and the reaction rates of the network using the available measurements. The full set of the estimates is utilised for model parameter identification for the network of known topology. An a priori model complexity test that indicates the feasibility of performance of the proposed algorithm is developed. Fisher information matrix (FIM) theory is used to address model identifiability issues. Two signalling pathway case studies, the caspase function in apoptosis and the MAP kinase cascade system, are considered. The MAP kinase cascade, with measurements restricted to protein complex concentrations, fails the a priori test and the SRP estimates are poor as expected. The apoptosis network structure used in this work has moderate complexity and is suitable for application of the proposed tools. Using a measurement set of seven protein concentrations, accurate estimates for all unknowns are obtained. Furthermore, the effects of measurement sampling frequency and quality of information in the measurement set on the performance of the identified model are described.

Timing Is Everything: Highly Specific and Transient Expression of a MAP Kinase Determines Auxin-Induced Leaf Venation Patterns in Arabidopsis

PubMed Central

Stanko, Vera; Giuliani, Concetta; Retzer, Katarzyna; Djamei, Armin; Wahl, Vanessa; Wurzinger, Bernhard; Wilson, Cathal; Heberle-Bors, Erwin; Teige, Markus; Kragler, Friedrich

2014-01-01

Mitogen-activated protein kinase (MAPK) cascades are universal signal transduction modules present in all eukaryotes. In plants, MAPK cascades were shown to regulate cell division, developmental processes, stress responses, and hormone pathways. The subgroup A of Arabidopsis MAPKs consists of AtMPK3, AtMPK6, and AtMPK10. AtMPK3 and AtMPK6 are activated by their upstream MAP kinase kinases (MKKs) AtMKK4 and AtMKK5 in response to biotic and abiotic stress. In addition, they were identified as key regulators of stomatal development and patterning. AtMPK10 has long been considered as a pseudo-gene, derived from a gene duplication of AtMPK6. Here we show that AtMPK10 is expressed highly but very transiently in seedlings and at sites of local auxin maxima leaves. MPK10 encodes a functional kinase and interacts with the upstream MAP kinase kinase (MAPKK) AtMKK2. mpk10 mutants are delayed in flowering in long-day conditions and in continuous light. Moreover, cotyledons of mpk10 and mkk2 mutants have reduced vein complexity, which can be reversed by inhibiting polar auxin transport (PAT). Auxin does not affect AtMPK10 expression while treatment with the PAT inhibitor HFCA extends the expression in leaves and reverses the mpk10 mutant phenotype. These results suggest that the AtMKK2–AtMPK10 MAPK module regulates venation complexity by altering PAT efficiency. PMID:25064848

The aortic ring model of angiogenesis: a quarter century of search and discovery

PubMed Central

Nicosia, R F

2009-01-01

The aortic ring model has become one of the most widely used methods to study angiogenesis and its mechanisms. Many factors have contributed to its popularity including reproducibility, cost effectiveness, ease of use and good correlation with in vivo studies. In this system aortic rings embedded in biomatrix gels and cultured under chemically defined conditions generate arborizing vascular outgrowths which can be stimulated or inhibited with angiogenic regulators. Originally based on the rat aorta, the aortic ring model was later adapted to the mouse for the evaluation of specific molecular alterations in genetically modified animals. Viral transduction of the aortic rings has enabled investigators to overexpress genes of interest in the aortic cultures. Experiments on angiogenic mechanisms have demonstrated that formation of neovessels in aortic cultures is regulated by macrophages, pericytes and fibroblasts through a complex molecular cascade involving growth factors, inflammatory cytokines, axonal guidance cues, extracellular matrix (ECM) molecules and matrix-degrading proteolytic enzymes. These studies have shown that endothelial sprouting can be effectively blocked by depleting the aortic explants of macrophages or by interfering with the angiogenic cascade at multiple levels including growth factor signalling, cell adhesion and proteolytic degradation of the ECM. In this paper, we review the literature in this field and retrace the journey from our first morphological descriptions of the aortic outgrowths to the latest breakthroughs in the cellular and molecular regulation of aortic vessel growth and regression. PMID:19725916

Mapping the CgrA regulon of Rhodospirillum centenum reveals a hierarchal network controlling Gram-negative cyst development.

PubMed

Dong, Qian; Fang, Mingxu; Roychowdhury, Sugata; Bauer, Carl E

2015-12-16

Several Gram-negative species undergo development leading to the formation of metabolically dormant desiccation resistant cysts. Recent analysis of cyst development has revealed that ~20 % of the Rhodospirillum centenum transcriptome undergo temporal changes in expression as cells transition from vegetative to cyst forms. It has also been established that one trigger for cyst formation is the synthesis of the signaling nucleotide 3', 5'- cyclic guanosine monophosphate (cGMP) that is sensed by a homolog of the catabolite repressor protein called CgrA. CgrA in the presence of cGMP initiate a cascade of gene expression leading to the development of cysts. In this study, we have used RNA-seq and chromatin immunoprecipitation (ChIP-Seq) techniques to define the CgrA-cGMP regulon. Our results indicate that disruption of CgrA leads to altered expression of 258 genes, 131 of which have been previously reported to be involved in cyst development. ChIP-seq analysis combined with transcriptome data also demonstrates that CgrA directly regulates the expression of numerous sigma factors and transcription factors several of which are known to be involved in cyst cell development. This analysis reveals the presence of CgrA binding sites upstream of many developmentally regulated genes including many transcription factors and signal transduction components. CgrA thus functions as master controller of the cyst development by initiating a hierarchal cascade of downstream transcription factors that induces temporal expression of encystment genes.

Artificial sRNAs activating the Gac/Rsm signal transduction pathway in Pseudomonas fluorescens.

PubMed

Valverde, Claudio

2009-04-01

In Pseudomonas fluorescens CHA0, the synthesis of antifungal compounds is post-transcriptionally activated by the Gac/Rsm cascade. The two-component system GacS/GacA promotes transcription of three small regulatory RNAs (i.e., sRNAs), RsmX, RsmY, and RsmZ, which remove the regulatory proteins RsmA and RsmE from the ribosome-binding sites of exoproduct-related mRNAs. The GacS/GacA-dependent accumulation of RsmX/Y/Z and formation of RsmX/Y/Z-RsmA/E complexes relieve mRNA translational repression. Other bacteria as E. coli and Vibrio spp. utilize similar sRNA-protein based systems to adjust mRNA translation (e.g., the E. coli Csr system for carbon storage, motility and biofilm regulation). The Rsm/Csr sRNAs are remarkably similar in that they contain several stem-loops with an invariant GGA trinucleotide exposed in the hairpin loop that would be the characteristic structural-sequence motifs relevant for sRNA activity and stability. Here it is shown that the dysfunctional Gac/Rsm cascade of P. fluorescens DeltarsmXYZ mutants could be restored by appropriate transcription levels of artificial genes encoding RNAs with unrelated primary sequence but with two or more hairpins displaying the RsmA/E binding motifs. The results support the hypothesis that the molecular mimicry of Rsm/Csr sRNAs is based on proper secondary structures that expose critical binding motifs irrespective of their overall sequence.

Modulating inflammation through the negative regulation of NF-κB signaling.

PubMed

Rothschild, Daniel E; McDaniel, Dylan K; Ringel-Scaia, Veronica M; Allen, Irving C

2018-02-01

Immune system activation is essential to thwart the invasion of pathogens and respond appropriately to tissue damage. However, uncontrolled inflammation can result in extensive collateral damage underlying a diverse range of auto-inflammatory, hyper-inflammatory, and neoplastic diseases. The NF-κB signaling pathway lies at the heart of the immune system and functions as a master regulator of gene transcription. Thus, this signaling cascade is heavily targeted by mechanisms designed to attenuate overzealous inflammation and promote resolution. Mechanisms associated with the negative regulation of NF-κB signaling are currently under intense investigation and have yet to be fully elucidated. Here, we provide an overview of mechanisms that negatively regulate NF-κB signaling through either attenuation of signal transduction, inhibition of posttranscriptional signaling, or interference with posttranslational modifications of key pathway components. While the regulators discussed for each group are far from comprehensive, they exemplify common mechanistic approaches that inhibit this critical biochemical signaling cascade. Despite their diversity, a commonality among these regulators is their selection of specific targets at key inflection points in the pathway, such as TNF-receptor-associated factor family members or essential kinases. A better understanding of these negative regulatory mechanisms will be essential to gain greater insight related to the maintenance of immune system homeostasis and inflammation resolution. These processes are vital elements of disease pathology and have important implications for targeted therapeutic strategies. ©2018 Society for Leukocyte Biology.

Short temporal asynchrony disrupts visual object recognition

PubMed Central

Singer, Jedediah M.; Kreiman, Gabriel

2014-01-01

Humans can recognize objects and scenes in a small fraction of a second. The cascade of signals underlying rapid recognition might be disrupted by temporally jittering different parts of complex objects. Here we investigated the time course over which shape information can be integrated to allow for recognition of complex objects. We presented fragments of object images in an asynchronous fashion and behaviorally evaluated categorization performance. We observed that visual recognition was significantly disrupted by asynchronies of approximately 30 ms, suggesting that spatiotemporal integration begins to break down with even small deviations from simultaneity. However, moderate temporal asynchrony did not completely obliterate recognition; in fact, integration of visual shape information persisted even with an asynchrony of 100 ms. We describe the data with a concise model based on the dynamic reduction of uncertainty about what image was presented. These results emphasize the importance of timing in visual processing and provide strong constraints for the development of dynamical models of visual shape recognition. PMID:24819738

Ultra-high contrast retinal display system for single photoreceptor psychophysics

PubMed Central

Domdei, Niklas; Domdei, Lennart; Reiniger, Jenny L.; Linden, Michael; Holz, Frank G.; Roorda, Austin; Harmening, Wolf M.

2017-01-01

Due to the enormous dynamic range of human photoreceptors in response to light, studying their visual function in the intact retina challenges the stimulation hardware, specifically with regard to the displayable luminance contrast. The adaptive optics scanning laser ophthalmoscope (AOSLO) is an optical platform that focuses light to extremely small retinal extents, approaching the size of single photoreceptor cells. However, the current light modulation techniques produce spurious visible backgrounds which fundamentally limit experimental options. To remove unwanted background light and to improve contrast for high dynamic range visual stimulation in an AOSLO, we cascaded two commercial fiber-coupled acousto-optic modulators (AOMs) and measured their combined optical contrast. By compensating for zero-point differences in the individual AOMs, we demonstrate a multiplicative extinction ratio in the cascade that was in accordance with the extinction ratios of both single AOMs. When latency differences in the AOM response functions were individually corrected, single switch events as short as 50 ns with radiant power contrasts up to 1:1010 were achieved. This is the highest visual contrast reported for any display system so far. We show psychophysically that this contrast ratio is sufficient to stimulate single foveal photoreceptor cells with small and bright enough visible targets that do not contain a detectable background. Background-free stimulation will enable photoreceptor testing with custom adaptation lights. Furthermore, a larger dynamic range in displayable light levels can drive photoreceptor responses in cones as well as in rods. PMID:29359094

Visualisation of the flow at the tip of a high speed axial flow turbine rotor: An assessment of flow visualisation techniques and the requirement of the experimental turbine

NASA Astrophysics Data System (ADS)

Bindon, J.; Alder, D.; Ianovici, I.

1987-11-01

The field of flow visualization has been reviewed and its application to the study of the flow near the tip of an unshrouded axial turbine rotor discussed in detail. The logical conceptualization of experiments which could lead to a final understanding of the flow structure was developed and how this leads to test turbine design philosophy is suggested. The rotor periodicity shed by the stator requires that particle of pulse tracing is needed rather than the more universal continuous streamline trace which arises from a continuous tracer injection at a point in a flow. While the whole field of flow visualization at a rotor tip is demanding because of its very nature, pulse tracking will place a greater demand on the development of new skills and techniques. Since streamline tracking is somewhat more standard, these demands will not be as great. A fundamental choice does however need to be made between the two methods. The suggested experimental turbine should thus, always with the facility of infinitely variable Mach number, model the following: (1) Stationary annular cascade with tip clearance inside a stationary outer endwall; (2) Stationary annular cascade with tip clearance inside a moving endwall; (3) The transfer of flow visualization techniques developed into the rotating frame; (4) Fully rotating rotor with no inlet periodicity; (5) Fully rotating rotor with inlet periodicity.

Whole-field visual motion drives swimming in larval zebrafish via a stochastic process

PubMed Central

Portugues, Ruben; Haesemeyer, Martin; Blum, Mirella L.; Engert, Florian

2015-01-01

ABSTRACT Caudo-rostral whole-field visual motion elicits forward locomotion in many organisms, including larval zebrafish. Here, we investigate the dependence on the latency to initiate this forward swimming as a function of the speed of the visual motion. We show that latency is highly dependent on speed for slow speeds (<10 mm s−1) and then plateaus for higher values. Typical latencies are >1.5 s, which is much longer than neuronal transduction processes. What mechanisms underlie these long latencies? We propose two alternative, biologically inspired models that could account for this latency to initiate swimming: an integrate and fire model, which is history dependent, and a stochastic Poisson model, which has no history dependence. We use these models to predict the behavior of larvae when presented with whole-field motion of varying speed and find that the stochastic process shows better agreement with the experimental data. Finally, we discuss possible neuronal implementations of these models. PMID:25792753

Whole-field visual motion drives swimming in larval zebrafish via a stochastic process.

PubMed

Portugues, Ruben; Haesemeyer, Martin; Blum, Mirella L; Engert, Florian

2015-05-01

Caudo-rostral whole-field visual motion elicits forward locomotion in many organisms, including larval zebrafish. Here, we investigate the dependence on the latency to initiate this forward swimming as a function of the speed of the visual motion. We show that latency is highly dependent on speed for slow speeds (<10 mm s(-1)) and then plateaus for higher values. Typical latencies are >1.5 s, which is much longer than neuronal transduction processes. What mechanisms underlie these long latencies? We propose two alternative, biologically inspired models that could account for this latency to initiate swimming: an integrate and fire model, which is history dependent, and a stochastic Poisson model, which has no history dependence. We use these models to predict the behavior of larvae when presented with whole-field motion of varying speed and find that the stochastic process shows better agreement with the experimental data. Finally, we discuss possible neuronal implementations of these models. © 2015. Published by The Company of Biologists Ltd.

Incidence loss for fan turbine rotor blade in two-dimensional cascade

NASA Technical Reports Server (NTRS)

Kline, J. F.; Moffitt, T. P.; Stabe, R. G.

1983-01-01

The effect of incidence angle on the aerodynamic performance of a fan turbine rotor blade was investigated experimentally in a two dimensional cascade. The test covered a range of incidence angles from -15 deg to 10 deg and exit ideal critical velocity ratios from 0.75 to 0.95. The principal measurements were blade-surface static pressures and cross-channel survey of exit total pressure, static pressure, and flow angle. Flow adjacent to surfaces was examined using a visualization technique. The results of the investigation include blade-surface velocity distribution and overall kinetic energy loss coefficients for the incidence angles and exit velocity ratios tested. The measured losses are compared with those from a reference core turbine rotor blade and also with two common analytical methods of predicting incidence loss.

Vitiligo: A Possible Model of Degenerative Diseases

PubMed Central

Bellei, Barbara; Pitisci, Angela; Ottaviani, Monica; Ludovici, Matteo; Cota, Carlo; Luzi, Fabiola; Dell'Anna, Maria Lucia; Picardo, Mauro

2013-01-01

Vitiligo is characterized by the progressive disappearance of pigment cells from skin and hair follicle. Several in vitro and in vivo studies show evidence of an altered redox status, suggesting that loss of cellular redox equilibrium might be the pathogenic mechanism in vitiligo. However, despite the numerous data supporting a pathogenic role of oxidative stress, there is still no consensus explanation underlying the oxidative stress-driven disappear of melanocytes from the epidermis. In this study, in vitro characterization of melanocytes cultures from non-lesional vitiligo skin revealed at the cellular level aberrant function of signal transduction pathways common with neurodegenerative diseases including modification of lipid metabolism, hyperactivation of mitogen-activated protein kinase (MAPK) and cAMP response element-binding protein (CREB), constitutive p53-dependent stress signal transduction cascades, and enhanced sensibility to pro-apoptotic stimuli. Notably, these long-term effects of subcytotoxic oxidative stress are also biomarkers of pre-senescent cellular phenotype. Consistent with this, vitiligo cells showed a significant increase in p16 that did not correlate with the chronological age of the donor. Moreover, vitiligo melanocytes produced many biologically active proteins among the senescence-associated secretory phenotype (SAPS), such as interleukin-6 (IL-6), matrix metallo proteinase-3 (MMP3), cyclooxygenase-2 (Cox-2), insulin-like growth factor-binding protein-3 and 7 (IGFBP3, IGFBP7). Together, these data argue for a complicated pathophysiologic puzzle underlying melanocytes degeneration resembling, from the biological point of view, neurodegenerative diseases. Our results suggest new possible targets for intervention that in combination with current therapies could correct melanocytes intrinsic defects. PMID:23555779

Eukaryotic-like Ser/Thr Protein Kinases SpkC/F/K Are Involved in Phosphorylation of GroES in the Cyanobacterium Synechocystis

PubMed Central

Zorina, Anna; Stepanchenko, Natalia; Novikova, Galina V.; Sinetova, Maria; Panichkin, Vladimir B.; Moshkov, Igor E.; Zinchenko, Vladislav V.; Shestakov, Sergey V.; Suzuki, Iwane; Murata, Norio; Los, Dmitry A.

2011-01-01

Serine/threonine protein kinases (STPKs) are the major participants in intracellular signal transduction in eukaryotes, such as yeasts, fungi, plants, and animals. Genome sequences indicate that these kinases are also present in prokaryotes, such as cyanobacteria. However, their roles in signal transduction in prokaryotes remain poorly understood. We have attempted to identify the roles of STPKs in response to heat stress in the prokaryotic cyanobacterium Synechocystis sp. PCC 6803, which has 12 genes for STPKs. Each gene was individually inactivated to generate a gene-knockout library of STPKs. We applied in vitro Ser/Thr protein phosphorylation and phosphoproteomics and identified the methionyl-tRNA synthetase, large subunit of RuBisCO, 6-phosphogluconate dehydrogenase, translation elongation factor Tu, heat-shock protein GrpE, and small chaperonin GroES as the putative targets for Ser/Thr phosphorylation. The expressed and purified GroES was used as an external substrate to screen the protein extracts of the individual mutants for their Ser/Thr kinase activities. The mutants that lack one of the three protein kinases, SpkC, SpkF, and SpkK, were unable to phosphorylate GroES in vitro, suggesting possible interactions between them towards their substrate. Complementation of the mutated SpkC, SpkF, and SpkK leads to the restoration of the ability of cells to phosphorylate the GroES. This suggests that these three STPKs are organized in a sequential order or a cascade and they work one after another to finally phosphorylate the GroES. PMID:21551175

LIM-domain protein AJUBA suppresses malignant mesothelioma cell proliferation via Hippo signaling cascade.

PubMed

Tanaka, I; Osada, H; Fujii, M; Fukatsu, A; Hida, T; Horio, Y; Kondo, Y; Sato, A; Hasegawa, Y; Tsujimura, T; Sekido, Y

2015-01-02

Malignant mesothelioma (MM) is one of the most aggressive neoplasms usually associated with asbestos exposure and is highly refractory to current therapeutic modalities. MMs show frequent activation of a transcriptional coactivator Yes-associated protein (YAP), which is attributed to the neurofibromatosis type 2 (NF2)-Hippo pathway dysfunction, leading to deregulated cell proliferation and acquisition of a malignant phenotype. However, the whole mechanism of disordered YAP activation in MMs has not yet been well clarified. In the present study, we investigated various components of the NF2-Hippo pathway, and eventually found that MM cells frequently showed downregulation of LIM-domain protein AJUBA, a binding partner of large tumor suppressor type 2 (LATS2), which is one of the last-step kinases of the NF2-Hippo pathway. Although loss of AJUBA expression was independent of the alteration status of other Hippo pathway components, MM cell lines with AJUBA inactivation showed a more dephosphorylated (activated) level of YAP. Immunohistochemical analysis showed frequent downregulation of AJUBA in primary MMs, which was associated with YAP constitutive activation. We found that AJUBA transduction into MM cells significantly suppressed promoter activities of YAP-target genes, and the suppression of YAP activity by AJUBA was remarkably canceled by knockdown of LATS2. In connection with these results, transduction of AJUBA-expressing lentivirus significantly inhibited the proliferation and anchorage-independent growth of the MM cells that harbored ordinary LATS family expression. Taken together, our findings indicate that AJUBA negatively regulates YAP activity through the LATS family, and inactivation of AJUBA is a novel key mechanism in MM cell proliferation.

Cholinergic chemosensory cells of the thymic medulla express the bitter receptor Tas2r131.

PubMed

Soultanova, Aichurek; Voigt, Anja; Chubanov, Vladimir; Gudermann, Thomas; Meyerhof, Wolfgang; Boehm, Ulrich; Kummer, Wolfgang

2015-11-01

The thymus is the site of T cell maturation which includes positive selection in the cortex and negative selection in the medulla. Acetylcholine is locally produced in the thymus and cholinergic signaling influences the T cell development. We recently described a distinct subset of medullary epithelial cells in the murine thymus which express the acetylcholine-synthesizing enzyme choline acetyltransferase (ChAT) and components of the canonical taste transduction cascade, i.e. transient receptor potential melastatin-like subtype 5 channel (TRPM5), phospholipase Cβ(2), and Gα-gustducin. Such a chemical phenotype is characteristic for chemosensory cells of mucosal surfaces which utilize bitter receptors for detection of potentially hazardous compounds and cholinergic signaling to initiate avoidance reflexes. We here demonstrate mRNA expression of bitter receptors Tas2r105, Tas2r108, and Tas2r131 in the murine thymus. Using a Tas2r131-tauGFP reporter mouse we localized the expression of this receptor to cholinergic cells expressing the downstream elements of the taste transduction pathway. These cells are distinct from the medullary thymic epithelial cells which promiscuously express tissue-restricted self-antigens during the process of negative selection, since double-labeling immunofluorescence showed no colocalization of autoimmune regulator (AIRE), the key mediator of negative selection, and TRPM5. These data demonstrate the presence of bitter taste-sensing signaling in cholinergic epithelial cells in the thymic medulla and opens a discussion as to what is the physiological role of this pathway. Copyright © 2015 Elsevier B.V. All rights reserved.

Cannabinoid inhibition of adenylate cyclase-mediated signal transduction and interleukin 2 (IL-2) expression in the murine T-cell line, EL4.IL-2.

PubMed

Condie, R; Herring, A; Koh, W S; Lee, M; Kaminski, N E

1996-05-31

Cannabinoid receptors negatively regulate adenylate cyclase through a pertussis toxin-sensitive GTP-binding protein. In the present studies, signaling via the adenylate cyclase/cAMP pathway was investigated in the murine thymoma-derived T-cell line, EL4.IL-2. Northern analysis of EL4.IL-2 cells identified the presence of 4-kilobase CB2 but not CB1 receptor-subtype mRNA transcripts. Southern analysis of genomic DNA digests for the CB2 receptor demonstrated identical banding patterns for EL4.IL-2 cells and mouse-derived DNA, both of which were dissimilar to DNA isolated from rat. Treatment of EL4.IL-2 cells with either cannabinol or Delta9-THC disrupted the adenylate cyclase signaling cascade by inhibiting forskolin-stimulated cAMP accumulation which consequently led to a decrease in protein kinase A activity and the binding of transcription factors to a CRE consensus sequence. Likewise, an inhibition of phorbol 12-myristate 13-acetate (PMA)/ionomycin-induced interleukin 2 (IL-2) protein secretion, which correlated to decreased IL-2 gene transcription, was induced by both cannabinol and Delta9-THC. Further, cannabinoid treatment also decreased PMA/ionomycin-induced nuclear factor binding to the AP-1 proximal site of the IL-2 promoter. Conversely, forskolin enhanced PMA/ionomycin-induced AP-1 binding. These findings suggest that inhibition of signal transduction via the adenylate cyclase/cAMP pathway induces T-cell dysfunction which leads to a diminution in IL-2 gene transcription.

Phosducin-like protein: an ethanol-responsive potential modulator of guanine nucleotide-binding protein function.

PubMed

Miles, M F; Barhite, S; Sganga, M; Elliott, M

1993-11-15

Acute and chronic exposure to ethanol produces specific changes in several signal transduction cascades. Such alterations in signaling are thought to be a crucial aspect of the central nervous system's adaptive response, which occurs with chronic exposure to ethanol. We have recently identified and isolated several genes whose expression is specifically induced by ethanol in neural cell cultures. The product of one of these genes has extensive sequence homology to phosducin, a phosphoprotein expressed in retina and pineal gland that modulates trimeric guanine nucleotide-binding protein (G protein) function by binding to G-protein beta gamma subunits. We identified from a rat brain cDNA library an isolate encoding the phosducin-like protein (PhLP), which has 41% identity and 65% amino acid homology to phosducin. PhLP cDNA is expressed in all tissues screened by RNA blot-hybridization analysis and shows marked evolutionary conservation on Southern hybridization. We have identified four forms of PhLP cDNA varying only in their 5' ends, probably due to alternative splicing. This 5'-end variation generates two predicted forms of PhLP protein that differ by 79 aa at the NH2 terminus. Treatment of NG108-15 cells for 24 hr with concentrations of ethanol seen in actively drinking alcoholics (25-100 mM) causes up to a 3-fold increase in PhLP mRNA levels. Induction of PhLP by ethanol could account for at least some of the widespread alterations in signal transduction and G-protein function that are known to occur with chronic exposure to ethanol.

Nitric oxide and phytohormone interactions: current status and perspectives

PubMed Central

Freschi, Luciano

2013-01-01

Nitric oxide (NO) is currently considered a ubiquitous signal in plant systems, playing significant roles in a wide range of responses to environmental and endogenous cues. During the signaling events leading to these plant responses, NO frequently interacts with plant hormones and other endogenous molecules, at times originating remarkably complex signaling cascades. Accumulating evidence indicates that virtually all major classes of plant hormones may influence, at least to some degree, the endogenous levels of NO. In addition, studies conducted during the induction of diverse plant responses have demonstrated that NO may also affect biosynthesis, catabolism/conjugation, transport, perception, and/or transduction of different phytohormones, such as auxins, gibberellins, cytokinins, abscisic acid, ethylene, salicylic acid, jasmonates, and brassinosteroids. Although still not completely elucidated, the mechanisms underlying the interaction between NO and plant hormones have recently been investigated in a number of species and plant responses. This review specifically focuses on the current knowledge of the mechanisms implicated in NO–phytohormone interactions during the regulation of developmental and metabolic plant events. The modifications triggered by NO on the transcription of genes encoding biosynthetic/degradative enzymes as well as proteins involved in the transport and signal transduction of distinct plant hormones will be contextualized during the control of developmental, metabolic, and defense responses in plants. Moreover, the direct post-translational modification of phytohormone biosynthetic enzymes and receptors through S-nitrosylation will also be discussed as a key mechanism for regulating plant physiological responses. Finally, some future perspectives toward a more complete understanding of NO–phytohormone interactions will also be presented and discussed. PMID:24130567

ATRIAL NATRIURETIC FACTOR RECEPTOR GUANYLATE CYCLASE SIGNALING: NEW ATP- REGULATED TRANSDUCTION MOTIF

PubMed Central

Duda, Teresa; Bharill, Shashank; Wojtas, Ireneusz; Yadav, Prem; Gryczynski, Ignacy; Gryczynski, Zygmunt; Sharma, Rameshwar K.

2010-01-01

ANF-RGC$ membrane guanylate cyclase is the receptor for the hypotensive peptide hormones, atrial natriuretic factor (ANF) and type B natriuretic peptide (BNP). It is a single transmembrane spanning protein. Binding the hormone to the extracellular domain activates its intracellular catalytic domain. This results in accelerated production of cyclic GMP, a second messenger in controlling blood pressure, cardiac vasculature and fluid secretion. ATP is the obligatory transducer of the ANF signal. It works through its ATP regulated module, ARM, which is juxtaposed to the C-terminal side of the transmembrane domain. Upon interaction, ATP induces a cascade of temporal and spatial changes in the ARM, which, finally, result in activation of the catalytic module. Although the exact nature and the details of these changes are not known, some of these have been stereographed in the simulated three-dimensional model of the ARM and validated biochemically. Through comprehensive techniques ofsteady-state, time-resolved tryptophan fluorescence and Forster Resonance Energy Transfer (FRET), site-directed and deletion-mutagenesis, and reconstitution, the present study validates and explains themechanism of the model-based predicted transduction role of the ARM’s structural motif, 669WTAPELL675. This motif is critical in the ATP-dependent ANF signaling. Molecular modeling shows that ATP binding exposes the 669WTAPELL675 motif, the exposure, in turn, facilitates its interaction and activation of the catalytic module. These principles of the model have been experimentally validated. This knowledge brings us a step closer to our understanding of the mechanism by which the ATP-dependent spatial changes within the ARM cause ANF signaling of ANF-RGC. PMID:19137266

Towards systems biology of the gravity response of higher plants -multiscale analysis of Arabidopsis thaliana root growth

NASA Astrophysics Data System (ADS)

Palme, Klaus; Aubry, D.; Bensch, M.; Schmidt, T.; Ronneberger, O.; Neu, C.; Li, X.; Wang, H.; Santos, F.; Wang, B.; Paponov, I.; Ditengou, F. A.; Teale, W. T.; Volkmann, D.; Baluska, F.; Nonis, A.; Trevisan, S.; Ruperti, B.; Dovzhenko, A.

Gravity plays a fundamental role in plant growth and development. Up to now, little is known about the molecular organisation of the signal transduction cascades and networks which co-ordinate gravity perception and response. By using an integrated systems biological approach, a systems analysis of gravity perception and the subsequent tightly-regulated growth response is planned in the model plant Arabidopsis thaliana. This approach will address questions such as: (i) what are the components of gravity signal transduction pathways? (ii) what are the dynamics of these components? (iii) what is their spatio-temporal regulation in different tis-sues? Using Arabidopsis thaliana as a model-we use root growth to obtain insights in the gravity response. New techniques enable identification of the individual genes affected by grav-ity and further integration of transcriptomics and proteomics data into interaction networks and cell communication events that operate during gravitropic curvature. Using systematic multiscale analysis we have identified regulatory networks consisting of transcription factors, the protein degradation machinery, vesicle trafficking and cellular signalling during the gravire-sponse. We developed approach allowing to incorporate key features of the root system across all relevant spatial and temporal scales to describe gene-expression patterns and correlate them with individual gene and protein functions. Combination of high-resolution microscopy and novel computational tools resulted in development of the root 3D model in which quantitative descriptions of cellular network properties and of multicellular interactions important in root growth and gravitropism can be integrated for the first time.

An Odor-Specific Threshold Deficit Implicates Abnormal Intracellular Cyclic AMP Signaling in Schizophrenia

PubMed Central

Turetsky, Bruce I.; Moberg, Paul J.

2012-01-01

Objective Although olfactory deficits are common in schizophrenia, their underlying pathophysiology remains unknown. Recent evidence has suggested that cAMP signaling may be disrupted in schizophrenia. Since cAMP mediates signal transduction in olfactory receptor neurons, this could contribute to the etiology of observed olfactory deficits. This study was designed to test this hypothesis by determining odor detection threshold sensitivities to two odorants that differ in their relative activations of this intracellular cAMP signaling cascade. Method Thirty schizophrenia patients, 25 healthy comparison subjects, and 19 unaffected first-degree relatives of schizophrenia patients were studied. Odor detection threshold sensitivities were measured for the two odorants citralva and lyral. Although both have fruity/floral scents, citralva strongly activates adenylyl cyclase to increase cAMP levels, while lyral is a very weak activator of adenylyl cyclase. Results There was a significant group-by-odor interaction. Both schizophrenia patients and unaffected first-degree relatives were impaired in their ability to detect lyral versus citralva. Comparison subjects were equally sensitive to both odorants. This selective deficit could not be explained by differences in age, sex, smoking, clinical symptom profile, or medication use. Conclusions This study establishes the presence of an odor-specific hyposmia that may denote a disruption of cAMP-mediated signal transduction in schizophrenia. The presence of a parallel deficit in the patients’ unaffected first-degree relatives suggests that this deficit is genetically mediated. Although additional physiological studies are needed to confirm the underlying mechanism, these results offer strong inferential support for the hypothesis that cAMP signaling is dys-regulated in schizophrenia. PMID:19074977

An odor-specific threshold deficit implicates abnormal intracellular cyclic AMP signaling in schizophrenia.

PubMed

Turetsky, Bruce I; Moberg, Paul J

2009-02-01

Although olfactory deficits are common in schizophrenia, their underlying pathophysiology remains unknown. Recent evidence has suggested that cAMP signaling may be disrupted in schizophrenia. Since cAMP mediates signal transduction in olfactory receptor neurons, this could contribute to the etiology of observed olfactory deficits. This study was designed to test this hypothesis by determining odor detection threshold sensitivities to two odorants that differ in their relative activations of this intracellular cAMP signaling cascade. Thirty schizophrenia patients, 25 healthy comparison subjects, and 19 unaffected first-degree relatives of schizophrenia patients were studied. Odor detection threshold sensitivities were measured for the two odorants citralva and lyral. Although both have fruity/floral scents, citralva strongly activates adenylyl cyclase to increase cAMP levels, while lyral is a very weak activator of adenylyl cyclase. There was a significant group-by-odor interaction. Both schizophrenia patients and unaffected first-degree relatives were impaired in their ability to detect lyral versus citralva. Comparison subjects were equally sensitive to both odorants. This selective deficit could not be explained by differences in age, sex, smoking, clinical symptom profile, or medication use. This study establishes the presence of an odor-specific hyposmia that may denote a disruption of cAMP-mediated signal transduction in schizophrenia. The presence of a parallel deficit in the patients' unaffected first-degree relatives suggests that this deficit is genetically mediated. Although additional physiological studies are needed to confirm the underlying mechanism, these results offer strong inferential support for the hypothesis that cAMP signaling is dysregulated in schizophrenia.

Human Chorionic Gonadotropin: Unknown about Known.

PubMed

Borisova, M A; Moiseenko, D Yu; Smirnova, O V

2017-01-01

The last two decade discoveries shift the accent from consideration of human chorionic gonadotripin (hCG) as a hormone, that controls progesterone production by corpus luteum cells, to a powerful paracrine regulator which'in the tandem with its hyperglycozilated analog (hCG-H) induces successful implantation and coordinated dialog between blastocyst and uterus tissues. Ability of hCG to interact with TSH receptor and hCG-H with TGF-beta-RII extend significantly the spectrum of processes controlled by these molecules. Differences between intracellular pathways of signal transduction between hCG and LH mediated by the same receptor (LH/hCG-R) impugn unity of their effector mechanisms previously considered as obvious. Paracine properties-of hCG comprise control of fusing of trophoblasts into syncytiotrophoblasts, angiogenesis, immunity regulation and endometrium predisposition to implantation. Angiogenesis is associated with LH/hCG-R expressed on mural cells of uterine spiral arteries as well as induced secretion of soluble VEGF type by endometrial cells. hCG.regulates ratio between different forms of T-helper cells in maternal organism on the initial gestation stage determining high level of Th2 cells. hCG supports local immunotolerance acting as chemoattractant for T-suppressors (T-Treg) and apoptotic factor for T-lymphocytes. Endometrial susceptibility arises from activation of osteopantin secretion and decline of mucin secretion by epithelial cells. hCG-H acts on the same tissues as hCG as a paracrine agent regulating multiple cascades of cytokines. hCG-H plays the key role in trophoblast invasion into,uterine decidua as a result of gelatinase secretion by these cells.The degree of angiogenic effect of hCG-H is compatiblewith hCG but its signal transduction is mediated by TGF-beta signal transduction pathway that stimulates mural cell proliferation. hCG-H acts as mitogen on NK-cells and is able to activate them and direct to angiogenesis maintenance. In this article the attempt was made to elucidate the most important discoveries about the role of hCG and its hyperglycosylated analog yet accomplished and still upcoming.

Visualization of boundary-layer development on turbomachine blades with liquid crystals

NASA Technical Reports Server (NTRS)

Vanzante, Dale E.; Okiishi, Theodore H.

1991-01-01

This report documents a study of the use of liquid crystals to visualize boundary layer development on a turbomachine blade. A turbine blade model in a linear cascade of blades was used for the tests involved. Details of the boundary layer development on the suction surface of the turbine blade model were known from previous research. Temperature sensitive and shear sensitive liquid crystals were tried as visual agents. The temperature sensitive crystals were very effective in their ability to display the location of boundary layer flow separation and reattachment. Visualization of natural transition from laminar to turbulent boundary layer flow with the temperature sensitive crystals was possible but subtle. The visualization of separated flow reattachment with the shear sensitive crystals was easily accomplished when the crystals were allowed to make a transition from the focal-conic to a Grandjean texture. Visualization of flow reattachment based on the selective reflection properties of shear sensitive crystals was achieved only marginally because of the larger surface shear stress and shear stress gradient levels required for more dramatic color differences.

Single Quantum Dot Tracking Reveals that an Individual Multivalent HIV-1 Tat Protein Transduction Domain Can Activate Machinery for Lateral Transport and Endocytosis

PubMed Central

Roy, Chandra Nath; Promjunyakul, Warunya; Hatakeyama, Hiroyasu; Gonda, Kohsuke; Imamura, Junji; Vasudevanpillai, Biju; Ohuchi, Noriaki; Kanzaki, Makoto; Higuchi, Hideo; Kaku, Mitsuo

2013-01-01

The mechanisms underlying the cellular entry of the HIV-1 Tat protein transduction domain (TatP) and the molecular information necessary to improve the transduction efficiency of TatP remain unclear due to the technical limitations for direct visualization of TatP's behavior in cells. Using confocal microscopy, total internal reflection fluorescence microscopy, and four-dimensional microscopy, we developed a single-molecule tracking assay for TatP labeled with quantum dots (QDs) to examine the kinetics of TatP initially and immediately before, at the beginning of, and immediately after entry into living cells. We report that even when the number of multivalent TatP (mTatP)-QDs bound to a cell was low, each single mTatP-QD first locally induced the cell's lateral transport machinery to move the mTatP-QD toward the center of the cell body upon cross-linking of heparan sulfate proteoglycans. The centripetal and lateral movements were linked to the integrity and flow of actomyosin and microtubules. Individual mTatP underwent lipid raft-mediated temporal confinement, followed by complete immobilization, which ultimately led to endocytotic internalization. However, bivalent TatP did not sufficiently promote either cell surface movement or internalization. Together, these findings provide clues regarding the mechanisms of TatP cell entry and indicate that increasing the valence of TatP on nanoparticles allows them to behave as cargo delivery nanomachines. PMID:23732912

Footprint area analysis of binary imaged Cupriavidus necator cells to study PHB production at balanced, transient, and limited growth conditions in a cascade process.

PubMed

Vadlja, Denis; Koller, Martin; Novak, Mario; Braunegg, Gerhart; Horvat, Predrag

2016-12-01

Statistical distribution of cell and poly[3-(R)-hydroxybutyrate] (PHB) granule size and number of granules per cell are investigated for PHB production in a five-stage cascade (5CSTR). Electron microscopic pictures of cells from individual cascade stages (R1-R5) were converted to binary pictures to visualize footprint areas for polyhydroxyalkanoate (PHA) and non-PHA biomass. Results for each stage were correlated to the corresponding experimentally determined kinetics (specific growth rate μ and specific productivity π). Log-normal distribution describes PHA granule size dissimilarity, whereas for R1 and R4, gamma distribution best reflects the situation. R1, devoted to balanced biomass synthesis, predominately contains cells with rather small granules, whereas with increasing residence time τ, maximum and average granule sizes by trend increase, approaching an upper limit determined by the cell's geometry. Generally, an increase of intracellular PHA content and ratio of granule to cell area slow down along the cascade. Further, the number of granules per cell decreases with increasing τ. Data for μ and π obtained by binary picture analysis correlate well with the experimental results. The work describes long-term continuous PHA production under balanced, transient, and nutrient-deficient conditions, as well as their reflection on the granules size, granule number, and cell structure on the microscopic level.

Local measurement and numerical modeling of mass/heat transfer from a turbine blade in a linear cascade with tip clearance

NASA Astrophysics Data System (ADS)

Jin, Peitong

2000-11-01

Local mass/heat transfer measurements from the turbine blade near-tip and the tip surfaces are performed using the naphthalene sublimation technique. The experiments are conducted in a linear cascade consisting of five high-pressure blades with a central test-blade configuration. The incoming flow conditions are close to those of the gas turbine engine environment (boundary layer displacement thickness is about 0.01 of chord) with an exit Reynolds number of 6.2 x 105. The effects of tip clearance level (0.86%--6.90% of chord), mainstream Reynolds number and turbulence intensity (0.2 and 12.0%) are investigated. Two methods of flow visualization---oil and lampblack, laser light sheet smoke wire---as well as static pressure measurement on the blade surface are used to study the tip leakage flow and vortex in the cascade. In addition, numerical modeling of the flow and heat transfer processes in the linear cascade with different tip clearances is conducted using commercial software incorporating advanced turbulence models. The present study confirms many important results on the tip leakage flow and vortex from the literature, contributes to the current understanding in the effects of tip leakage flow and vortex on local heat transfer from the blade near-tip and the tip surfaces, and provides detailed local and average heat/mass transfer data applicable to turbine blade tip cooling design.

Action Video Gaming and Cognitive Control: Playing First Person Shooter Games Is Associated with Improved Action Cascading but Not Inhibition

PubMed Central

Steenbergen, Laura; Sellaro, Roberta; Stock, Ann-Kathrin; Beste, Christian; Colzato, Lorenza S.

2015-01-01

There is a constantly growing interest in developing efficient methods to enhance cognitive functioning and/or to ameliorate cognitive deficits. One particular line of research focuses on the possibly cognitive enhancing effects that action video game (AVG) playing may have on game players. Interestingly, AVGs, especially first person shooter games, require gamers to develop different action control strategies to rapidly react to fast moving visual and auditory stimuli, and to flexibly adapt their behaviour to the ever-changing context. This study investigated whether and to what extent experience with such videogames is associated with enhanced performance on cognitive control tasks that require similar abilities. Experienced action videogame-players (AVGPs) and individuals with little to no videogame experience (NVGPs) performed a stop-change paradigm that provides a relatively well-established diagnostic measure of action cascading and response inhibition. Replicating previous findings, AVGPs showed higher efficiency in response execution, but not improved response inhibition (i.e. inhibitory control), as compared to NVGPs. More importantly, compared to NVGPs, AVGPs showed enhanced action cascading processes when an interruption (stop) and a change towards an alternative response were required simultaneously, as well as when such a change had to occur after the completion of the stop process. Our findings suggest that playing AVGs is associated with enhanced action cascading and multi-component behaviour without affecting inhibitory control. PMID:26655929

Action Video Gaming and Cognitive Control: Playing First Person Shooter Games Is Associated with Improved Action Cascading but Not Inhibition.

PubMed

Steenbergen, Laura; Sellaro, Roberta; Stock, Ann-Kathrin; Beste, Christian; Colzato, Lorenza S

2015-01-01

There is a constantly growing interest in developing efficient methods to enhance cognitive functioning and/or to ameliorate cognitive deficits. One particular line of research focuses on the possibly cognitive enhancing effects that action video game (AVG) playing may have on game players. Interestingly, AVGs, especially first person shooter games, require gamers to develop different action control strategies to rapidly react to fast moving visual and auditory stimuli, and to flexibly adapt their behaviour to the ever-changing context. This study investigated whether and to what extent experience with such videogames is associated with enhanced performance on cognitive control tasks that require similar abilities. Experienced action videogame-players (AVGPs) and individuals with little to no videogame experience (NVGPs) performed a stop-change paradigm that provides a relatively well-established diagnostic measure of action cascading and response inhibition. Replicating previous findings, AVGPs showed higher efficiency in response execution, but not improved response inhibition (i.e. inhibitory control), as compared to NVGPs. More importantly, compared to NVGPs, AVGPs showed enhanced action cascading processes when an interruption (stop) and a change towards an alternative response were required simultaneously, as well as when such a change had to occur after the completion of the stop process. Our findings suggest that playing AVGs is associated with enhanced action cascading and multi-component behaviour without affecting inhibitory control.

GenePublisher: Automated analysis of DNA microarray data.

PubMed

Knudsen, Steen; Workman, Christopher; Sicheritz-Ponten, Thomas; Friis, Carsten

2003-07-01

GenePublisher, a system for automatic analysis of data from DNA microarray experiments, has been implemented with a web interface at http://www.cbs.dtu.dk/services/GenePublisher. Raw data are uploaded to the server together with a specification of the data. The server performs normalization, statistical analysis and visualization of the data. The results are run against databases of signal transduction pathways, metabolic pathways and promoter sequences in order to extract more information. The results of the entire analysis are summarized in report form and returned to the user.

Effects of harvest on carbon and nitrogen dynamics in a Pacific Northwest forest catchment

Treesearch

Alex Abdelnour; Robert B. McKane; Marc Stieglitz; Feifei Pan; Yiwei Cheng

2013-01-01

We used a new ecohydrological model, Visualizing Ecosystems for Land Management Assessments (VELMA), to analyze the effects of forest harvest on catchment carbon and nitrogen dynamics. We applied the model to a 10 ha headwater catchment in the western Oregon Cascade Range where two major disturbance events have occurred during the past 500 years: a stand-replacing fire...

Finding Brown's peony a sweet attraction

Treesearch

Nan. Vance

2012-01-01

I first encountered Brown’s peony (Paeonia brownie) with its verdant, lavender-tinged leaves and elegantly nodding maroon flowers growing among bitterbrush and bunchgrass on the eastern flank of the Oregon Cascades. My first thought was “What is a plant like you doing in a place like this?” It would be natural to visualize this native wild peony as...

R4 RGS Proteins: Regulation of G Protein Signaling and Beyond

PubMed Central

Bansal, Geetanjali; Druey, Kirk M.; Xie, Zhihui

2007-01-01

The Regulators of G protein Signaling (RGS) proteins were initially characterized as inhibitors of signal transduction cascades initiated by G-protein-coupled receptors (GPCRs) because of their ability to increase the intrinsic GTPase activity of heterotrimeric G proteins. This GTPase accelerating (GAP) activity enhances G protein deactivation and promotes desensitization. However, in addition to this signature trait, emerging data have revealed an expanding network of proteins, lipids, and ions that interact with RGS proteins and confer additional regulatory functions. This review highlights recent advances in our understanding of the physiological functions of one subfamily of RGS proteins with a high degree of homology (B/R4) gleaned from recent studies of knockout mice or cells with reduced RGS expression. We also discuss some of the newly-appreciated interactions of RGS proteins with cellular factors that suggest RGS control of several components of G-protein-mediated pathways as well as a diverse array of non-GPCR-mediated biological responses. PMID:18006065

Mechanisms implicated in the effects of boron on wound healing.

PubMed

Nzietchueng, Rosine Mayap; Dousset, Brigitte; Franck, Patricia; Benderdour, Mohamed; Nabet, Pierre; Hess, Ketsia

2002-01-01

Recently, we demonstrated that boron modulates the turnover of the extracellular matrix and increases TNFalpha release. In the present study, we used an in vitro test to investigate the direct effect of boron on specific enzymes (elastase, trypsin-like enzymes, collagenase and alkaline phosphatase) implicated in extracellular matrix turnover. Boron decreased the elastase and alkaline phosphatase activity, but had no effect on trypsin and collagenase activities. The effect of boron on the enzyme activities was also tested in fibroblasts considered as an in vivo test. In contrast to the results obtained in vitro, boron enhanced the trypsin-like, collagenase, and cathepsin D activities in fibroblasts. Boron did not modify the generation of free radicals compared to the control and did not seem to act on the intracellular alkaline phosphatase activity, However, as it did enhance phosphorylation, it can be hypothesized that boron may affect living cells via a mediator, which could be TNFalpha whose transduction signal involves a cascade of phosphorylations.

Conformational stability of the epidermal growth factor (EGF) receptor as influenced by glycosylation, dimerization and EGF hormone binding.

PubMed

Taylor, Eric S; Pol-Fachin, Laercio; Lins, Roberto D; Lower, Steven K

2017-04-01

The epidermal growth factor receptor (EGFR) is an important transmembrane glycoprotein kinase involved the initiation or perpetuation of signal transduction cascades within cells. These processes occur after EGFR binds to a ligand [epidermal growth factor (EGF)], thus inducing its dimerization and tyrosine autophosphorylation. Previous publications have highlighted the importance of glycosylation and dimerization for promoting proper function of the receptor and conformation in membranes; however, the effects of these associations on the protein conformational stability have not yet been described. Molecular dynamics simulations were performed to characterize the conformational preferences of the monomeric and dimeric forms of the EGFR extracellular domain upon binding to EGF in the presence and absence of N-glycan moieties. Structural stability analyses revealed that EGF provides the most conformational stability to EGFR, followed by glycosylation and dimerization, respectively. The findings also support that EGF-EGFR binding takes place through a large-scale induced-fitting mechanism. Proteins 2017; 85:561-570. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Brain-derived neurotrophic factor and its clinical implications

PubMed Central

Bathina, Siresha

2015-01-01

Brain-derived neurotrophic factor (BDNF) plays an important role in neuronal survival and growth, serves as a neurotransmitter modulator, and participates in neuronal plasticity, which is essential for learning and memory. It is widely expressed in the CNS, gut and other tissues. BDNF binds to its high affinity receptor TrkB (tyrosine kinase B) and activates signal transduction cascades (IRS1/2, PI3K, Akt), crucial for CREB and CBP production, that encode proteins involved in β cell survival. BDNF and insulin-like growth factor-1 have similar downstream signaling mechanisms incorporating both p-CAMK and MAPK that increase the expression of pro-survival genes. Brain-derived neurotrophic factor regulates glucose and energy metabolism and prevents exhaustion of β cells. Decreased levels of BDNF are associated with neurodegenerative diseases with neuronal loss, such as Parkinson's disease, Alzheimer's disease, multiple sclerosis and Huntington's disease. Thus, BDNF may be useful in the prevention and management of several diseases including diabetes mellitus. PMID:26788077

FGF-mediated mesoderm induction involves the Src-family kinase Laloo.

PubMed

Weinstein, D C; Marden, J; Carnevali, F; Hemmati-Brivanlou, A

1998-08-27

During embryogenesis, inductive interactions underlie the development of much of the body plan. In Xenopus laevis, factors secreted from the vegetal pole induce mesoderm in the adjacent marginal zone; members of both the transforming growth factor-beta (TGF-beta) and fibroblast growth factor (FGF) ligand families seem to have critical roles in this process. Here we report the identification and characterization of laloo, a novel participant in the signal transduction cascade linking extracellular, mesoderm-inducing signals to the nucleus, where alteration of cell fate is driven by changes in gene expression. Overexpression of laloo, a member of the Src-related gene family, in Xenopus embryos gives rise to ectopic posterior structures that frequently contain axial tissue. Laloo induces mesoderm in Xenopus ectodermal explants; this induction is blocked by reagents that disrupt the FGF signalling pathway. Conversely, expression of a dominant-inhibitory Laloo mutant blocks mesoderm induction by FGF and causes severe posterior truncations in vivo. This work provides the first evidence that a Src-related kinase is involved in vertebrate mesoderm induction.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Becker, Verena; Sengupta, D; Ketteler, Robin

The formation of signal-promoting dimeric or oligomeric receptor complexes at the cell surface is modulated by self-interaction of their transmembrane (TM) domains. To address the importance of TM domain packing density for receptor functionality, we examined a set of asparagine mutants in the TM domain of the erythropoietin receptor (EpoR). We identified EpoR-T242N as a receptor variant that is present at the cell surface similar to wild-type EpoR but lacks visible localization in vesicle-like structures and is impaired in efficient activation of specific signaling cascades. Analysis by a molecular modeling approach indicated an increased interhelical distance for the EpoR-T242N TMmore » dimer. By employing the model, we designed additional mutants with increased or decreased packing volume and confirmed a correlation between packing volume and biological responsiveness. These results propose that the packing density of the TM domain provides a novel layer for fine-tuned regulation of signal transduction and cellular decisions.« less

Strigolactones: new plant hormones in action.

PubMed

Zwanenburg, Binne; Pospíšil, Tomáš; Ćavar Zeljković, Sanja

2016-06-01

The key step in the mode of action of strigolactones is the enzymatic detachment of the D-ring. The thus formed hydroxy butenolide induces conformational changes of the receptor pocket which trigger a cascade of reactions in the signal transduction. Strigolactones (SLs) constitute a new class of plant hormones which are of increasing importance in plant science. For the last 60 years, they have been known as germination stimulants for parasitic plants. Recently, several new bio-properties of SLs have been discovered such as the branching factor for arbuscular mycorrhizal fungi, regulation of plant architecture (inhibition of bud outgrowth and of shoot branching) and the response to abiotic factors, etc. To broaden horizons and encourage new ideas for identifying and synthesising new and structurally simple SLs, this review is focused on molecular aspects of this new class of plant hormones. Special attention has been given to structural features, the mode of action of these phytohormones in various biological actions, the design of SL analogs and their applications.

Emerging roles of Hippo signaling in inflammation and YAP-driven tumor immunity.

PubMed

Zhou, Yuhang; Huang, Tingting; Zhang, Jinglin; Cheng, Alfred S L; Yu, Jun; Kang, Wei; To, Ka Fai

2018-07-10

Initially identified as a cell and organ size controller, Hippo pathway turns into a hotspot for researchers. Within recent years, more and more mechanisms about Hippo pathway were uncovered. Even though Hippo signaling has been revealed to exert controversial roles according to different cell context and microenvironment, which is because of its diversified interplays with a great variety of signaling transduction cascades; mechanisms other than size-limitation, however, remain to be elucidated. Recently, a growing number of studies tend to put Hippo on inflammatory and immunological focus: its antimicrobial role in flies, its pro- or anti-inflammation in mammals, as well as its relevance to cancerous immunity. From inflammation to tumor immunogenicity, Hippo has been gradually justified to play a crucial role. This review summarized the latest findings regarding the involvement of Hippo pathway in immunity, and a more comprehensive understanding of Hippo pathway will shed light on clinical translational potential even precision medicine. Copyright © 2018 Elsevier B.V. All rights reserved.

Intramuscular triacylglycerol and insulin resistance: guilty as charged or wrongly accused?

PubMed

Muoio, Deborah M

2010-03-01

The term lipotoxicity elicits visions of steatotic liver, fat laden skeletal muscles and engorged lipid droplets that spawn a number of potentially harmful intermediates that can wreak havoc on signal transduction and organ function. Prominent among these so-called lipotoxic mediators are signaling molecules such as long chain acyl-CoAs, ceramides and diacyglycerols; each of which is thought to engage serine kinases that disrupt the insulin signaling cascade, thereby causing insulin resistance. Defects in skeletal muscle fat oxidation have been implicated as a driving factor contributing to systemic lipid imbalance, whereas exercise-induced enhancement of oxidative potential is considered protective. The past decade of diabetes research has focused heavily on the foregoing scenario, and indeed the model is grounded in strong experimental evidence, albeit largely correlative. This review centers on mechanisms that connect lipid surplus to insulin resistance in skeletal muscle, as well as those that underlie the antilipotoxic actions of exercise. Emphasis is placed on recent studies that challenge accepted paradigms. Copyright (c) 2009. Published by Elsevier B.V.

Intramuscular triacylglycerol and insulin resistance: Guilty as charged or wrongly accused?

PubMed Central

Muoio, Deborah M.

2015-01-01

The term lipotoxicity elicits visions of steatotic liver, fat laden skeletal muscles and engorged lipid droplets that spawn a number of potentially harmful intermediates that can wreak havoc on signal transduction and organ function. Prominent among these so-called lipotoxic mediators are signaling molecules such as long chain acyl-CoAs, ceramides and diacyglycerols; each of which is thought to engage serine kinases that disrupt the insulin signaling cascade, thereby causing insulin resistance. Defects in skeletal muscle fat oxidation have been implicated as a driving factor contributing to systemic lipid imbalance, whereas exercise-induced enhancement of oxidative potential is considered protective. The past decade of diabetes research has focused heavily on the foregoing scenario, and indeed the model is grounded in strong experimental evidence, albeit largely correlative. This review centers on mechanisms that connect lipid surplus to insulin resistance in skeletal muscle, as well as those that underlie the antilipotoxic actions of exercise. Emphasis is placed on recent studies that challenge accepted paradigms. PMID:19958841

Regulation of EGFR signal transduction by analogue-to-digital conversion in endosomes

PubMed Central

Villaseñor, Roberto; Nonaka, Hidenori; Del Conte-Zerial, Perla; Kalaidzidis, Yannis; Zerial, Marino

2015-01-01

An outstanding question is how receptor tyrosine kinases (RTKs) determine different cell-fate decisions despite sharing the same signalling cascades. Here, we uncovered an unexpected mechanism of RTK trafficking in this process. By quantitative high-resolution FRET microscopy, we found that phosphorylated epidermal growth factor receptor (p-EGFR) is not randomly distributed but packaged at constant mean amounts in endosomes. Cells respond to higher EGF concentrations by increasing the number of endosomes but keeping the mean p-EGFR content per endosome almost constant. By mathematical modelling, we found that this mechanism confers both robustness and regulation to signalling output. Different growth factors caused specific changes in endosome number and size in various cell systems and changing the distribution of p-EGFR between endosomes was sufficient to reprogram cell-fate decision upon EGF stimulation. We propose that the packaging of p-RTKs in endosomes is a general mechanism to ensure the fidelity and specificity of the signalling response. DOI: http://dx.doi.org/10.7554/eLife.06156.001 PMID:25650738

PI3K Phosphorylation Is Linked to Improved Electrical Excitability in an In Vitro Engineered Heart Tissue Disease Model System.

PubMed

Kana, Kujaany; Song, Hannah; Laschinger, Carol; Zandstra, Peter W; Radisic, Milica

2015-09-01

Myocardial infarction, a prevalent cardiovascular disease, is associated with cardiomyocyte cell death, and eventually heart failure. Cardiac tissue engineering has provided hopes for alternative treatment options, and high-fidelity tissue models for drug discovery. The signal transduction mechanisms relayed in response to mechanoelectrical (physical) stimulation or biochemical stimulation (hormones, cytokines, or drugs) in engineered heart tissues (EHTs) are poorly understood. In this study, an EHT model was used to elucidate the signaling mechanisms involved when insulin was applied in the presence of electrical stimulation, a stimulus that mimics functional heart tissue environment in vitro. EHTs were insulin treated, electrically stimulated, or applied in combination (insulin and electrical stimulation). Electrical excitability parameters (excitation threshold and maximum capture rate) were measured. Protein kinase B (AKT) and phosphatidylinositol-3-kinase (PI3K) phosphorylation revealed that insulin and electrical stimulation relayed electrical excitability through two separate signaling cascades, while there was a negative crosstalk between sustained activation of AKT and PI3K.

FLCN Maintains the Leucine Level in Lysosome to Stimulate mTORC1

PubMed Central

Chen, Zhi; Ji, Xin; Qiao, Xianfeng; Jin, Yaping; Liu, Wei

2016-01-01

The intracellular amino acid pool within lysosome is a signal that stimulates the nutrient-sensing mTORC1 signalling pathway. The signal transduction cascade has garnered much attention, but little is known about the sequestration of the signalling molecules within the lysosome. Using human HEK293 cells as a model, we found that suppression of the BHD syndrome gene FLCN reduced the leucine level in lysosome, which correlated with decreased mTORC1 activity. Both consequences could be reversed by supplementation with high levels of leucine, but not other tested amino acids. Conversely, overexpressed FLCN could sequester lysosomal leucine and stimulate mTORC1 in an amino acid limitation environment. These results identify a novel function of FLCN: it controls mTORC1 by modulating the leucine signal in lysosome. Furthermore, we provided evidence that FLCN exerted this role by inhibiting the accumulation of the amino acid transporter PAT1 on the lysosome surface, thereby maintaining the signal level within the organelle. PMID:27280402

Synthetic Biomaterials to Rival Nature's Complexity-a Path Forward with Combinatorics, High-Throughput Discovery, and High-Content Analysis.

PubMed

Zhang, Douglas; Lee, Junmin; Kilian, Kristopher A

2017-10-01

Cells in tissue receive a host of soluble and insoluble signals in a context-dependent fashion, where integration of these cues through a complex network of signal transduction cascades will define a particular outcome. Biomaterials scientists and engineers are tasked with designing materials that can at least partially recreate this complex signaling milieu towards new materials for biomedical applications. In this progress report, recent advances in high throughput techniques and high content imaging approaches that are facilitating the discovery of efficacious biomaterials are described. From microarrays of synthetic polymers, peptides and full-length proteins, to designer cell culture systems that present multiple biophysical and biochemical cues in tandem, it is discussed how the integration of combinatorics with high content imaging and analysis is essential to extracting biologically meaningful information from large scale cellular screens to inform the design of next generation biomaterials. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Cyclic-nucleotide–gated cation current and Ca2+-activated Cl current elicited by odorant in vertebrate olfactory receptor neurons

PubMed Central

Li, Rong-Chang; Ben-Chaim, Yair; Yau, King-Wai; Lin, Chih-Chun

2016-01-01

Olfactory transduction in vertebrate olfactory receptor neurons (ORNs) involves primarily a cAMP-signaling cascade that leads to the opening of cyclic-nucleotide–gated (CNG), nonselective cation channels. The consequent Ca2+ influx triggers adaptation but also signal amplification, the latter by opening a Ca2+-activated Cl channel (ANO2) to elicit, unusually, an inward Cl current. Hence the olfactory response has inward CNG and Cl components that are in rapid succession and not easily separable. We report here success in quantitatively separating these two currents with respect to amplitude and time course over a broad range of odorant strengths. Importantly, we found that the Cl current is the predominant component throughout the olfactory dose–response relation, down to the threshold of signaling to the brain. This observation is very surprising given a recent report by others that the olfactory-signal amplification effected by the Ca2+-activated Cl current does not influence the behavioral olfactory threshold in mice. PMID:27647918

Evolution of olfactory receptors.

PubMed

Hoover, Kara C

2013-01-01

Olfactory receptors are a specialized set of receptor cells responsible for the detection of odors. These cells are G protein-coupled receptors and expressed in the cell membranes of olfactory sensory neurons. Once a cell is activated by a ligand, it initiates a signal transduction cascade that produces a nerve impulse to the brain where odor perception is processed. Vertebrate olfactory evolution is characterized by birth-and-death events, a special case of the stochastic continuous time Markov process. Vertebrate fish have three general types of receptor cells (two dedicated to pheromones). Terrestrial animals have different epithelial biology due to the specialized adaptation to detecting airborne odors. Two general classes of olfactory receptor gene reflect the vertebrate marine heritage (Class I) and the derived amphibian, reptile, and mammal terrestrial heritage (Class II). While we know much about olfactory receptor cells, there are still areas where our knowledge is insufficient, such as intra-individual diversity throughout the life time, epigenetic processes acting on olfactory receptors, and association of ligands to specific cells.

The Cryptococcus neoformans Capsule: a Sword and a Shield

PubMed Central

O'Meara, Teresa R.

2012-01-01

Summary: The human fungal pathogen Cryptococcus neoformans is characterized by its ability to induce a distinct polysaccharide capsule in response to a number of host-specific environmental stimuli. The induction of capsule is a complex biological process encompassing regulation at multiple steps, including the biosynthesis, transport, and maintenance of the polysaccharide at the cell surface. By precisely regulating the composition of its cell surface and secreted polysaccharides, C. neoformans has developed intricate ways to establish chronic infection and dormancy in the human host. The plasticity of the capsule structure in response to various host conditions also underscores the complex relationship between host and parasite. Much of this precise regulation of capsule is achieved through the transcriptional responses of multiple conserved signaling pathways that have been coopted to regulate this C. neoformans-specific virulence-associated phenotype. This review focuses on specific host stimuli that trigger the activation of the signal transduction cascades and on the downstream transcriptional responses that are required for robust encapsulation around the cell. PMID:22763631

Targeting the fibroblast growth factor receptors for the treatment of cancer.

PubMed

Lemieux, Steven M; Hadden, M Kyle

2013-06-01

Receptor tyrosine kinases (RTKs) are transmembrane proteins that play a critical role in stimulating signal transduction cascades to influence cell proliferation, growth, and differentiation and they have also been shown to promote angiogenesis when they are up-regulated or mutated. For this reason, their dysfunction has been implicated in the development of human cancer. Over the past decade, much attention has been devoted to developing inhibitors and antibodies against several classes of RTKs, including vascular endothelial growth factor receptors (VEGFRs), epidermal growth factor receptors (EGFRs), and platelet-derived growth factor receptors (PDGFRs). More recently, interest in the fibroblast growth factor receptor (FGFR) class of RTKs as a drug target for the treatment of cancer has emerged. Signaling through FGFRs is critical for normal cellular function and their dysregulation has been linked to various malignancies such as breast and prostate cancer. This review will focus on the current state of both small molecules and antibodies as FGFR inhibitors to provide insight into their development and future potential as anti-cancer agents.

How vertebrate and invertebrate visual pigments differ in their mechanism of photoactivation

PubMed Central

Nakagawa, Masashi; Iwasa, Tatsuo; Kikkawa, Satoshi; Tsuda, Motoyuki; Ebrey, Thomas G.

1999-01-01

In vertebrate visual pigments, a glutamic acid serves as a negative counterion to the positively charged chromophore, a protonated Schiff base of retinal. When photoisomerization leads to the Schiff base deprotonating, the anionic glutamic acid becomes protonated, forming a neutral species that activates the visual cascade. We show that in octopus rhodopsin, the glutamic acid has no anionic counterpart. Thus, the “counterion” is already neutral, so no protonated form of an initially anionic group needs to be created to activate. This helps to explain another observation—that the active photoproduct of octopus rhodopsin can be formed without its Schiff base deprotonating. In this sense, the mechanism of light activation of octopus rhodopsin is simpler than for vertebrates, because it eliminates one of the steps required for vertebrate rhodopsins to achieve their activating state. PMID:10339563

Streakline flow visualization study of a horseshoe vortex in a large-scale, two-dimensional turbine stator cascade

NASA Technical Reports Server (NTRS)

Gaugler, R. E.; Russell, L. M.

1980-01-01

Neutrally buoyant helium-filled bubbles were observed as they followed the streamlines in a horseshoe vortex system around the vane leading edge in a large-scale, two-dimensional, turbine stator cascade. Bubbles were introduced into the endwall boundary layer through a slot upstream of the vane leading edge. The paths of the bubbles were recorded photographically as streaklines on 16-mm movie film. Individual frames from the film have been selected, and overlayed to show the details of the horseshoe vortex around the leading edge. The transport of the vortex across the passage near the leading edge is clearly seen when compared to the streaks formed by bubbles carried in the main stream. Limiting streamlines on the endwall surface were traced by the flow of oil drops.

Integrated Circuits for Rapid Sample Processing and Electrochemical Detection of Biomarkers

NASA Astrophysics Data System (ADS)

Besant, Justin

The trade-off between speed and sensitivity of detection is a fundamental challenge in the design of point-of-care diagnostics. As the relevant molecules in many diseases exist natively at extremely low levels, many gold-standard diagnostic tests are designed with high sensitivity at the expense of long incubations needed to amplify the target analytes. The central aim of this thesis is to design new strategies to detect biologically relevant analytes with both high speed and sensitivity. The response time of a biosensor is limited by the ability of the target analyte to accumulate to detectable levels at the sensor surface. We overcome this limitation by designing a range of integrated devices to optimize the flux of the analyte to the sensor by increasing the effective analyte concentration, shortening the required diffusion distance, and confining the analyte in close proximity to the sensor. We couple these devices with novel ultrasensitive electrochemical transduction strategies to convert rare analytes into a detectable signal. We showcase the clinical utility of these approaches with several applications including cancer diagnosis, bacterial identification, and antibiotic susceptibility profiling. We design and optimize a device to isolate rare cancer cells from the bloodstream with near 100% efficiency and 10 000-fold specificity. We analyse pathogen specific nucleic acids by lysing bacteria in close proximity to an electrochemical sensor and find that this approach has 10-fold higher sensitivity than standard lysis in bulk solution. We design an electronic chip to readout the antibiotic susceptibility profile with an hour-long incubation by concentrating bacteria into nanoliter chambers with integrated electrodes. Finally, we report a strategy for ultrasensitive visual readout of nucleic acids as low as 100 fM within 10 minutes using an amplification cascade. The strategies presented could guide the development of fast, sensitive and low-cost diagnostics for diseases not previously detectable at the point-of-care.

Inherited macular degeneration-associated mutations in CNGB3 increase the ligand sensitivity and spontaneous open probability of cone cyclic nucleotide-gated channels

PubMed Central

Meighan, Peter C.; Peng, Changhong; Varnum, Michael D.

2015-01-01

Cyclic nucleotide gated (CNG) channels are a critical component of the visual transduction cascade in the vertebrate retina. Mutations in the genes encoding these channels have been associated with a spectrum of inherited retinal disorders. To gain insight into their pathophysiological mechanisms, we have investigated the functional consequences of several CNGB3 mutations, previously associated with macular degeneration (Y469D and L595F) or complete achromatopsia (S156F, P309L, and G558C), by expressing these subunits in combination with wild-type CNGA3 in Xenopus oocytes and characterizing them using patch-clamp recordings in the inside-out configuration. These mutations did not prevent the formation of functional heteromeric channels, as indicated by sensitivity to block by L-cis-diltiazem. With the exception of S156F, each of the mutant channels displayed electrophysiological properties reflecting enhanced channel activity at physiological concentrations of cGMP (i.e., a gain-of-function phenotype). The increased channel activity produced by these mutations resulted from either increased functional expression levels, or increased sensitivity to cyclic nucleotides. Furthermore, L595F increased the spontaneous open probability in the absence of activating ligand, signifying a ligand independent gain-of-function change. In addition to the CNGB3 disease-associate mutations, we characterized the effects of several common CNGB3 and CNGA3 single-nucleotide polymorphisms (SNPs) on heteromeric CNGA3+CNGB3 channel function. Two of the SNPs examined (A3-T153M, and B3-W234C) produced decreased ligand sensitivity for heteromeric CNG channels. These changes may contribute to background disease susceptibility when combined with other genetic or non-genetic factors. Together, these studies help to define the underlying molecular phenotype for mutations relating to CNG channel disease pathogenesis. PMID:26106334

Investigation of Positively Curved Blade in Compressor Cascade Based on Transition Model

NASA Astrophysics Data System (ADS)

Chen, Shaowen; Lan, Yunhe; Zhou, Zhihua; Wang, Songtao

2016-06-01

Experiment and numerical simulation of flow transition in a compressor cascade with positively curved blade is carried out in a low speed. In the experimental investigation, the outlet aerodynamic parameters are measured using a five-hole aerodynamic probe, and an ink-trace flow visualization is applied to the cascade surface. The effects of transition flow on the boundary layer development, three-dimensional flow separation and aerodynamic performance are studied. The feasibility of a commercial computational fluid dynamic code is validated and the numerical results show a good agreement with experimental data. The blade-positive curving intensifies the radial force from the endwalls to the mid-span near the suction surface, which leads to the smaller scope of the intermittent region, the lesser extents of turbulence intensity and the shorter radial height of the separation bubble near the endwalls, but has little influence on the flow near the mid-span. The large passage vortex is divided into two smaller shedding vortexes under the impact of the radial pressure gradient due to the positively curved blade. The new concentrated shedding vortex results in an increase in the turbulence intensity and secondary flow loss of the corresponding region.

CRISPR-Cas-Mediated Phage Resistance Enhances Horizontal Gene Transfer by Transduction.

PubMed

Watson, Bridget N J; Staals, Raymond H J; Fineran, Peter C

2018-02-13

A powerful contributor to prokaryotic evolution is horizontal gene transfer (HGT) through transformation, conjugation, and transduction, which can be advantageous, neutral, or detrimental to fitness. Bacteria and archaea control HGT and phage infection through CRISPR-Cas (clustered regularly interspaced short palindromic repeats-CRISPR-associated proteins) adaptive immunity. Although the benefits of resisting phage infection are evident, this can come at a cost of inhibiting the acquisition of other beneficial genes through HGT. Despite the ability of CRISPR-Cas to limit HGT through conjugation and transformation, its role in transduction is largely overlooked. Transduction is the phage-mediated transfer of bacterial DNA between cells and arguably has the greatest impact on HGT. We demonstrate that in Pectobacterium atrosepticum , CRISPR-Cas can inhibit the transduction of plasmids and chromosomal loci. In addition, we detected phage-mediated transfer of a large plant pathogenicity genomic island and show that CRISPR-Cas can inhibit its transduction. Despite these inhibitory effects of CRISPR-Cas on transduction, its more common role in phage resistance promotes rather than diminishes HGT via transduction by protecting bacteria from phage infection. This protective effect can also increase transduction of phage-sensitive members of mixed populations. CRISPR-Cas systems themselves display evidence of HGT, but little is known about their lateral dissemination between bacteria and whether transduction can contribute. We show that, through transduction, bacteria can acquire an entire chromosomal CRISPR-Cas system, including cas genes and phage-targeting spacers. We propose that the positive effect of CRISPR-Cas phage immunity on enhancing transduction surpasses the rarer cases where gene flow by transduction is restricted. IMPORTANCE The generation of genetic diversity through acquisition of DNA is a powerful contributor to microbial evolution and occurs through transformation, conjugation, and transduction. Of these, transduction, the phage-mediated transfer of bacterial DNA, is arguably the major route for genetic exchange. CRISPR-Cas adaptive immune systems control gene transfer by conjugation and transformation, but transduction has been mostly overlooked. Our results indicate that CRISPR-Cas can impede, but typically enhances the transduction of plasmids, chromosomal genes, and pathogenicity islands. By limiting wild-type phage replication, CRISPR-Cas immunity increases transduction in both phage-resistant and -sensitive members of mixed populations. Furthermore, we demonstrate mobilization of a chromosomal CRISPR-Cas system containing phage-targeting spacers by generalized transduction, which might partly account for the uneven distribution of these systems in nature. Overall, the ability of CRISPR-Cas to promote transduction reveals an unexpected impact of adaptive immunity on horizontal gene transfer, with broader implications for microbial evolution. Copyright © 2018 Watson et al.

Vascular Cell Induction Culture System Using Arabidopsis Leaves (VISUAL) Reveals the Sequential Differentiation of Sieve Element-Like Cells.

PubMed

Kondo, Yuki; Nurani, Alif Meem; Saito, Chieko; Ichihashi, Yasunori; Saito, Masato; Yamazaki, Kyoko; Mitsuda, Nobutaka; Ohme-Takagi, Masaru; Fukuda, Hiroo

2016-06-01

Cell differentiation is a complex process involving multiple steps, from initial cell fate specification to final differentiation. Procambial/cambial cells, which act as vascular stem cells, differentiate into both xylem and phloem cells during vascular development. Recent studies have identified regulatory cascades for xylem differentiation. However, the molecular mechanism underlying phloem differentiation is largely unexplored due to technical challenges. Here, we established an ectopic induction system for phloem differentiation named Vascular Cell Induction Culture System Using Arabidopsis Leaves (VISUAL). Our results verified similarities between VISUAL-induced Arabidopsis thaliana phloem cells and in vivo sieve elements. We performed network analysis using transcriptome data with VISUAL to dissect the processes underlying phloem differentiation, eventually identifying a factor involved in the regulation of the master transcription factor gene APL Thus, our culture system opens up new avenues not only for genetic studies of phloem differentiation, but also for future investigations of multidirectional differentiation from vascular stem cells. © 2016 American Society of Plant Biologists. All rights reserved.

HOG MAP kinase regulation of alternariol biosynthesis in Alternaria alternata is important for substrate colonization.

PubMed

Graf, Eva; Schmidt-Heydt, Markus; Geisen, Rolf

2012-07-16

Strains of the genus Alternaria are ubiquitously present and frequently found on fruits, vegetables and cereals. One of the most commonly found species from this genus is A. alternata which is able to produce the mycotoxin alternariol among others. To date only limited knowledge is available about the regulation of the biosynthesis of alternariol, especially under conditions relevant to food. Tomatoes are a typical substrate of A. alternata and have a high water activity. On the other hand cereals with moderate water activity are also frequently colonized by A. alternata. In the current analysis it was demonstrated that even minor changes in the osmotic status of the substrate affect the alternariol biosynthesis of strains from vegetables resulting in nearly complete inhibition. High osmolarity in the environment is usually transmitted to the transcriptional level of downstream regulated genes by the HOG signal cascade (high osmolarity glycerol cascade) which is a MAP kinase transduction pathway. The phosphorylation status of the A. alternata HOG (AaHOG) was determined. Various concentrations of NaCl induce the phosphorylation of AaHOG in a concentration, time and strain dependent manner. A strain with a genetically inactivated aahog gene was no longer able to produce alternariol indicating that the activity of the aahog gene is required for alternariol biosynthesis. Further experiments revealed that the biosynthesis of alternariol is important for the fungus to colonize tomato tissue. The tight water activity dependent regulation of alternariol biosynthesis ensures alternariol biosynthesis at conditions which indicate an optimal colonization substrate for the fungus. Copyright © 2012 Elsevier B.V. All rights reserved.

Role of Akt signaling in resistance to DNA-targeted therapy

PubMed Central

Avan, Abolfazl; Narayan, Ravi; Giovannetti, Elisa; Peters, Godefridus J

2016-01-01

The Akt signal transduction pathway controls most hallmarks of cancer. Activation of the Akt cascade promotes a malignant phenotype and is also widely implicated in drug resistance. Therefore, the modulation of Akt activity is regarded as an attractive strategy to enhance the efficacy of cancer therapy and irradiation. This pathway consists of phosphatidylinositol 3 kinase (PI3K), mammalian target of rapamycin, and the transforming serine-threonine kinase Akt protein isoforms, also known as protein kinase B. DNA-targeted agents, such as platinum agents, taxanes, and antimetabolites, as well as radiation have had a significant impact on cancer treatment by affecting DNA replication, which is aberrantly activated in malignancies. However, the caveat is that they may also trigger the activation of repairing mechanisms, such as upstream and downstream cascade of Akt survival pathway. Thus, each target can theoretically be inhibited in view of improving the potency of conventional treatment. Akt inhibitors, e.g., MK-2206 and perifosine, or PI3K modulators, e.g., LY294002 and Wortmannin, have shown some promising results in favor of sensitizing the cancer cells to the therapy in vitro and in vivo, which have provided the rationale for incorporation of these novel agents into multimodality treatment of different malignancies. Nevertheless, despite the acceptable safety profile of some of these agents in the clinical studies, with regard to the efficacy, the results are still too preliminary. Hence, we need to wait for the upcoming data from the ongoing trials before utilizing them into the standard care of cancer patients. PMID:27777878

Rofecoxib modulates multiple gene expression pathways in a clinical model of acute inflammatory pain

PubMed Central

Wang, Xiao-Min; Wu, Tian-Xia; Hamza, May; Ramsay, Edward S.; Wahl, Sharon M.; Dionne, Raymond A.

2007-01-01

New insights into the biological properties of cyclooxygenase-2 (COX-2) and its response pathway challenge the hypothesis that COX-2 is simply pro-inflammatory and inhibition of COX-2 solely prevents the development of inflammation and ameliorates inflammatory pain. The present study performed a comprehensive analysis of gene/protein expression induced by a selective inhibitor of COX-2, rofecoxib, compared with a non-selective COX inhibitor, ibuprofen, and placebo in a clinical model of acute inflammatory pain (the surgical extraction of impacted third molars) using microarray analysis followed by quantitative RT-PCR verification and Western blotting. Inhibition of COX-2 modulated gene expression related to inflammation and pain, the arachidonic acid pathway, apoptosis/angiogenesis, cell adhesion and signal transduction. Compared to placebo, rofecoxib treatment increased the gene expression of ANXA3 (annexin 3), SOD2 (superoxide dismutase 2), SOCS3 (suppressor of cytokine signaling 3) and IL1RN (IL1 receptor antagonist) which are associated with inhibition of phospholipase A2 and suppression of cytokine signaling cascades, respectively. Both rofecoxib and ibuprofen treatment increased the gene expression of the pro-inflammatory mediators, IL6 and CCL2 (chemokine C-C motif ligand 2), following tissue injury compared to the placebo treatment. These results indicate a complex role for COX-2 in the inflammatory cascade in addition to the well-characterized COX-dependent pathway, as multiple pathways are also involved in rofecoxib-induced anti-inflammatory and analgesic effects at the gene expression level. These findings may also suggest an alternative hypothesis for the adverse effects attributed to selective inhibition of COX-2. PMID:17070997

Leptin attenuates BACE1 expression and Amyloid-β genesis via the activation of SIRT1 signaling pathway

PubMed Central

Marwarha, Gurdeep; Raza, Shaneabbas; Meiers, Craig; Ghribi, Othman

2014-01-01

The aspartyl protease β-site AβPP-cleaving enzyme 1 (BACE1) catalyzes the rate-limiting step in Aβ production, a peptide at the nexus of neurodegenerative cascades in Alzheimer Disease (AD). The adipocytokine leptin has been demonstrated to reduce Aβ production and decrease BACE1 activity and expression levels. However, the signaling cascades involved in the leptin-induced mitigation in Aβ levels and BACE1 expression levels have not been elucidated. We have demonstrated that the transcription factor nuclear factor – kappa B (NF-κB) positively regulates BACE1 transcription. NF-κB activity is tightly regulated by the mammalian sirtuin SIRT1. Multiple studies have cogently evinced that leptin activates the metabolic master regulator SIRT1. In this study, we determined the extent to which SIRT1 expression and activity regulate the leptin-induced attenuation in BACE1 expression and Aβ levels in cultured human neuroblastoma SH-SY5Y cells. This study also elucidated and delineated the signal transduction pathways involved in the leptin induced mitigation in BACE1 expression. Our results demonstrate for the first time that leptin attenuates the activation and transcriptional activity of NF-κB by reducing the acetylation of the p65 subunit in a SIRT1-dependent manner. Furthermore, our data shows that leptin reduces the NF-κB – mediated transcription of BACE1 and consequently reduces Amyloid-β genesis. Our study provides a valuable insight and a novel mechanism by which leptin reduces BACE1 expression and Amyloid-β production and may help design potential therapeutic interventions. PMID:24874077

Cholinergic urethral brush cells are widespread throughout placental mammals.

PubMed

Deckmann, Klaus; Krasteva-Christ, Gabriela; Rafiq, Amir; Herden, Christine; Wichmann, Judy; Knauf, Sascha; Nassenstein, Christina; Grevelding, Christoph G; Dorresteijn, Adriaan; Chubanov, Vladimir; Gudermann, Thomas; Bschleipfer, Thomas; Kummer, Wolfgang

2015-11-01

We previously identified a population of cholinergic epithelial cells in murine, human and rat urethrae that exhibits a structural marker of brush cells (villin) and expresses components of the canonical taste transduction signaling cascade (α-gustducin, phospholipase Cβ2 (PLCβ2), transient receptor potential cation channel melanostatin 5 (TRPM5)). These cells serve as sentinels, monitoring the chemical composition of the luminal content for potentially hazardous compounds such as bacteria, and initiate protective reflexes counteracting further ingression. In order to elucidate cross-species conservation of the urethral chemosensory pathway we investigated the occurrence and molecular make-up of urethral brush cells in placental mammals. We screened 11 additional species, at least one in each of the five mammalian taxonomic units primates, carnivora, perissodactyla, artiodactyla and rodentia, for immunohistochemical labeling of the acetylcholine synthesizing enzyme, choline acetyltransferase (ChAT), villin, and taste cascade components (α-gustducin, PLCβ2, TRPM5). Corresponding to findings in previously investigated species, urethral epithelial cells with brush cell shape were immunolabeled in all 11 mammals. In 8 species, immunoreactivities against all marker proteins and ChAT were observed, and double-labeling immunofluorescence confirmed the cholinergic nature of villin-positive and chemosensory (TRPM5-positive) cells. In cat and horse, these cells were not labeled by the ChAT antiserum used in this study, and unspecific reactions of the secondary antiserum precluded conclusions about ChAT-expression in the bovine epithelium. These data indicate that urethral brush cells are widespread throughout the mammalian kingdom and evolved not later than about 64.5millionyears ago. Copyright © 2015 Elsevier B.V. All rights reserved.

Parallel neural pathways in higher visual centers of the Drosophila brain that mediate wavelength-specific behavior

PubMed Central

Otsuna, Hideo; Shinomiya, Kazunori; Ito, Kei

2014-01-01

Compared with connections between the retinae and primary visual centers, relatively less is known in both mammals and insects about the functional segregation of neural pathways connecting primary and higher centers of the visual processing cascade. Here, using the Drosophila visual system as a model, we demonstrate two levels of parallel computation in the pathways that connect primary visual centers of the optic lobe to computational circuits embedded within deeper centers in the central brain. We show that a seemingly simple achromatic behavior, namely phototaxis, is under the control of several independent pathways, each of which is responsible for navigation towards unique wavelengths. Silencing just one pathway is enough to disturb phototaxis towards one characteristic monochromatic source, whereas phototactic behavior towards white light is not affected. The response spectrum of each demonstrable pathway is different from that of individual photoreceptors, suggesting subtractive computations. A choice assay between two colors showed that these pathways are responsible for navigation towards, but not for the detection itself of, the monochromatic light. The present study provides novel insights about how visual information is separated and processed in parallel to achieve robust control of an innate behavior. PMID:24574974

Metarhodopsin control by arrestin, light-filtering screening pigments, and visual pigment turnover in invertebrate microvillar photoreceptors.

PubMed

Stavenga, Doekele G; Hardie, Roger C

2011-03-01

The visual pigments of most invertebrate photoreceptors have two thermostable photo-interconvertible states, the ground state rhodopsin and photo-activated metarhodopsin, which triggers the phototransduction cascade until it binds arrestin. The ratio of the two states in photoequilibrium is determined by their absorbance spectra and the effective spectral distribution of illumination. Calculations indicate that metarhodopsin levels in fly photoreceptors are maintained below ~35% in normal diurnal environments, due to the combination of a blue-green rhodopsin, an orange-absorbing metarhodopsin and red transparent screening pigments. Slow metarhodopsin degradation and rhodopsin regeneration processes further subserve visual pigment maintenance. In most insect eyes, where the majority of photoreceptors have green-absorbing rhodopsins and blue-absorbing metarhodopsins, natural illuminants are predicted to create metarhodopsin levels greater than 60% at high intensities. However, fast metarhodopsin decay and rhodopsin regeneration also play an important role in controlling metarhodopsin in green receptors, resulting in a high rhodopsin content at low light intensities and a reduced overall visual pigment content in bright light. A simple model for the visual pigment-arrestin cycle is used to illustrate the dependence of the visual pigment population states on light intensity, arrestin levels and pigment turnover.

Mechanotransduction signaling in podocytes from fluid flow shear stress.

PubMed

Srivastava, Tarak; Dai, Hongying; Heruth, Daniel P; Alon, Uri S; Garola, Robert E; Zhou, Jianping; Duncan, R Scott; El-Meanawy, Ashraf; McCarthy, Ellen T; Sharma, Ram; Johnson, Mark L; Savin, Virginia J; Sharma, Mukut

2018-01-01

Recently, we and others have found that hyperfiltration-associated increase in biomechanical forces, namely, tensile stress and fluid flow shear stress (FFSS), can directly and distinctly alter podocyte structure and function. The ultrafiltrate flow over the major processes and cell body generates FFSS to podocytes. Our previous work suggests that the cyclooxygenase-2 (COX-2)-PGE 2 -PGE 2 receptor 2 (EP2) axis plays an important role in mechanoperception of FFSS in podocytes. To address mechanotransduction of the perceived stimulus through EP2, cultured podocytes were exposed to FFSS (2 dyn/cm 2 ) for 2 h. Total RNA from cells at the end of FFSS treatment, 2-h post-FFSS, and 24-h post-FFSS was used for whole exon array analysis. Differentially regulated genes ( P < 0.01) were analyzed using bioinformatics tools Enrichr and Ingenuity Pathway Analysis to predict pathways/molecules. Candidate pathways were validated using Western blot analysis and then further confirmed to be resulting from a direct effect of PGE 2 on podocytes. Results show that FFSS-induced mechanotransduction as well as exogenous PGE 2 activate the Akt-GSK3β-β-catenin (Ser552) and MAPK/ERK but not the cAMP-PKA signal transduction cascades. These pathways are reportedly associated with FFSS-induced and EP2-mediated signaling in other epithelial cells as well. The current regimen for treating hyperfiltration-mediated injury largely depends on targeting the renin-angiotensin-aldosterone system. The present study identifies specific transduction mechanisms and provides novel information on the direct effect of FFSS on podocytes. These results suggest that targeting EP2-mediated signaling pathways holds therapeutic significance for delaying progression of chronic kidney disease secondary to hyperfiltration.

Genome-Wide Survey and Expression Profile Analysis of the Mitogen-Activated Protein Kinase (MAPK) Gene Family in Brassica rapa

PubMed Central

Yu, Hao; Qu, Cunmin; Tang, Zhanglin; Li, Jiana; Chai, Yourong; Liang, Ying

2015-01-01

Mitogen-activated protein kinase (MAPK) cascades are fundamental signal transduction modules in plants, controlling cell division, development, hormone signaling, and biotic and abiotic stress responses. Although MAPKs have been investigated in several plant species, a comprehensive analysis of the MAPK gene family has hitherto not been performed in Brassica rapa. In this study, we identified 32 MAPKs in the B. rapa genome by conducting BLASTP and syntenic block analyses, and screening for the essential signature motif (TDY or TEY) of plant MAPK proteins. Of the 32 BraMAPK genes retrieved from the Brassica Database, 13 exhibited exon splicing errors, excessive splicing of the 5' sequence, excessive retention of the 5' sequence, and sequencing errors of the 3' end. Phylogenetic trees of the 32 corrected MAPKs from B. rapa and of MAPKs from other plants generated by the neighbor-joining and maximum likelihood methods suggested that BraMAPKs could be divided into four groups (groups A, B, C, and D). Gene number expansion was observed for BraMAPK genes in groups A and D, which may have been caused by the tandem duplication and genome triplication of the ancestral genome of the Brassica progenitor. Except for five members of the BraMAPK10 subfamily, the identified BraMAPKs were expressed in most of the tissues examined, including callus, root, stem, leaf, flower, and silique. Quantitative real-time PCR demonstrated that at least six and five BraMAPKs were induced or repressed by various abiotic stresses and hormone treatments, respectively, suggesting their potential roles in the abiotic stress response and various hormone signal transduction pathways in B. rapa. This study provides valuable insight into the putative physiological and biochemical functions of MAPK genes in B. rapa. PMID:26173020

Mitogenic signals and transforming potential of Nyk, a newly identified neural cell adhesion molecule-related receptor tyrosine kinase.

PubMed Central

Ling, L; Kung, H J

1995-01-01

Nyk/Mer is a recently identified receptor tyrosine kinase with neural cell adhesion molecule-like structure (two immunoglobulin G-like domains and two fibronectin III-like domains) in its extracellular region and belongs to the Ufo/Axl family of receptors. The ligand for Nyk/Mer is presently unknown, as are the signal transduction pathways mediated by this receptor. We constructed and expressed a chimeric receptor (Fms-Nyk) composed of the extracellular domain of the human colony-stimulating factor 1 receptor (Fms) and the transmembrane and cytoplasmic domains of human Nyk/Mer in NIH 3T3 fibroblasts in order to investigate the mitogenic signaling and biochemical properties of Nyk/Mer. Colony-stimulating factor 1 stimulation of the Fms-Nyk chimeric receptor in transfected NIH 3T3 fibroblasts leads to a transformed phenotype and generates a proliferative response in the absence of other growth factors. We show that phospholipase C gamma, phosphatidylinositol 3-kinase/p70 S6 kinase, Shc, Grb2, Raf-1, and mitogen-activated protein kinase are downstream components of the Nyk/Mer signal transduction pathways. In addition, Nyk/Mer weakly activates p90rsk, while stress-activated protein kinase, Ras GTPase-activating protein (GAP), and GAP-associated p62 and p190 proteins are not activated or tyrosine phosphorylated by Nyk/Mer. An analysis comparing the Nyk/Mer signal cascade with that of the epidermal growth factor receptor indicates substrate preferences by these two receptors. Our results provide a detailed description of the Nyk/Mer signaling pathways. Given the structural similarity between the Ufo/Axl family receptors, some of the information may also be applied to other members of this receptor tyrosine kinase family. PMID:8524223

Cell envelope stress in mycobacteria is regulated by the novel signal transduction ATPase IniR in response to trehalose

PubMed Central

van Winden, Vincent J. C.; Sparrius, Marion; van de Weerd, Robert; Speer, Alexander; Ummels, Roy; Sherman, David R.

2017-01-01

The cell envelope of mycobacteria is a highly unique and complex structure that is functionally equivalent to that of Gram-negative bacteria to protect the bacterial cell. Defects in the integrity or assembly of this cell envelope must be sensed to allow the induction of stress response systems. The promoter that is specifically and most strongly induced upon exposure to ethambutol and isoniazid, first line drugs that affect cell envelope biogenesis, is the iniBAC promoter. In this study, we set out to identify the regulator of the iniBAC operon in Mycobacterium marinum using an unbiased transposon mutagenesis screen in a constitutively iniBAC-expressing mutant background. We obtained multiple mutants in the mce1 locus as well as mutants in an uncharacterized putative transcriptional regulator (MMAR_0612). This latter gene was shown to function as the iniBAC regulator, as overexpression resulted in constitutive iniBAC induction, whereas a knockout mutant was unable to respond to the presence of ethambutol and isoniazid. Experiments with the M. tuberculosis homologue (Rv0339c) showed identical results. RNAseq experiments showed that this regulatory gene was exclusively involved in the regulation of the iniBAC operon. We therefore propose to name this dedicated regulator iniBAC Regulator (IniR). IniR belongs to the family of signal transduction ATPases with numerous domains, including a putative sugar-binding domain. Upon testing different sugars, we identified trehalose as an activator and metabolic cue for iniBAC activation, which could also explain the effect of the mce1 mutations. In conclusion, cell envelope stress in mycobacteria is regulated by IniR in a cascade that includes trehalose. PMID:29281637

Toward high fidelity spectral sensing and RF signal processing in silicon photonic and nano-opto-mechanical platforms

NASA Astrophysics Data System (ADS)

Siddiqui, Aleem; Reinke, Charles; Shin, Heedeuk; Jarecki, Robert L.; Starbuck, Andrew L.; Rakich, Peter

2017-05-01

The performance of electronic systems for radio-frequency (RF) spectrum analysis is critical for agile radar and communications systems, ISR (intelligence, surveillance, and reconnaissance) operations in challenging electromagnetic (EM) environments, and EM-environment situational awareness. While considerable progress has been made in size, weight, and power (SWaP) and performance metrics in conventional RF technology platforms, fundamental limits make continued improvements increasingly difficult. Alternatively, we propose employing cascaded transduction processes in a chip-scale nano-optomechanical system (NOMS) to achieve a spectral sensor with exceptional signal-linearity, high dynamic range, narrow spectral resolution and ultra-fast sweep times. By leveraging the optimal capabilities of photons and phonons, the system we pursue in this work has performance metrics scalable well beyond the fundamental limitations inherent to all electronic systems. In our device architecture, information processing is performed on wide-bandwidth RF-modulated optical signals by photon-mediated phononic transduction of the modulation to the acoustical-domain for narrow-band filtering, and then back to the optical-domain by phonon-mediated phase modulation (the reverse process). Here, we rely on photonics to efficiently distribute signals for parallel processing, and on phononics for effective and flexible RF-frequency manipulation. This technology is used to create RF-filters that are insensitive to the optical wavelength, with wide center frequency bandwidth selectivity (1-100GHz), ultra-narrow filter bandwidth (1-100MHz), and high dynamic range (70dB), which we will present. Additionally, using this filter as a building block, we will discuss current results and progress toward demonstrating a multichannel-filter with a bandwidth of < 10MHz per channel, while minimizing cumulative optical/acoustic/optical transduced insertion-loss to ideally < 10dB. These proposed metric represent significant improvements over RF-platforms.

A Dynamic Response Regulator Protein Modulates G-Protein–Dependent Polarity in the Bacterium Myxococcus xanthus

PubMed Central

Zhang, Yong; Guzzo, Mathilde; Ducret, Adrien; Li, Yue-Zhong; Mignot, Tâm

2012-01-01

Migrating cells employ sophisticated signal transduction systems to respond to their environment and polarize towards attractant sources. Bacterial cells also regulate their polarity dynamically to reverse their direction of movement. In Myxococcus xanthus, a GTP-bound Ras-like G-protein, MglA, activates the motility machineries at the leading cell pole. Reversals are provoked by pole-to-pole switching of MglA, which is under the control of a chemosensory-like signal transduction cascade (Frz). It was previously known that the asymmetric localization of MglA at one cell pole is regulated by MglB, a GTPase Activating Protein (GAP). In this process, MglB specifically localizes at the opposite lagging cell pole and blocks MglA localization at that pole. However, how MglA is targeted to the leading pole and how Frz activity switches the localizations of MglA and MglB synchronously remained unknown. Here, we show that MglA requires RomR, a previously known response regulator protein, to localize to the leading cell pole efficiently. Specifically, RomR-MglA and RomR-MglB complexes are formed and act complementarily to establish the polarity axis, segregating MglA and MglB to opposite cell poles. Finally, we present evidence that Frz signaling may regulate MglA localization through RomR, suggesting that RomR constitutes a link between the Frz-signaling and MglAB polarity modules. Thus, in Myxococcus xanthus, a response regulator protein governs the localization of a small G-protein, adding further insight to the polarization mechanism and suggesting that motility regulation evolved by recruiting and combining existing signaling modules of diverse origins. PMID:22916026

A novel siglec (CgSiglec-1) from the Pacific oyster (Crassostrea gigas) with broad recognition spectrum and inhibitory activity to apoptosis, phagocytosis and cytokine release.

PubMed

Liu, Conghui; Jiang, Shuai; Wang, Mengqiang; Wang, Lingling; Chen, Hao; Xu, Jiachao; Lv, Zhao; Song, Linsheng

2016-08-01

Sialic acid binding immunoglobulin-type lectin (siglec) belongs to the immunoglobulin superfamily (IgSF), which acts as regulator involved in glycan recognition and signal transduction in the immune and nervous systems. In the present study, a siglec gene (designated CgSiglec-1) was characterized from the Pacific oyster, Crassostrea gigas. The cDNA of CgSiglec-1 was of 1251 bp encoding a predicted polypeptide of 416 amino acids. CgSiglec-1 was composed of two I-set immunoglobulin (Ig) domains, one transmembrane (TM) domain and two ITIM motifs, sharing a sequence similarity with vertebrate CD22 homologs. The mRNA expression of CgSiglec-1 could be detected in all the selected tissues, with the highest level in hemocytes and labial palps. The confocal analysis revealed that CgSiglec-1 mainly distributed on the cytoplasmic membrane of the oyster hemocytes. In addition, the mRNA transcripts of CgSiglec-1 in hemocytes increased significantly (4.29-fold to that of control group, p < 0.05) after Vibrio splendidus stimulation. The recombinant CgSiglec-1 protein (rCgSiglec-1) could bind to poly sialic acid (pSIAS), lipopolysaccharides (LPS) and peptidoglycan (PGN) in a dose-dependent manner. The blockade of CgSiglec-1 by specific polyclonal antibodies could enhance the LPS-induced cell apoptosis, phagocytosis towards V. splendidus and the release of cytokines, such as CgTNF-1, CgIFNLP and CgIL-17. The results collectively indicated that CgSiglec-1 could act as a bridge molecule between invader recognition and signal transduction cascade, and modulate the immune response by inhibiting various important processes of immunity in oyster. Copyright © 2016 Elsevier Ltd. All rights reserved.

A food-derived synergist of NGF signaling: identification of protein tyrosine phosphatase 1B as a key regulator of NGF receptor-initiated signal transduction.

PubMed

Shibata, Takahiro; Nakahara, Hiroko; Kita, Narumi; Matsubara, Yui; Han, Chunguang; Morimitsu, Yasujiro; Iwamoto, Noriko; Kumagai, Yoshito; Nishida, Motohiro; Kurose, Hitoshi; Aoki, Naohito; Ojika, Makoto; Uchida, Koji

2008-12-01

Neurotrophins, such as the nerve growth factor (NGF), play an essential role in the growth, development, survival and functional maintenance of neurons in the central and peripheral systems. They also prevent neuronal cell death under various stressful conditions, such as ischemia and neurodegenerative disorders. NGF induces cell differentiation and neurite outgrowth by binding with and activating the NGF receptor tyrosine kinase followed by activation of a variety of signaling cascades. We have investigated the NGF-dependent neuritogenesis enhancer potential of a food-derived small molecule contained in Brassica vegetables and identified the protein tyrosine phosphatase (PTP) 1B as a key regulator of the NGF receptor-initiated signal transduction. Based on an extensive screening of Brassica vegetable extracts for the neuritogenic-promoting activity in the rat pheochromocytoma cell line PC12, we found the Japanese horseradish, wasabi (Wasabia japonica, syn. Eutrema wasabi), as the richest source and identified 6-methylsulfinylhexyl isothiocyanate (6-HITC), an analogue of sulforaphane isolated from broccoli, as one of the major neuritogenic enhancers in the wasabi. 6-HITC strongly enhanced the neurite outgrowth and neurofilament expression elicited by a low-concentration of NGF that alone was insufficient to induce neuronal differentiation. 6-HITC also facilitated the sustained-phosphorylation of the extracellular signal-regulated kinase and the autophosphorylation of the NGF receptor TrkA. It was found that PTP1B act as a phosphatase capable of dephosphorylating Tyr-490 of TrkA and was inactivated by 6-HITC in a redox-dependent manner. The identification of PTP1B as a regulator of NGF signaling may provide new clues about the chemoprotective potential of food components, such as isothiocyanates.

Nature of extracellular signal that triggers RhoA/ROCK activation for the basal internal anal sphincter tone in humans

PubMed Central

Singh, Jagmohan; Kumar, Sumit; Phillips, Benjamin

2015-01-01

The extracellular signal that triggers activation of rho-associated kinase (RhoA/ROCK), the major molecular determinant of basal internal anal sphincter (IAS) smooth muscle tone, is not known. Using human IAS tissues, we identified the presence of the biosynthetic machineries for angiotensin II (ANG II), thromboxane A2 (TXA2), and prostaglandin F2α (PGF2α). These end products of the renin-angiotensin system (RAS) (ANG II) and arachidonic acid (TXA2 and PGF2α) pathways and their effects in human IAS vs. rectal smooth muscle (RSM) were studied. A multipronged approach utilizing immunocytochemistry, Western blot analyses, and force measurements was implemented. Additionally, in a systematic analysis of the effects of respective inhibitors along different steps of biosynthesis and those of antagonists, their end products were evaluated either individually or in combination. To further describe the molecular mechanism for the IAS tone via these pathways, we monitored RhoA/ROCK activation and its signal transduction cascade. Data showed characteristically higher expression of biosynthetic machineries of RAS and AA pathways in the IAS compared with the RSM. Additionally, specific inhibition of the arachidonic acid (AA) pathway caused ∼80% decrease in the IAS tone, whereas that of RAS lead to ∼20% decrease. Signal transduction studies revealed that the end products of both AA and RAS pathways cause increase in the IAS tone via activation of RhoA/ROCK. Both AA and RAS (via the release of their end products TXA2, PGF2α, and ANG II, respectively), provide extracellular signals which activate RhoA/ROCK for the maintenance of the basal tone in human IAS. PMID:25882611

Aging-associated oxidative stress leads to decrease in IAS tone via RhoA/ROCK downregulation.

PubMed

Singh, Jagmohan; Kumar, Sumit; Krishna, Chadalavada Vijay; Rattan, Satish

2014-06-01

Internal anal sphincter (IAS) tone plays an important role in rectoanal incontinence (RI). IAS tone may be compromised during aging, leading to RI in certain patients. We examined the influence of oxidative stress in the aging-associated decrease in IAS tone (AADI). Using adult (4-6 mo old) and aging (24-30 mo old) rats, we determined the effect of oxidative stress on IAS tone and the regulatory RhoA/ROCK signal transduction cascade. We determined the effect of the oxidative stress inducer LY83583, which produces superoxide anions (O2 (·-)), on basal and stimulated IAS tone before and after treatment of intact smooth muscle strips and smooth muscle cells with the O2 (·-) scavenger SOD. Our data showed that AADI was associated with a decrease in RhoA/ROCK expression at the transcriptional and translational levels. Oxidative stress with a LY83583-mediated decrease in IAS tone and relaxation of IAS smooth muscle cells was associated with a decrease in RhoA/ROCK signal transduction, which was reversible by SOD. In addition, LY83583 caused a significant decrease in IAS contraction produced by the RhoA activator and a known RhoA/ROCK agonist, U46619, that was also reversible by SOD. The inhibitory effects of LY83583 and the ROCK inhibitor Y27632 on the U46619-induced increase in IAS tone were similar. We conclude that an increase in oxidative stress plays an important role in AADI in the elderly and may be one of the underlying mechanisms of RI in certain aging patients. Copyright © 2014 the American Physiological Society.

Aging-associated oxidative stress leads to decrease in IAS tone via RhoA/ROCK downregulation

PubMed Central

Singh, Jagmohan; Kumar, Sumit; Krishna, Chadalavada Vijay

2014-01-01

Internal anal sphincter (IAS) tone plays an important role in rectoanal incontinence (RI). IAS tone may be compromised during aging, leading to RI in certain patients. We examined the influence of oxidative stress in the aging-associated decrease in IAS tone (AADI). Using adult (4–6 mo old) and aging (24–30 mo old) rats, we determined the effect of oxidative stress on IAS tone and the regulatory RhoA/ROCK signal transduction cascade. We determined the effect of the oxidative stress inducer LY83583, which produces superoxide anions (O2·−), on basal and stimulated IAS tone before and after treatment of intact smooth muscle strips and smooth muscle cells with the O2·− scavenger SOD. Our data showed that AADI was associated with a decrease in RhoA/ROCK expression at the transcriptional and translational levels. Oxidative stress with a LY83583-mediated decrease in IAS tone and relaxation of IAS smooth muscle cells was associated with a decrease in RhoA/ROCK signal transduction, which was reversible by SOD. In addition, LY83583 caused a significant decrease in IAS contraction produced by the RhoA activator and a known RhoA/ROCK agonist, U46619, that was also reversible by SOD. The inhibitory effects of LY83583 and the ROCK inhibitor Y27632 on the U46619-induced increase in IAS tone were similar. We conclude that an increase in oxidative stress plays an important role in AADI in the elderly and may be one of the underlying mechanisms of RI in certain aging patients. PMID:24742984

The cytosolic domain of T-cell receptor ζ associates with membranes in a dynamic equilibrium and deeply penetrates the bilayer.

PubMed

Zimmermann, Kerstin; Eells, Rebecca; Heinrich, Frank; Rintoul, Stefanie; Josey, Brian; Shekhar, Prabhanshu; Lösche, Mathias; Stern, Lawrence J

2017-10-27

Interactions between lipid bilayers and the membrane-proximal regions of membrane-associated proteins play important roles in regulating membrane protein structure and function. The T-cell antigen receptor is an assembly of eight single-pass membrane-spanning subunits on the surface of T lymphocytes that initiates cytosolic signaling cascades upon binding antigens presented by MHC-family proteins on antigen-presenting cells. Its ζ-subunit contains multiple cytosolic immunoreceptor tyrosine-based activation motifs involved in signal transduction, and this subunit by itself is sufficient to couple extracellular stimuli to intracellular signaling events. Interactions of the cytosolic domain of ζ (ζ cyt ) with acidic lipids have been implicated in the initiation and regulation of transmembrane signaling. ζ cyt is unstructured in solution. Interaction with acidic phospholipids induces structure, but its disposition when bound to lipid bilayers is controversial. Here, using surface plasmon resonance and neutron reflection, we characterized the interaction of ζ cyt with planar lipid bilayers containing mixtures of acidic and neutral lipids. We observed two binding modes of ζ cyt to the bilayers in dynamic equilibrium: one in which ζ cyt is peripherally associated with lipid headgroups and one in which it penetrates deeply into the bilayer. Such an equilibrium between the peripherally bound and embedded forms of ζ cyt apparently controls accessibility of the immunoreceptor tyrosine-based activation signal transduction pathway. Our results reconcile conflicting findings of the ζ structure reported in previous studies and provide a framework for understanding how lipid interactions regulate motifs to tyrosine kinases and may regulate the T-cell antigen receptor biological activities for this cell-surface receptor system.

Altered Protein Interactions of the Endogenous Interactome of PTPIP51 towards MAPK Signaling

PubMed Central

Brobeil, Alexander; Chehab, Rajaa; Dietel, Eric; Gattenlöhner, Stefan; Wimmer, Monika

2017-01-01

Protein–protein interactions play a pivotal role in normal cellular functions as well as in carcinogenesis. The protein–protein interactions form functional clusters during signal transduction. To elucidate the fine calibration of the protein–protein interactions of protein tyrosine phosphatase interacting protein 51 (PTPIP51) a small molecule drug, namely LDC-3, directly targeting PTPIP51 is now available. Therefore, LDC-3 allows for the studying of the regulation of the endogenous interactome by modulating PTPIP51 binding capacity. Small interfering ribonucleic acid (siRNA) experiments show that the modification in PTPIP51 binding capacity is induced by LDC-3. Application of LDC-3 annuls the known regulatory phosphorylation mechanisms for PTPIP51 and consequently, significantly alters the assembly of the PTPIP51 associated protein complexes. The treatment of human keratinocytes (HaCaT cells) with LDC-3 induces an altered protein–protein interaction profile of the endogenous interactome of PTPIP51. In addition, LDC-3 stabilizes PTPIP51 within a mitogen activated protein kinase (MAPK) complex composed of Raf-1 and the scaffold protein 14-3-3, independent of the phosphorylation status of PTPIP51. Of note, under LDC-3 treatment the regulatory function of the PTP1B on PTPIP51 fails to impact the PTPIP51 interaction characteristics, as reported for the HaCaT cell line. In summary, LDC-3 gives the unique opportunity to directly modulate PTPIP51 in malignant cells, thus targeting potential dysregulated signal transduction pathways such as the MAPK cascade. The provided data give critical insights in the therapeutic potential of PTPIP51 protein interactions and thus are basic for possible targeted therapy regimens. PMID:28754031

Primate Torpor: Regulation of Stress-activated Protein Kinases During Daily Torpor in the Gray Mouse Lemur, Microcebus murinus.

PubMed

Biggar, Kyle K; Wu, Cheng-Wei; Tessier, Shannon N; Zhang, Jing; Pifferi, Fabien; Perret, Martine; Storey, Kenneth B

2015-04-01

Very few selected species of primates are known to be capable of entering torpor. This exciting discovery means that the ability to enter a natural state of dormancy is an ancestral trait among primates and, in phylogenetic terms, is very close to the human lineage. To explore the regulatory mechanisms that underlie primate torpor, we analyzed signal transduction cascades to discover those involved in coordinating tissue responses during torpor. The responses of mitogen-activated protein kinase (MAPK) family members to primate torpor were compared in six organs of control (aroused) versus torpid gray mouse lemurs, Microcebus murinus. The proteins examined include extracellular signal-regulated kinases (ERKs), c-jun NH2-terminal kinases (JNKs), MAPK kinase (MEK), and p38, in addition to stress-related proteins p53 and heat shock protein 27 (HSP27). The activation of specific MAPK signal transduction pathways may provide a mechanism to regulate the expression of torpor-responsive genes or the regulation of selected downstream cellular processes. In response to torpor, each MAPK subfamily responded differently during torpor and each showed organ-specific patterns of response. For example, skeletal muscle displayed elevated relative phosphorylation of ERK1/2 during torpor. Interestingly, adipose tissues showed the highest degree of MAPK activation. Brown adipose tissue displayed an activation of ERK1/2 and p38, whereas white adipose tissue showed activation of ERK1/2, p38, MEK, and JNK during torpor. Importantly, both adipose tissues possess specialized functions that are critical for torpor, with brown adipose required for non-shivering thermogenesis and white adipose utilized as the primary source of lipid fuel for torpor. Overall, these data indicate crucial roles of MAPKs in the regulation of primate organs during torpor. Copyright © 2015. Production and hosting by Elsevier Ltd.

Visual reliability and information rate in the retina of a nocturnal bee.

PubMed

Frederiksen, Rikard; Wcislo, William T; Warrant, Eric J

2008-03-11

Nocturnal animals relying on vision typically have eyes that are optically and morphologically adapted for both increased sensitivity and greater information capacity in dim light. Here, we investigate whether adaptations for increased sensitivity also are found in their photoreceptors by using closely related and fast-flying nocturnal and diurnal bees as model animals. The nocturnal bee Megalopta genalis is capable of foraging and homing by using visually discriminated landmarks at starlight intensities. Megalopta's near relative, Lasioglossum leucozonium, performs these tasks only in bright sunshine. By recording intracellular responses to Gaussian white-noise stimuli, we show that photoreceptors in Megalopta actually code less information at most light levels than those in Lasioglossum. However, as in several other nocturnal arthropods, Megalopta's photoreceptors possess a much greater gain of transduction, indicating that nocturnal photoreceptors trade information capacity for sensitivity. By sacrificing photoreceptor signal-to-noise ratio and information capacity in dim light for an increased gain and, thus, an increased sensitivity, this strategy can benefit nocturnal insects that use neural summation to improve visual reliability at night.

The Tuberculosis Cascade of Care in India’s Public Sector: A Systematic Review and Meta-analysis

PubMed Central

Satyanarayana, Srinath; Pai, Madhukar; Thomas, Beena E.; Chadha, Vineet K.; Swaminathan, Soumya; Mayer, Kenneth H.

2016-01-01

Background India has 23% of the global burden of active tuberculosis (TB) patients and 27% of the world’s “missing” patients, which includes those who may not have received effective TB care and could potentially spread TB to others. The “cascade of care” is a useful model for visualizing deficiencies in case detection and retention in care, in order to prioritize interventions. Methods and Findings The care cascade constructed in this paper focuses on the Revised National TB Control Programme (RNTCP), which treats about half of India’s TB patients. We define the TB cascade as including the following patient populations: total prevalent active TB patients in India, TB patients who reach and undergo evaluation at RNTCP diagnostic facilities, patients successfully diagnosed with TB, patients who start treatment, patients retained to treatment completion, and patients who achieve 1-y recurrence-free survival. We estimate each step of the cascade for 2013 using data from two World Health Organization (WHO) reports (2014–2015), one WHO dataset (2015), and three RNTCP reports (2014–2016). In addition, we conduct three targeted systematic reviews of the scientific literature to identify 39 unique articles published from 2000–2015 that provide additional data on five indicators that help estimate different steps of the TB cascade. We construct separate care cascades for the overall population of patients with active TB and for patients with specific forms of TB—including new smear-positive, new smear-negative, retreatment smear-positive, and multidrug-resistant (MDR) TB. The WHO estimated that there were 2,700,000 (95%CI: 1,800,000–3,800,000) prevalent TB patients in India in 2013. Of these patients, we estimate that 1,938,027 (72%) TB patients were evaluated at RNTCP facilities; 1,629,906 (60%) were successfully diagnosed; 1,417,838 (53%) got registered for treatment; 1,221,764 (45%) completed treatment; and 1,049,237 (95%CI: 1,008,775–1,083,243), or 39%, of 2,700,000 TB patients achieved the optimal outcome of 1-y recurrence-free survival. The separate cascades for different forms of TB highlight different patterns of patient attrition. Pretreatment loss to follow-up of diagnosed patients and post-treatment TB recurrence were major points of attrition in the new smear-positive TB cascade. In the new smear-negative and MDR TB cascades, a substantial proportion of patients who were evaluated at RNTCP diagnostic facilities were not successfully diagnosed. Retreatment smear-positive and MDR TB patients had poorer treatment outcomes than the general TB population. Limitations of our analysis include the lack of available data on the cascade of care in the private sector and substantial uncertainty regarding the 1-y period prevalence of TB in India. Conclusions Increasing case detection is critical to improving outcomes in India’s TB cascade of care, especially for smear-negative and MDR TB patients. For new smear-positive patients, pretreatment loss to follow-up and post-treatment TB recurrence are considerable points of attrition that may contribute to ongoing TB transmission. Future multisite studies providing more accurate information on key steps in the public sector TB cascade and extension of this analysis to private sector patients may help to better target interventions and resources for TB control in India. PMID:27780217

The Tuberculosis Cascade of Care in India's Public Sector: A Systematic Review and Meta-analysis.

PubMed

Subbaraman, Ramnath; Nathavitharana, Ruvandhi R; Satyanarayana, Srinath; Pai, Madhukar; Thomas, Beena E; Chadha, Vineet K; Rade, Kiran; Swaminathan, Soumya; Mayer, Kenneth H

2016-10-01

India has 23% of the global burden of active tuberculosis (TB) patients and 27% of the world's "missing" patients, which includes those who may not have received effective TB care and could potentially spread TB to others. The "cascade of care" is a useful model for visualizing deficiencies in case detection and retention in care, in order to prioritize interventions. The care cascade constructed in this paper focuses on the Revised National TB Control Programme (RNTCP), which treats about half of India's TB patients. We define the TB cascade as including the following patient populations: total prevalent active TB patients in India, TB patients who reach and undergo evaluation at RNTCP diagnostic facilities, patients successfully diagnosed with TB, patients who start treatment, patients retained to treatment completion, and patients who achieve 1-y recurrence-free survival. We estimate each step of the cascade for 2013 using data from two World Health Organization (WHO) reports (2014-2015), one WHO dataset (2015), and three RNTCP reports (2014-2016). In addition, we conduct three targeted systematic reviews of the scientific literature to identify 39 unique articles published from 2000-2015 that provide additional data on five indicators that help estimate different steps of the TB cascade. We construct separate care cascades for the overall population of patients with active TB and for patients with specific forms of TB-including new smear-positive, new smear-negative, retreatment smear-positive, and multidrug-resistant (MDR) TB. The WHO estimated that there were 2,700,000 (95%CI: 1,800,000-3,800,000) prevalent TB patients in India in 2013. Of these patients, we estimate that 1,938,027 (72%) TB patients were evaluated at RNTCP facilities; 1,629,906 (60%) were successfully diagnosed; 1,417,838 (53%) got registered for treatment; 1,221,764 (45%) completed treatment; and 1,049,237 (95%CI: 1,008,775-1,083,243), or 39%, of 2,700,000 TB patients achieved the optimal outcome of 1-y recurrence-free survival. The separate cascades for different forms of TB highlight different patterns of patient attrition. Pretreatment loss to follow-up of diagnosed patients and post-treatment TB recurrence were major points of attrition in the new smear-positive TB cascade. In the new smear-negative and MDR TB cascades, a substantial proportion of patients who were evaluated at RNTCP diagnostic facilities were not successfully diagnosed. Retreatment smear-positive and MDR TB patients had poorer treatment outcomes than the general TB population. Limitations of our analysis include the lack of available data on the cascade of care in the private sector and substantial uncertainty regarding the 1-y period prevalence of TB in India. Increasing case detection is critical to improving outcomes in India's TB cascade of care, especially for smear-negative and MDR TB patients. For new smear-positive patients, pretreatment loss to follow-up and post-treatment TB recurrence are considerable points of attrition that may contribute to ongoing TB transmission. Future multisite studies providing more accurate information on key steps in the public sector TB cascade and extension of this analysis to private sector patients may help to better target interventions and resources for TB control in India.

The Islamic State Battle Plan: Press Release Natural Language Processing

DTIC Science & Technology

2016-06-01

Processing, text mining , corpus, generalized linear model, cascade, R Shiny, leaflet, data visualization 15. NUMBER OF PAGES 83 16. PRICE CODE...Terrorism and Responses to Terrorism TDM Term Document Matrix TF Term Frequency TF-IDF Term Frequency-Inverse Document Frequency tm text mining (R...package=leaflet. Feinerer I, Hornik K (2015) Text Mining Package “tm,” Version 0.6-2. (Jul 3) https://cran.r-project.org/web/packages/tm/tm.pdf

Streakline flow visualization study of a horseshoe vortex in a large-scale, two-dimensional turbine stator cascade

NASA Technical Reports Server (NTRS)

Gaugler, R. E.; Russell, L. M.

1979-01-01

Neutrally bouyant helium-filled bubbles were observed as they followed the streamlines in a horseshoe vortex system around the vane leading edge in a large scale, two dimensional, turbine stator cascade. Inlet Reynolds number, based on true chord, ranged between 100,000 to 300,000. Bubbles were introduced into the endwall boundary layer through a slot upstream of the vane leading edge. The paths of the bubbles were recorded photographically as streaklines on 16 mm movie film. Individual frames from the film were selected, and overlayed to show the details of the horseshoe vortex around the leading edge. The transport of the vortex across the passage near the leading edge is clearly seen when compared to the streaks formed by bubbles carried in the main stream. Limiting streamlines on the endwall surface were traced by the flow of oil drops.

Role of substance use in HIV care cascade outcomes among people who inject drugs in Russia.

PubMed

Idrisov, Bulat; Lunze, Karsten; Cheng, Debbie M; Blokhina, Elena; Gnatienko, Natalia; Quinn, Emily; Bridden, Carly; Walley, Alexander Y; Bryant, Kendall J; Lioznov, Dmitry; Krupitsky, Evgeny; Samet, Jeffrey H

2017-12-04

Engaging people who drink alcohol or inject drugs in HIV care can be challenging, particularly in Eastern Europe. Healthcare facilities in Russia are organized by specialty; therefore linking patients from addiction care to HIV hospitals has been difficult. The HIV care cascade outlines stages of HIV care (e.g., linkage to care, prescribed antiretroviral therapy [ART], and achieving HIV viral suppression). We hypothesized that unhealthy alcohol use, injection drug use, and opioid craving are associated with unfavorable HIV care cascade outcomes. We analyzed data from a cohort (n = 249) of HIV-positive Russians who have been in addiction hospital treatment in the past year and had a lifetime history of injection drug use (IDU). We evaluated the association between unhealthy alcohol use (AUDIT score > 7 [both hazardous drinking and dependence]), past-month injection drug use (IDU), and opioid craving (visual analogue scale from 1 to 100) with HIV care cascade outcomes. The primary outcome was linkage to HIV care within 12 months. Other outcomes were prescription of ART (secondary) and achievement of undetectable HIV viral load (HVL < 500 copies/mL) within 12 months (exploratory); the latter was analyzed on a subset in which HVL was measured (n = 48). We assessed outcomes via medical record review (linkage, ART) and serum tests (HVL). To examine the primary outcome, we used multiple logistic regression models controlling for potential confounders. Among 249 study participants, unhealthy alcohol use (n = 148 [59%]) and past-month IDU (n = 130 [52%]) were common. The mean opioid craving score was 49 (SD: 38). We were unable to detect significant associations between the independent variables (i.e., unhealthy alcohol use, IDU and opioid craving) and any HIV care cascade outcomes in unadjusted and adjusted analyses. In this cohort of HIV-positive Russians with a history of IDU, individual substance use factors were not significantly associated with achieving HIV care cascade milestones (i.e., linkage to HIV care; prescription for ART; or suppressed viral load). Given no detection of an association of cascade outcomes with recent unhealthy use of alcohol or injection drugs in this cohort, examining systemic factors to understand determinants of HIV care engagement for people with drug use would be important.

Expansion of signal transduction by G proteins The second 15 years or so: From 3 to 16 α subunits plus βγ dimers

PubMed Central

Birnbaumer, Lutz

2007-01-01

The first 15 years, or so, brought the realization that there existed a G protein coupled signal transduction mechanism by which hormone receptors regulate adenylyl cyclases and the light receptor rhodopsin activates visual phosphodiesterase. Three G proteins, Gs, Gi and transducin (T) had been characterized as αβγ heterotrimers, and Gsα-GTP and Tα-GTP had been identified as the sigaling arms of Gs and T. These discoveries were made using classical biochemical approaches, and culminated in the purification of these G proteins. The second 15 years, or so, are the subject of the present review. This time coincided with the advent of powerful recombinant DNA techniques. Combined with the classical approaches, the field expanded the repertoire of G proteins from 3 to 16, discovered the superfamily of seven transmembrane G protein coupled receptors (GPCRs) – which is not addressed in this article – and uncovered an amazing repertoire of effector functions regulated not only by αGTP complexes but also by βγ dimers. Emphasis is placed in presenting how the field developed with the hope of conveying why many of the new findings were made. PMID:17258171

Visualization in simulation tools: requirements and a tool specification to support the teaching of dynamic biological processes.

PubMed

Jørgensen, Katarina M; Haddow, Pauline C

2011-08-01

Simulation tools are playing an increasingly important role behind advances in the field of systems biology. However, the current generation of biological science students has either little or no experience with such tools. As such, this educational glitch is limiting both the potential use of such tools as well as the potential for tighter cooperation between the designers and users. Although some simulation tool producers encourage their use in teaching, little attempt has hitherto been made to analyze and discuss their suitability as an educational tool for noncomputing science students. In general, today's simulation tools assume that the user has a stronger mathematical and computing background than that which is found in most biological science curricula, thus making the introduction of such tools a considerable pedagogical challenge. This paper provides an evaluation of the pedagogical attributes of existing simulation tools for cell signal transduction based on Cognitive Load theory. Further, design recommendations for an improved educational simulation tool are provided. The study is based on simulation tools for cell signal transduction. However, the discussions are relevant to a broader biological simulation tool set.

MARS15

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mokhov, Nikolai

MARS is a Monte Carlo code for inclusive and exclusive simulation of three-dimensional hadronic and electromagnetic cascades, muon, heavy-ion and low-energy neutron transport in accelerator, detector, spacecraft and shielding components in the energy range from a fraction of an electronvolt up to 100 TeV. Recent developments in the MARS15 physical models of hadron, heavy-ion and lepton interactions with nuclei and atoms include a new nuclear cross section library, a model for soft pion production, the cascade-exciton model, the quark gluon string models, deuteron-nucleus and neutrino-nucleus interaction models, detailed description of negative hadron and muon absorption and a unified treatment ofmore » muon, charged hadron and heavy-ion electromagnetic interactions with matter. New algorithms are implemented into the code and thoroughly benchmarked against experimental data. The code capabilities to simulate cascades and generate a variety of results in complex media have been also enhanced. Other changes in the current version concern the improved photo- and electro-production of hadrons and muons, improved algorithms for the 3-body decays, particle tracking in magnetic fields, synchrotron radiation by electrons and muons, significantly extended histograming capabilities and material description, and improved computational performance. In addition to direct energy deposition calculations, a new set of fluence-to-dose conversion factors for all particles including neutrino are built into the code. The code includes new modules for calculation of Displacement-per-Atom and nuclide inventory. The powerful ROOT geometry and visualization model implemented in MARS15 provides a large set of geometrical elements with a possibility of producing composite shapes and assemblies and their 3D visualization along with a possible import/export of geometry descriptions created by other codes (via the GDML format) and CAD systems (via the STEP format). The built-in MARS-MAD Beamline Builder (MMBLB) was redesigned for use with the ROOT geometry package that allows a very efficient and highly-accurate description, modeling and visualization of beam loss induced effects in arbitrary beamlines and accelerator lattices. The MARS15 code includes links to the MCNP-family codes for neutron and photon production and transport below 20 MeV, to the ANSYS code for thermal and stress analyses and to the STRUCT code for multi-turn particle tracking in large synchrotrons and collider rings.« less

Identification of bovine leukemia virus tax function associated with host cell transcription, signaling, stress response and immune response pathway by microarray-based gene expression analysis

PubMed Central

2012-01-01

Background Bovine leukemia virus (BLV) is associated with enzootic bovine leukosis and is closely related to human T-cell leukemia virus type I. The Tax protein of BLV is a transcriptional activator of viral replication and a key contributor to oncogenic potential. We previously identified interesting mutant forms of Tax with elevated (TaxD247G) or reduced (TaxS240P) transactivation effects on BLV replication and propagation. However, the effects of these mutations on functions other than transcriptional activation are unknown. In this study, to identify genes that play a role in the cascade of signal events regulated by wild-type and mutant Tax proteins, we used a large-scale host cell gene-profiling approach. Results Using a microarray containing approximately 18,400 human mRNA transcripts, we found several alterations after the expression of Tax proteins in genes involved in many cellular functions such as transcription, signal transduction, cell growth, apoptosis, stress response, and immune response, indicating that Tax protein has multiple biological effects on various cellular environments. We also found that TaxD247G strongly regulated more genes involved in transcription, signal transduction, and cell growth functions, contrary to TaxS240P, which regulated fewer genes. In addition, the expression of genes related to stress response significantly increased in the presence of TaxS240P as compared to wild-type Tax and TaxD247G. By contrast, the largest group of downregulated genes was related to immune response, and the majority of these genes belonged to the interferon family. However, no significant difference in the expression level of downregulated genes was observed among the Tax proteins. Finally, the expression of important cellular factors obtained from the human microarray results were validated at the RNA and protein levels by real-time quantitative reverse transcription-polymerase chain reaction and western blotting, respectively, after transfecting Tax proteins into bovine cells and human HeLa cells. Conclusion A comparative analysis of wild-type and mutant Tax proteins indicates that Tax protein exerts a significant impact on cellular functions as diverse as transcription, signal transduction, cell growth, stress response and immune response. Importantly, our study is the first report that shows the extent to which BLV Tax regulates the innate immune response. PMID:22455445

Use of Pressure Sensitive Paint for Diagnostics in Turbomachinery Flows With Shocks

NASA Technical Reports Server (NTRS)

Lepicovsky, Jan; Bencic, Timothy J.

2001-01-01

The technology of pressure sensitive paint (PSP) is well established in external aerodynamics. In internal flows in narrow channels and in turbomachinery cascades, however, there are still unresolved problems. In particular, the internal flows with complex shock structures inside highly curved channels present a challenge. It is not always easy and straightforward to distinguish between true signals and "ghost" images due to multiple internal reflections in narrow channels. To address some of the problems, investigations were first carried out in a narrow supersonic channel of Mach number 2.5. A single wedge or a combination of two wedges were used to generate a complex shock wave structure in the flow. The experience gained in a small supersonic channel was used for surface pressure measurements on the stator vane of a supersonic throughflow fan. The experimental results for several fan operating conditions are shown in a concise form, including performance map points, midspan static tap pressure distributions, and vane suction side pressure fields. Finally, the PSP technique was used in the NASA transonic flutter cascade to compliment flow visualization data and to acquire backwall pressure fields to assess the cascade flow periodicity. A summary of shortcomings of the pressure sensitive paint technology for internal flow application and lessons learned are presented in the conclusion of the paper.

Hemodynamic Response Alteration As a Function of Task Complexity and Expertise—An fNIRS Study in Jugglers

PubMed Central

Carius, Daniel; Andrä, Christian; Clauß, Martina; Ragert, Patrick; Bunk, Michael; Mehnert, Jan

2016-01-01

Detailed knowledge about online brain processing during the execution of complex motor tasks with a high motion range still remains elusive. The aim of the present study was to investigate the hemodynamic responses within sensorimotor networks as well as in visual motion area during the execution of a complex visuomotor task such as juggling. More specifically, we were interested in how far the hemodynamic response as measured with functional near infrared spectroscopy (fNIRS) adapts as a function of task complexity and the level of the juggling expertise. We asked expert jugglers to perform different juggling tasks with different levels of complexity such as a 2-ball juggling, 3- and 5-ball juggling cascades. We here demonstrate that expert jugglers show an altered neurovascular response with increasing task complexity, since a 5-ball juggling cascade showed enhanced hemodynamic responses for oxygenated hemoglobin as compared to less complex tasks such as a 3- or 2-ball juggling pattern. Moreover, correlations between the hemodynamic response and the level of the juggling expertise during the 5-ball juggling cascade, acquired by cinematographic video analysis, revealed only a non-significant trend in primary motor cortex, indicating that a higher level of expertise might be associated with lower hemodynamic responses. PMID:27064925

Use of pressure-sensitive paint for diagnostics in turbomachinery flows with shocks

NASA Astrophysics Data System (ADS)

Lepicovsky, J.; Bencic, T. J.

2002-07-01

The technology of pressure-sensitive paint (PSP) is well established in external aerodynamics. In internal flows in narrow channels and in turbomachinery cascades, however, there are still unresolved problems. In particular, the internal flows with complex shock structures inside highly curved channels present a challenge. It is not always easy and straightforward to distinguish between true signals and 'ghost' images due to multiple internal reflections in narrow channels. To address some of the problems, investigations were first carried out in a narrow supersonic channel of Mach number 2.5. A single wedge or a combination of two wedges was used to generate a complex shock wave structure in the flow. The experience gained in a small supersonic channel was used for surface pressure measurements on the stator vane of a supersonic throughflow fan. The experimental results for several fan operating conditions are shown in a concise form, including performance map test points, midspan static tap pressure distributions, and vane suction side pressure fields. Finally, the PSP technique was used in the NASA transonic flutter cascade to compliment flow visualization data and to acquire backwall pressure fields to assess the cascade flow periodicity. Lessons learned from this investigation and shortcomings of the PSP technology for internal flow application are presented in the conclusion of the paper.

Quantum dot coating of baculoviral vectors enables visualization of transduced cells and tissues

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhao, Ying; Lo, Seong Loong; Zheng, Yuangang

2013-04-26

Highlights: •The use of quantum dot (QD)-labeled viral vectors for in vivo imaging is not well investigated. •A new method to label enveloped baculovirus with glutathione-capped CdTe QDs is developed. •The labeling enables the identification of transduced, cultured cells based on fluorescence. •The labeling also allows evaluation of viral transduction in a real-time manner in living mice. •The method has the potential to assess viral vector-based gene therapy protocols in future. -- Abstract: Imaging of transduced cells and tissues is valuable in developing gene transfer vectors and evaluating gene therapy efficacy. We report here a simple method to use brightmore » and photostable quantum dots to label baculovirus, an emerging gene therapy vector. The labeling was achieved through the non-covalent interaction of glutathione-capped CdTe quantum dots with the virus envelope, without the use of chemical conjugation. The quantum dot labeling was nondestructive to viral transduction function and enabled the identification of baculoviral vector-transduced, living cells based on red fluorescence. When the labeled baculoviral vectors were injected intravenously or intraventricularly for in vivo delivery of a transgene into mice, quantum dot fluorescence signals allow us monitor whether or not the injected tissues were transduced. More importantly, using a dual-color whole-body imaging technology, we demonstrated that in vivo viral transduction could be evaluated in a real-time manner in living mice. Thus, our method of labeling a read-to-use gene delivery vector with quantum dots could be useful towards the improvement of vector design and will have the potential to assess baculovirus-based gene therapy protocols in future.« less

Genetic Circuits that Govern Bisexual and Unisexual Reproduction in Cryptococcus neoformans

PubMed Central

Feretzaki, Marianna; Heitman, Joseph

2013-01-01

Cryptococcus neoformans is a human fungal pathogen with a defined sexual cycle. Nutrient-limiting conditions and pheromones induce a dimorphic transition from unicellular yeast to multicellular hyphae and the production of infectious spores. Sexual reproduction involves cells of either opposite (bisexual) or one (unisexual) mating type. Bisexual and unisexual reproduction are governed by shared components of the conserved pheromone-sensing Cpk1 MAPK signal transduction cascade and by Mat2, the major transcriptional regulator of the pathway. However, the downstream targets of the pathway are largely unknown, and homology-based approaches have failed to yield downstream transcriptional regulators or other targets. In this study, we applied insertional mutagenesis via Agrobacterium tumefaciens transkingdom DNA delivery to identify mutants with unisexual reproduction defects. In addition to elements known to be involved in sexual development (Crg1, Ste7, Mat2, and Znf2), three key regulators of sexual development were identified by our screen: Znf3, Spo11, and Ubc5. Spo11 and Ubc5 promote sporulation during both bisexual and unisexual reproduction. Genetic and phenotypic analyses provide further evidence implicating both genes in the regulation of meiosis. Phenotypic analysis of sexual development showed that Znf3 is required for hyphal development during unisexual reproduction and also plays a central role during bisexual reproduction. Znf3 promotes cell fusion and pheromone production through a pathway parallel to and independent of the pheromone signaling cascade. Surprisingly, Znf3 participates in transposon silencing during unisexual reproduction and may serve as a link between RNAi silencing and sexual development. Our studies illustrate the power of unbiased genetic screens to reveal both novel and conserved circuits that operate sexual reproduction. PMID:23966871

Kinase cascades and ligand-directed signaling at the kappa opioid receptor.

PubMed

Bruchas, Michael R; Chavkin, Charles

2010-06-01

The dynorphin/kappa opioid receptor (KOR) system has been implicated as a critical component of the stress response. Stress-induced activation of dynorphin-KOR is well known to produce analgesia, and more recently, it has been implicated as a mediator of stress-induced responses including anxiety, depression, and reinstatement of drug seeking. Drugs selectively targeting specific KOR signaling pathways may prove potentially useful as therapeutic treatments for mood and addiction disorders. KOR is a member of the seven transmembrane spanning (7TM) G-protein coupled receptor (GPCR) superfamily. KOR activation of pertussis toxin-sensitive G proteins leads to Galphai/o inhibition of adenylyl cyclase production of cAMP and releases Gbetagamma, which modulates the conductances of Ca(+2) and K(+) channels. In addition, KOR agonists activate kinase cascades including G-protein coupled Receptor Kinases (GRK) and members of the mitogen-activated protein kinase (MAPK) family: ERK1/2, p38 and JNK. Recent pharmacological data suggests that GPCRs exist as dynamic, multi-conformational protein complexes that can be directed by specific ligands towards distinct signaling pathways. Ligand-induced conformations of KOR that evoke beta-arrestin-dependent p38 MAPK activation result in aversion; whereas ligand-induced conformations that activate JNK without activating arrestin produce long-lasting inactivation of KOR signaling. In this review, we discuss the current status of KOR signal transduction research and the data that support two novel hypotheses: (1) KOR selective partial agonists that do not efficiently activate p38 MAPK may be useful analgesics without producing the dysphoric or hallucinogenic effects of selective, highly efficacious KOR agonists and (2) KOR antagonists that do not activate JNK may be effective short-acting drugs that may promote stress-resilience.

Inhibiting Src family tyrosine kinase activity blocks glutamate signalling to ERK1/2 and Akt/PKB but not JNK in cultured striatal neurones.

PubMed

Crossthwaite, Andrew J; Valli, Haseeb; Williams, Robert J

2004-03-01

Glutamate receptor activation of mitogen-activated protein (MAP) kinase signalling cascades has been implicated in diverse neuronal functions such as synaptic plasticity, development and excitotoxicity. We have previously shown that Ca2+-influx through NMDA receptors in cultured striatal neurones mediates the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) and Akt/protein kinase B (PKB) through a phosphatidylinositol 3-kinase (PI 3-kinase)-dependent pathway. Exposing neurones to the Src family tyrosine kinase inhibitor PP2, but not the inactive analogue PP3, inhibited NMDA receptor-induced phosphorylation of ERK1/2 and Akt/PKB in a concentration-dependent manner, and reduced cAMP response element-binding protein (CREB) phosphorylation. To establish a link between Src family tyrosine kinase-mediated phosphorylation and PI 3-kinase signalling, affinity precipitation experiments were performed with the SH2 domains of the PI 3-kinase regulatory subunit p85. This revealed a Src-dependent phosphorylation of a focal adhesion kinase (FAK)-p85 complex on glutamate stimulation. Demonstrating that PI3-kinase is not ubiquitously involved in NMDA receptor signal transduction, the PI 3-kinase inhibitors wortmannin and LY294002 did not prevent NMDA receptor Ca2+-dependent phosphorylation of c-Jun N-terminal kinase 1/2 (JNK1/2). Further, inhibiting Src family kinases increased NMDA receptor-dependent JNK1/2 phosphorylation, suggesting that Src family kinase-dependent cascades may physiologically limit signalling to JNK. These results demonstrate that Src family tyrosine kinases and PI3-kinase are pivotal regulators of NMDA receptor signalling to ERK/Akt and JNK in striatal neurones.

[Rbf1 (RPG-box binding factor), a transcription factor involved in yeast-hyphal transition of Candida albicans].

PubMed

Aoki, Y; Ishii, N; Watanabe, M; Yoshihara, F; Arisawa, M

1998-01-01

The major fungal pathogen for fungal diseases which have become a major medical problem in the last few years is Candida albicans, which can grow both in yeast and hyphae forms. This ability of C. albicans is thought to contribute to its colonization and dissemination within host tissues. In a recent few years, accompanying the introduction of molecular biological tools into C. albicans organism, several factors involved in the signal transduction pathway for yeast-hyphal transition have been identified. One MAP kinase pathway in C. albicans, similar to that leading to STE12 activation in Saccharomyces cerevisiae, has been reported. C. albicans strains mutant in these genes show retarded filamentous growth on a solid media but no impairment of filamentous growth in mice. These results suggest two scenarios that a kinase signaling cascade plays a part in stimulating the morphological transition in C. albicans, and that there would be another signaling pathway effective in animals. In this latter true hyphal pathway, although some candidate proteins, such as Efg1 (transcription factor), Int1 (integrin-like membrane protein), or Phr1 (pH-regulated membrane protein), have been identified, it is still too early to say that we understand the whole picture of that cascade. We have cloned a C. albicans gene encoding a novel DNA binding protein, Rbf1, that predominantly localizes in the nucleus, and shows transcriptional activation capability. Disruption of the functional RBF1 genes of C. albicans induced the filamentous growth on all solid and liquid media tested, suggesting that Rbf1 might be another candidate for the true hyphal pathway. Relationships with other factors described above, and the target (regulated) genes of Rbf1 is under investigation.

Genetic circuits that govern bisexual and unisexual reproduction in Cryptococcus neoformans.

PubMed

Feretzaki, Marianna; Heitman, Joseph

2013-01-01

Cryptococcus neoformans is a human fungal pathogen with a defined sexual cycle. Nutrient-limiting conditions and pheromones induce a dimorphic transition from unicellular yeast to multicellular hyphae and the production of infectious spores. Sexual reproduction involves cells of either opposite (bisexual) or one (unisexual) mating type. Bisexual and unisexual reproduction are governed by shared components of the conserved pheromone-sensing Cpk1 MAPK signal transduction cascade and by Mat2, the major transcriptional regulator of the pathway. However, the downstream targets of the pathway are largely unknown, and homology-based approaches have failed to yield downstream transcriptional regulators or other targets. In this study, we applied insertional mutagenesis via Agrobacterium tumefaciens transkingdom DNA delivery to identify mutants with unisexual reproduction defects. In addition to elements known to be involved in sexual development (Crg1, Ste7, Mat2, and Znf2), three key regulators of sexual development were identified by our screen: Znf3, Spo11, and Ubc5. Spo11 and Ubc5 promote sporulation during both bisexual and unisexual reproduction. Genetic and phenotypic analyses provide further evidence implicating both genes in the regulation of meiosis. Phenotypic analysis of sexual development showed that Znf3 is required for hyphal development during unisexual reproduction and also plays a central role during bisexual reproduction. Znf3 promotes cell fusion and pheromone production through a pathway parallel to and independent of the pheromone signaling cascade. Surprisingly, Znf3 participates in transposon silencing during unisexual reproduction and may serve as a link between RNAi silencing and sexual development. Our studies illustrate the power of unbiased genetic screens to reveal both novel and conserved circuits that operate sexual reproduction.

Transcriptional profiling in rat hair follicles following simulated Blast insult: a new diagnostic tool for traumatic brain injury.

PubMed

Zhang, Jing; Carnduff, Lisa; Norman, Grant; Josey, Tyson; Wang, Yushan; Sawyer, Thomas W; Martyniuk, Christopher J; Langlois, Valerie S

2014-01-01

With wide adoption of explosive-dependent weaponry during military activities, Blast-induced neurotrauma (BINT)-induced traumatic brain injury (TBI) has become a significant medical issue. Therefore, a robust and accessible biomarker system is in demand for effective and efficient TBI diagnosis. Such systems will also be beneficial to studies of TBI pathology. Here we propose the mammalian hair follicles as a potential candidate. An Advanced Blast Simulator (ABS) was developed to generate shock waves simulating traumatic conditions on brains of rat model. Microarray analysis was performed in hair follicles to identify the gene expression profiles that are associated with shock waves. Gene set enrichment analysis (GSEA) and sub-network enrichment analysis (SNEA) were used to identify cell processes and molecular signaling cascades affected by simulated bomb blasts. Enrichment analyses indicated that genes with altered expression levels were involved in central nervous system (CNS)/peripheral nervous system (PNS) responses as well as signal transduction including Ca2+, K+-transportation-dependent signaling, Toll-Like Receptor (TLR) signaling and Mitogen Activated Protein Kinase (MAPK) signaling cascades. Many of the pathways identified as affected by shock waves in the hair follicles have been previously reported to be TBI responsive in other organs such as brain and blood. The results suggest that the hair follicle has some common TBI responsive molecular signatures to other tissues. Moreover, various TBI-associated diseases were identified as preferentially affected using a gene network approach, indicating that the hair follicle may be capable of reflecting comprehensive responses to TBI conditions. Accordingly, the present study demonstrates that the hair follicle is a potentially viable system for rapid and non-invasive TBI diagnosis.

Developmental evolutionary biology of the vertebrate ear: conserving mechanoelectric transduction and developmental pathways in diverging morphologies

NASA Technical Reports Server (NTRS)

Fritzsch, B.; Beisel, K. W.; Bermingham, N. A.

2000-01-01

This brief overview shows that a start has been made to molecularly dissect vertebrate ear development and its evolutionary conservation to the development of the insect hearing organ. However, neither the patterning process of the ear nor the patterning process of insect sensory organs is sufficiently known at the moment to provide more than a first glimpse. Moreover, hardly anything is known about otocyst development of the cephalopod molluscs, another triploblast lineage that evolved complex 'ears'. We hope that the apparent conserved functional and cellular components present in the ciliated sensory neurons/hair cells will also be found in the genes required for vertebrate ear and insect sensory organ morphogenesis (Fig. 3). Likewise, we expect that homologous pre-patterning genes will soon be identified for the non-sensory cell development, which is more than a blocking of neuronal development through the Delta/Notch signaling system. Generation of the apparently unique ear could thus represent a multiplication of non-sensory cells by asymmetric and symmetric divisions as well as modification of existing patterning process by implementing novel developmental modules. In the final analysis, the vertebrate ear may come about by increasing the level of gene interactions in an already existing and highly conserved interactive cascade of bHLH genes. Since this was apparently achieved in all three lineages of triploblasts independently (Fig. 3), we now need to understand how much of the morphogenetic cascades are equally conserved across phyla to generate complex ears. The existing mutations in humans and mice may be able to point the direction of future research to understand the development of specific cell types and morphologies in the formation of complex arthropod, cephalopod, and vertebrate 'ears'.

Involvement of the cGMP pathway in mediating the insulin-inhibitory effect of melatonin in pancreatic beta-cells.

PubMed

Stumpf, Ina; Mühlbauer, Eckhard; Peschke, Elmar

2008-10-01

Recent investigations have demonstrated an influence of melatonin on insulin secretion in pancreatic beta-cells. The effects are receptor-mediated via two parallel signaling pathways. The aim of this study was to examine the relevance of a second melatonin receptor (MT2) as well as the involvement of a third signaling cascade in mediating melatonin effects, i.e. the cyclic guanosine monophosphate (cGMP) pathway. Our results demonstrate that the insulin-inhibiting effect of melatonin could be partly reversed by preincubation with the unspecific melatonin receptor antagonist luzindole as well as by the MT2-receptor-specific antagonist 4P-PDOT (4-phenyl-2-propionamidotetraline). As melatonin is known to modulate cGMP concentration via the MT2 receptor, these data indicate transmission of the melatonin effects via the cGMP transduction cascade. Molecular investigations established the presence of different types of guanylate cyclases, cGMP-specific phosphodiesterases and cyclic nucleotide-gated channels in rat insulinoma beta-cells (INS1). Moreover, variations in mRNA expression were found when comparing day and night values as well as different states of glucose metabolism. Incubation experiments provided evidence that 3-isobutyl-1-methylxanthine (IBMX)-stimulated cGMP concentrations were significantly decreased in INS1 cells exposed to melatonin for 1 hr in a dose- and time-dependent manner. This effect could also be reversed by application of luzindole and 4P-PDOT. Stimulation with 8-Br-cGMP resulted in significantly increased insulin production. In conclusion, it could be demonstrated that the melatonin receptor subtype MT2 as well as the cGMP signaling pathway are involved in mediating the insulin-inhibiting effect of melatonin.

Aggregatibacter actinomycetemcomitans regulates the expression of integrins and reduces cell adhesion via integrin α5 in human gingival epithelial cells.

PubMed

Kochi, Shinsuke; Yamashiro, Keisuke; Hongo, Shoichi; Yamamoto, Tadashi; Ugawa, Yuki; Shimoe, Masayuki; Kawamura, Mari; Hirata-Yoshihara, Chiaki; Ideguchi, Hidetaka; Maeda, Hiroshi; Takashiba, Shogo

2017-12-01

Gingival epithelial cells form a physiological barrier against bacterial invasion. Excessive bacterial invasion destroys the attachment between the tooth surface and the epithelium, resulting in periodontitis. Integrins play a significant role in cell attachment; therefore, we hypothesized that bacterial infection might decrease the expressions of these integrins in gingival epithelial cells, resulting in reduced cell adhesion. Immortalized human gingival epithelial cells were co-cultured with Aggregatibacter actinomycetemcomitans Y4 (Aa Y4), and the gene expression levels of IL-8, proliferating cell nuclear antigen (PCNA), and integrins (α2, α3, α5, β4, and β6) were measured using quantitative reverse transcription polymerase chain reaction. Expression of PCNA and integrins, except integrin α5, was significantly downregulated, while expression of IL-8 and integrin α5 was significantly upregulated in the cells co-cultured with Aa Y4. The number of adherent cells significantly decreased when co-cultured with Aa Y4, as determined using cell adhesion assays. In the cells co-cultured with Aa Y4 and an integrin α5 neutralizing antibody, there was no effect on the expression of IL-8 and PCNA, while the expressions of integrins α2, α3, β4, and β6, and the number of adherent cells did not decrease. The number of invading bacteria in the cells was reduced in the presence of the antibody and increased in the presence of TLR2/4 inhibitor. Therefore, integrin α5 might be involved in Aa Y4 invasion into gingival epithelial cells, and the resulting signal transduction cascade reduces cell adhesion by decreasing the expression of integrins, while the TLR2/4 signaling cascade regulates IL-8 expression.

Leptin attenuates BACE1 expression and amyloid-β genesis via the activation of SIRT1 signaling pathway.

PubMed

Marwarha, Gurdeep; Raza, Shaneabbas; Meiers, Craig; Ghribi, Othman

2014-09-01

The aspartyl protease β-site AβPP-cleaving enzyme 1 (BACE1) catalyzes the rate-limiting step in Aβ production, a peptide at the nexus of neurodegenerative cascades in Alzheimer Disease (AD). The adipocytokine leptin has been demonstrated to reduce Aβ production and decrease BACE1 activity and expression levels. However, the signaling cascades involved in the leptin-induced mitigation in Aβ levels and BACE1 expression levels have not been elucidated. We have demonstrated that the transcription factor nuclear factor - kappa B (NF-κB) positively regulates BACE1 transcription. NF-κB activity is tightly regulated by the mammalian sirtuin SIRT1. Multiple studies have cogently evinced that leptin activates the metabolic master regulator SIRT1. In this study, we determined the extent to which SIRT1 expression and activity regulate the leptin-induced attenuation in BACE1 expression and Aβ levels in cultured human neuroblastoma SH-SY5Y cells. This study also elucidated and delineated the signal transduction pathways involved in the leptin induced mitigation in BACE1 expression. Our results demonstrate for the first time that leptin attenuates the activation and transcriptional activity of NF-κB by reducing the acetylation of the p65 subunit in a SIRT1-dependent manner. Furthermore, our data shows that leptin reduces the NF-κB-mediated transcription of BACE1 and consequently reduces Amyloid-β genesis. Our study provides a valuable insight and a novel mechanism by which leptin reduces BACE1 expression and Amyloid-β production and may help design potential therapeutic interventions. Copyright © 2014 Elsevier B.V. All rights reserved.

Transcriptional Profiling in Rat Hair Follicles following Simulated Blast Insult: A New Diagnostic Tool for Traumatic Brain Injury

PubMed Central

Zhang, Jing; Carnduff, Lisa; Norman, Grant; Josey, Tyson; Wang, Yushan; Sawyer, Thomas W.; Martyniuk, Christopher J.; Langlois, Valerie S.

2014-01-01

With wide adoption of explosive-dependent weaponry during military activities, Blast-induced neurotrauma (BINT)-induced traumatic brain injury (TBI) has become a significant medical issue. Therefore, a robust and accessible biomarker system is in demand for effective and efficient TBI diagnosis. Such systems will also be beneficial to studies of TBI pathology. Here we propose the mammalian hair follicles as a potential candidate. An Advanced Blast Simulator (ABS) was developed to generate shock waves simulating traumatic conditions on brains of rat model. Microarray analysis was performed in hair follicles to identify the gene expression profiles that are associated with shock waves. Gene set enrichment analysis (GSEA) and sub-network enrichment analysis (SNEA) were used to identify cell processes and molecular signaling cascades affected by simulated bomb blasts. Enrichment analyses indicated that genes with altered expression levels were involved in central nervous system (CNS)/peripheral nervous system (PNS) responses as well as signal transduction including Ca2+, K+-transportation-dependent signaling, Toll-Like Receptor (TLR) signaling and Mitogen Activated Protein Kinase (MAPK) signaling cascades. Many of the pathways identified as affected by shock waves in the hair follicles have been previously reported to be TBI responsive in other organs such as brain and blood. The results suggest that the hair follicle has some common TBI responsive molecular signatures to other tissues. Moreover, various TBI-associated diseases were identified as preferentially affected using a gene network approach, indicating that the hair follicle may be capable of reflecting comprehensive responses to TBI conditions. Accordingly, the present study demonstrates that the hair follicle is a potentially viable system for rapid and non-invasive TBI diagnosis. PMID:25136963

Signaling Architectures that Transmit Unidirectional Information Despite Retroactivity.

PubMed

Shah, Rushina; Del Vecchio, Domitilla

2017-08-08

A signaling pathway transmits information from an upstream system to downstream systems, ideally in a unidirectional fashion. A key obstacle to unidirectional transmission is retroactivity, the additional reaction flux that affects a system once its species interact with those of downstream systems. This raises the fundamental question of whether signaling pathways have developed specialized architectures that overcome retroactivity and transmit unidirectional signals. Here, we propose a general procedure based on mathematical analysis that provides an answer to this question. Using this procedure, we analyze the ability of a variety of signaling architectures to transmit one-way (from upstream to downstream) signals, as key biological parameters are tuned. We find that single stage phosphorylation and phosphotransfer systems that transmit signals from a kinase show a stringent design tradeoff that hampers their ability to overcome retroactivity. Interestingly, cascades of these architectures, which are highly represented in nature, can overcome this tradeoff and thus enable unidirectional transmission. By contrast, phosphotransfer systems, and single and double phosphorylation cycles that transmit signals from a substrate, are unable to mitigate retroactivity effects, even when cascaded, and hence are not well suited for unidirectional information transmission. These results are largely independent of the specific reaction-rate constant values, and depend on the topology of the architectures. Our results therefore identify signaling architectures that, allowing unidirectional transmission of signals, embody modular processes that conserve their input/output behavior across multiple contexts. These findings can be used to decompose natural signal transduction networks into modules, and at the same time, they establish a library of devices that can be used in synthetic biology to facilitate modular circuit design. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Intracellular photoinduced oxidative stress by zinc phthalocyanine photosensitization: a study of the early events in real time using confocal microscopy

NASA Astrophysics Data System (ADS)

Alexandratou, Eleni; Yova, Dido; Handris, Panagiotis; Kletsas, Dimitris; Loukas, Spyros

2003-10-01

Oxidative stress has been implicated in several biological and pathological aspects. Reactive oxygen species (ROS) have been proposed to act as signal transduction molecules activating reactions leading to cell rescue or to cell apoptosis/necrosis. In the present study, oxidative stress was induced by photosensitization of zinc phthalocyanine (ZnPc) in human fibroblasts using a photodynamic dose that did not lead to apoptosis or necrosis. The induction of oxidative stress was performed at the microscope stage in preassigned time. The cascade of phenomena evoked was studied in real time and at the single cell level using confocal laser scanning microscopy. Using specific vital fluorescent probes, alterations induced by oxidative stress in mitochondria membrane potential, in intracellular pH and in calcium concentration were recorded. Image processing and analysis techniques were used to quantify the observed changes. Subcellular localization of the photosensitizer was studied in order to determine the primary and immediate ROS target. It was found that ZnPc is mainly localized in the mitochondria region.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ren, Jinqi; Cook, Aaron A.; Bergmeier, Wolfgang

The dynamic regulation of ERK1 and -2 (ERK1/2) is required for precise signal transduction controlling cell proliferation, differentiation, and survival. However, the underlying mechanisms regulating the activation of ERK1/2 are not completely understood. In this study, we show that phosphorylation of RasGRP2, a guanine nucleotide exchange factor (GEF), inhibits its ability to activate the small GTPase Rap1 that ultimately leads to decreased activation of ERK1/2 in cells. ERK2 phosphorylates RasGRP2 at Ser394 located in the linker region implicated in its autoinhibition. These studies identify RasGRP2 as a novel substrate of ERK1/2 and define a negative-feedback loop that regulates the BRaf–MEK–ERKmore » signaling cascade. This negative-feedback loop determines the amplitude and duration of active ERK1/2. -- Highlights: •ERK2 phosphorylates the guanine nucleotide exchange factor RasGRP2 at Ser394. •Phosphorylated RasGRP2 has decreased capacity to active Rap1b in vitro and in cells. •Phosphorylation of RasGRP2 by ERK1/2 introduces a negative-feedback loop into the BRaf-MEK-ERK pathway.« less

A review of natural and synthetic antioxidants important for health and longevity.

PubMed

Wojcik, M; Burzynska-Pedziwiatr, I; Wozniak, L A

2010-01-01

Reactive oxygen species (ROS) generated in the presence of O(2) by mitochondria, phagocytic cells, peroxisomes, and cytochrome P450 enzymes under physiological conditions, may play a dual function in the human organism. On the one hand, they participate in cell signal transduction cascades, leading to the activation of some transcription factors responsible for regulating of the expression of genes relevant for cell growth and differentiation. On the other hand, they cause oxidative damage of cellular DNA, protein and lipids, resulting in the initiation or development of numerous diseases such as cancer, cardiovascular diseases, type 2 diabetes mellitus, cataract, rheumatoid arthritis, or different neurodegenerative diseases. Both endogenous compounds (glutathione, ubiquinol, urate, bilirubin) and enzymes (superoxide dismutase, catalase, glutathione peroxidase) are engaged in the detoxification of ROS. In addition, numerous dietary components such as vitamin C, vitamin E, carotenoids, and polyphenols are thought to be involved in the antioxidant defense system. The present review article is focused on the summary and the assessment of research on the impact of dietary antioxidants in the prevention of chronic diseases, particularly cancer and cardiovascular diseases.

Gravity sensing and signal transduction in vascular plant primary roots.

PubMed

Baldwin, Katherine L; Strohm, Allison K; Masson, Patrick H

2013-01-01

During gravitropism, the potential energy of gravity is converted into a biochemical signal. How this transfer occurs remains one of the most exciting mysteries in plant cell biology. New experiments are filling in pieces of the puzzle. In this review, we introduce gravitropism and give an overview of what we know about gravity sensing in roots of vascular plants, with special highlight on recent papers. When plant roots are reoriented sideways, amyloplast resedimentation in the columella cells is a key initial step in gravity sensing. This process somehow leads to cytoplasmic alkalinization of these cells followed by relocalization of auxin efflux carriers (PINs). This changes auxin flow throughout the root, generating a lateral gradient of auxin across the cap that upon transmission to the elongation zone leads to differential cell elongation and gravibending. We will present the evidence for and against the following players having a role in transferring the signal from the amyloplast sedimentation into the auxin signaling cascade: mechanosensitive ion channels, actin, calcium ions, inositol trisphosphate, receptors/ligands, ARG1/ARL2, spermine, and the TOC complex. We also outline auxin transport and signaling during gravitropism.

mTORC1 Directly Phosphorylates and Regulates Human MAF1▿

PubMed Central

Michels, Annemieke A.; Robitaille, Aaron M.; Buczynski-Ruchonnet, Diane; Hodroj, Wassim; Reina, Jaime H.; Hall, Michael N.; Hernandez, Nouria

2010-01-01

mTORC1 is a central regulator of growth in response to nutrient availability, but few direct targets have been identified. RNA polymerase (pol) III produces a number of essential RNA molecules involved in protein synthesis, RNA maturation, and other processes. Its activity is highly regulated, and deregulation can lead to cell transformation. The human phosphoprotein MAF1 becomes dephosphorylated and represses pol III transcription after various stresses, but neither the significance of the phosphorylations nor the kinase involved is known. We find that human MAF1 is absolutely required for pol III repression in response to serum starvation or TORC1 inhibition by rapamycin or Torin1. The protein is phosphorylated mainly on residues S60, S68, and S75, and this inhibits its pol III repression function. The responsible kinase is mTORC1, which phosphorylates MAF1 directly. Our results describe molecular mechanisms by which mTORC1 controls human MAF1, a key repressor of RNA polymerase III transcription, and add a new branch to the signal transduction cascade immediately downstream of TORC1. PMID:20516213

mTORC1 directly phosphorylates and regulates human MAF1.

PubMed

Michels, Annemieke A; Robitaille, Aaron M; Buczynski-Ruchonnet, Diane; Hodroj, Wassim; Reina, Jaime H; Hall, Michael N; Hernandez, Nouria

2010-08-01

mTORC1 is a central regulator of growth in response to nutrient availability, but few direct targets have been identified. RNA polymerase (pol) III produces a number of essential RNA molecules involved in protein synthesis, RNA maturation, and other processes. Its activity is highly regulated, and deregulation can lead to cell transformation. The human phosphoprotein MAF1 becomes dephosphorylated and represses pol III transcription after various stresses, but neither the significance of the phosphorylations nor the kinase involved is known. We find that human MAF1 is absolutely required for pol III repression in response to serum starvation or TORC1 inhibition by rapamycin or Torin1. The protein is phosphorylated mainly on residues S60, S68, and S75, and this inhibits its pol III repression function. The responsible kinase is mTORC1, which phosphorylates MAF1 directly. Our results describe molecular mechanisms by which mTORC1 controls human MAF1, a key repressor of RNA polymerase III transcription, and add a new branch to the signal transduction cascade immediately downstream of TORC1.

Structural Dynamics in Ras and Related Proteins upon Nucleotide Switching.

PubMed

Harrison, Rane A; Lu, Jia; Carrasco, Martin; Hunter, John; Manandhar, Anuj; Gondi, Sudershan; Westover, Kenneth D; Engen, John R

2016-11-20

Structural dynamics of Ras proteins contributes to their activity in signal transduction cascades. Directly targeting Ras proteins with small molecules may rely on the movement of a conserved structural motif, switch II. To understand Ras signaling and advance Ras-targeting strategies, experimental methods to measure Ras dynamics are required. Here, we demonstrate the utility of hydrogen-deuterium exchange (HDX) mass spectrometry (MS) to measure Ras dynamics by studying representatives from two branches of the Ras superfamily, Ras and Rho. A comparison of differential deuterium exchange between active (GMPPNP-bound) and inactive (GDP-bound) proteins revealed differences between the families, with the most notable differences occurring in the phosphate-binding loop and switch II. The P-loop exchange signature correlated with switch II dynamics observed in molecular dynamics simulations focused on measuring main-chain movement. HDX provides a means of evaluating Ras protein dynamics, which may be useful for understanding the mechanisms of Ras signaling, including activated signaling of pathologic mutants, and for targeting strategies that rely on protein dynamics. Copyright © 2016 Elsevier Ltd. All rights reserved.

cGMP signalling in pre- and post-conditioning: the role of mitochondria.

PubMed

Costa, Alexandre D T; Pierre, Sandrine V; Cohen, Michael V; Downey, James M; Garlid, Keith D

2008-01-15

Much of cell death from ischaemia/reperfusion in heart and other tissues is generally thought to arise from mitochondrial permeability transition (MPT) in the first minutes of reperfusion. In ischaemic pre-conditioning, agonist binding to G(i) protein-coupled receptors prior to ischaemia triggers a signalling cascade that protects the heart from MPT. We believe that the cytosolic component of this trigger pathway terminates in activation of guanylyl cyclase resulting in increased production of cGMP and subsequent activation of protein kinase G (PKG). PKG phosphorylates a protein on the mitochondrial outer membrane (MOM), which then causes the mitochondrial K(ATP) channel (mitoK(ATP)) on the mitochondrial inner membrane to open, leading to increased production of reactive oxygen species (ROS) by the mitochondria. This implies that the protective signal is somehow transmitted from the MOM to its inner membrane. This is accomplished by a series of intermembrane signalling steps that includes protein kinase C (PKCepsilon) activation. The resulting ROS then activate a second PKC pool which, through another signal transduction pathway termed the mediator pathway, causes inhibition of MPT and reduction in cell death.

Stereochemical study of mouse muscone receptor MOR215-1 and vibrational theory based on statistical physics formalism.

PubMed

Ben Khemis, Ismahene; Mechi, Nesrine; Ben Lamine, Abdelmottaleb

2018-02-10

In the biosensor system, olfactory receptor sites could be activated by odorant molecules and then the biological interactions are converted into electrical signals by a signal transduction cascade that leads the toopening of ion channels, generating a current that leads into the cilia and depolarizes the membrane. The aim of this paper is to present a new investigation that allows determining the olfactory band using a monolayer adsorption with identical sites modeling which may also describe the static and the dynamic sensitivities through the expression of the olfactory response. Moreover, knowing the size of receptor site in olfactory sensory neurons provides valuable information about the relationship between molecular structure and biological activity. The determination of microreceptors and mesoreceptors is mostly carried out via physical adsorption and the radius is calculated using the Kelvin equation. The mean values of radius obtained from the maximum of the receptor size distributions peaks are 4 nm for ℓ-muscone and 6 nm for d-muscone. Copyright © 2018. Published by Elsevier Ltd.

Non-immune cells equipped with T cell receptor-like signaling for cancer cell ablation

PubMed Central

Kojima, Ryosuke; Scheller, Leo; Fussenegger, Martin

2017-01-01

The ability to engineer custom cell-contact-sensing output devices into human non-immune cells would be useful for extending the applicability of cell-based cancer therapies and avoiding risks associated with engineered immune cells. Here, we have developed a new class of synthetic T-cell receptor-like signal-transduction device that functions efficiently in human non-immune cells and triggers release of output molecules specifically upon sensing contact with a target cell. This device employs an interleukin signaling cascade, whose OFF/ON switching is controlled by biophysical segregation of a transmembrane signal-inhibitory protein from the sensor cell/target cell interface. We further showed that designer non-immune cells equipped with this device driving expression of a membrane-penetrator/prodrug-activating enzyme construct could specifically kill target cells in the presence of the prodrug, indicating its potential usefulness for target-cell-specific, cell-based enzyme-prodrug cancer therapy. Our study also contributes to advancement of synthetic biology by extending available design principles to transmit extracellular information to cells. PMID:29131143

Pharmacological preconditioning by milrinone: memory preserving and neuroprotective effect in ischemia-reperfusion injury in mice.

PubMed

Saklani, Reetu; Jaggi, Amteshwar; Singh, Nirmal

2010-07-01

We tested the neuroprotective effect of milrinone, a phosphodiesterase III inhibitor, in pharmacological preconditioning. Bilateral carotid artery occlusion for 12 min followed by reperfusion for 24 h produced ischemia-reperfusion (I/R) cerebral injury in male Swiss albino mice. Cerebral infarct size was measured using triphenyltetrazolium chloride staining. Memory was assessed using the Morris water maze test, and motor coordination was evaluated using the inclined beam walking test, rota-rod test, and lateral push test. Milrinone (50 microg/kg & 100 microg/kg i.v.) was administered 24 h before surgery in a separate group of animals to induce pharmacological preconditioning. I/R increased cerebral infarct size and impaired memory and motor coordination. Milrinone treatment significantly decreased cerebral infarct size and reversed I/R-induced impairments in memory and motor coordination. This neuroprotective effect was blocked by ruthenium red (3 mg/kg, s.c.), an intracellular ryanodine receptor blocker. These findings indicate that milrinone preconditioning exerts a marked neuroprotective effect on the ischemic brain, putatively due to increased intracellular calcium levels activating calcium-sensitive signal transduction cascades.

Antioxidant Supplementation: A Linchpin in Radiation-Induced Enteritis

PubMed Central

Anwar, Mumtaz; Ahmad, Shabeer; Akhtar, Reyhan; Mahmood, Akhtar

2017-01-01

Radiation enteritis is one of the most feared complications of abdominal and pelvic regions. Thus, radiation to abdominal or pelvic malignancies unavoidably injures the intestine. Because of rapid cell turnover, the intestine is highly sensitive to radiation injury, which is the limiting factor in the permissible dosage of irradiation. Bowel injuries such as fistulas, strictures, and chronic malabsorption are potentially life-threatening complications and have an impact on patient quality of life. The incidence of radiation enteritis is increasing because of the current trend of combined chemotherapy and radiation. The consequences of radiation damage to the intestine may result in considerable morbidity and even mortality. The observed effects of ionizing radiation are mediated mainly by oxygen-free radicals that are generated by its action on water and are involved in several steps of signal transduction cascade, leading to apoptosis. The oxyradicals also induce DNA strand breaks and protein oxidation. An important line of defense against free radical damage is the presence of antioxidants. Therefore, administration of antioxidants may ameliorate the radiation-induced damage to the intestine. PMID:28532242

Using oriented peptide array libraries to evaluate methylarginine-specific antibodies and arginine methyltransferase substrate motifs

PubMed Central

Gayatri, Sitaram; Cowles, Martis W.; Vemulapalli, Vidyasiri; Cheng, Donghang; Sun, Zu-Wen; Bedford, Mark T.

2016-01-01

Signal transduction in response to stimuli relies on the generation of cascades of posttranslational modifications that promote protein-protein interactions and facilitate the assembly of distinct signaling complexes. Arginine methylation is one such modification, which is catalyzed by a family of nine protein arginine methyltransferases, or PRMTs. Elucidating the substrate specificity of each PRMT will promote a better understanding of which signaling networks these enzymes contribute to. Although many PRMT substrates have been identified, and their methylation sites mapped, the optimal target motif for each of the nine PRMTs has not been systematically addressed. Here we describe the use of Oriented Peptide Array Libraries (OPALs) to methodically dissect the preferred methylation motifs for three of these enzymes – PRMT1, CARM1 and PRMT9. In parallel, we show that an OPAL platform with a fixed methylarginine residue can be used to validate the methyl-specific and sequence-specific properties of antibodies that have been generated against different PRMT substrates, and can also be used to confirm the pan nature of some methylarginine-specific antibodies. PMID:27338245

Leptin's effect on taste bud calcium responses and transmitter secretion.

PubMed

Meredith, Tricia L; Corcoran, Alan; Roper, Stephen D

2015-05-01

Leptin, a peptide hormone released by adipose tissue, acts on the hypothalamus to control cravings and appetite. Leptin also acts to decrease taste responses to sweet substances, though there is little detailed information regarding where leptin acts in the taste transduction cascade. The present study examined the effects of leptin on sweet-evoked responses and neuro transmitter release from isolated taste buds. Our results indicate that leptin moderately decreased sweet-evoked calcium mobilization in isolated mouse taste buds. We also employed Chinese hamster ovary biosensor cells to examine taste transmitter release from isolated taste buds. Leptin reduced ATP and increased serotonin release in response to sweet stimulation. However, leptin has no effect on bitter-evoked transmitter release, further showing that the action of leptin is sweet specific. Our results support those of previous studies, which state that leptin acts on taste tissue via the leptin receptor, most likely on Type II (Receptor) cells, but also possibly on Type III (Presynaptic) cells. © The Author 2014. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

DNA-independent PARP-1 activation by phosphorylated ERK2 increases Elk1 activity: a link to histone acetylation.

PubMed

Cohen-Armon, Malka; Visochek, Leonid; Rozensal, Dana; Kalal, Adi; Geistrikh, Ilona; Klein, Rodika; Bendetz-Nezer, Sarit; Yao, Zhong; Seger, Rony

2007-01-26

PolyADP-ribose polymerases (PARPs) catalyze a posttranslational modification of nuclear proteins by polyADP-ribosylation. The catalytic activity of the abundant nuclear protein PARP-1 is stimulated by DNA strand breaks, and PARP-1 activation is required for initiation of DNA repair. Here we show that PARP-1 also acts within extracellular signal-regulated kinase (ERK) signaling cascade that mediates growth and differentiation. The findings reveal an alternative mode of PARP-1 activation, which does not involve binding to DNA or DNA damage. In a cell-free system, recombinant PARP-1 was intensively activated and thereby polyADP-ribosylated by a direct interaction with phosphorylated ERK2, and the activated PARP-1 dramatically increased ERK2-catalyzed phosphorylation of the transcription factor Elk1. In cortical neurons treated with nerve growth factors and in stimulated cardiomyocytes, PARP-1 activation enhanced ERK-induced Elk1-phosphorylation, core histone acetylation, and transcription of the Elk1-target gene c-fos. These findings constitute evidence for PARP-1 activity within the ERK signal-transduction pathway.

The PIM kinases in hematological cancers.

PubMed

Alvarado, Yesid; Giles, Francis J; Swords, Ronan T

2012-02-01

The PIM genes represent a family of proto-oncogenes that encode three different serine/threonine protein kinases (PIM1, PIM2 and PIM3) with essential roles in the regulation of signal transduction cascades, which promote cell survival, proliferation and drug resistance. PIM kinases are overexpressed in several hematopoietic tumors and support in vitro and in vivo malignant cell growth and survival, through cell cycle regulation and inhibition of apoptosis. PIM kinases do not have an identified regulatory domain, which means that these proteins are constitutively active once transcribed. They appear to be critical downstream effectors of important oncoproteins and, when overexpressed, can mediate drug resistance to available agents, such as rapamycin. Recent crystallography studies reveal that, unlike other kinases, they possess a hinge region, which creates a unique binding pocket for ATP, offering a target for an increasing number of potent small-molecule PIM kinase inhibitors. Preclinical studies in models of various hematologic cancers indicate that these novel agents show promising activity and some of them are currently being evaluated in a clinical setting. In this review, we profile the PIM kinases as targets for therapeutics in hematologic malignancies.

The association of β-arrestin2 polymorphisms with response to antidepressant treatment in depressed patients.

PubMed

Petit, Anne-Cécile; El Asmar, Khalil; David, Denis J; Gardier, Alain M; Becquemont, Laurent; Fève, Bruno; Verstuyft, Céline; Corruble, Emmanuelle

2018-02-02

The study of genetic polymorphisms involved in antidepressants (AD) response is essential to provide a personalized medicine approach in the field of depression. β-arrestin 2 (ARRB2) is a candidate gene in the pharmacogenetics of AD as it is involved in the signaling cascade downstream of numerous neurotransmitter receptors. We investigated the association between five ARRB2 single nucleotide polymorphisms (SNPs): rs1045280, rs2036657, rs4790694, rs3786047 and rs452246, and response to AD treatment in a sample of 569 patients with a major depressive episode treated for 6months. We show that GG/GT patients for rs4522461 (n=534) and AA/AC patients for rs4790694 (n=244) have a lower response to AD than other genotype groups (HDRS score of 10.9 vs 8.0 after 6months, multivariate analysis: p=0.03; 12.2 vs 9.6, p=0.02, respectively). These data provide additional evidence that β-arrestin 2 is a regulator of intracellular signal transduction processes involved in AD treatment. Copyright © 2017 Elsevier Inc. All rights reserved.

Activated stress response pathways within multicellular aggregates utilize an autocrine component.

PubMed

Jack, Graham D; Cabrera, M Carla; Manning, Michael L; Slaughter, Stephen M; Potts, Malcolm; Helm, Richard F

2007-04-01

Multicellular aggregates (spheroids) of primary human foreskin fibroblasts (HFF-2) and a glioblastoma cell line (T98G) entered and exited from long term (2 weeks) metabolic arrest utilizing an autocrine response. Cytokine production (specifically IFN-gamma) activated a Gadd45alpha/p38 pathway that led to increased AP-1 (c-jun and ATF3) transcription factor levels, augmenting cytokine production in an autocrine fashion. Whereas HFF-2 aggregates were capable of surviving long term arrest and recovery during NF-kappaB inhibition independent of JNK activation, T98G aggregates were not. Such endogenous processes are not easily observed with adherent monolayer cell culturing systems, strongly suggesting that more emphasis needs to be placed on determining the operational signal transduction cascades within multicellular aggregates. Extracellular inputs such as spheroid formation, arrest, and regrowth as monolayers invoke intracellular signaling responses converging at the AP-1 transcription factor level. Variations in responses are both cell type and transformation state dependent and require an autocrine cytokine component. The data are discussed in relation to the wounding response and avascular tumor growth mechanisms.

Deciphering the function of the CNGB1b subunit in olfactory CNG channels.

PubMed

Nache, Vasilica; Wongsamitkul, Nisa; Kusch, Jana; Zimmer, Thomas; Schwede, Frank; Benndorf, Klaus

2016-07-11

Olfactory cyclic nucleotide-gated (CNG) ion channels are key players in the signal transduction cascade of olfactory sensory neurons. The second messengers cAMP and cGMP directly activate these channels, generating a depolarizing receptor potential. Olfactory CNG channels are composed of two CNGA2 subunits and two modulatory subunits, CNGA4, and CNGB1b. So far the exact role of the modulatory subunits for channel activation is not fully understood. By measuring ligand binding and channel activation simultaneously, we show that in functional heterotetrameric channels not only the CNGA2 subunits and the CNGA4 subunit but also the CNGB1b subunit binds cyclic nucleotides and, moreover, also alone translates this signal to open the pore. In addition, we show that the CNGB1b subunit is the most sensitive subunit in a heterotetrameric channel to cyclic nucleotides and that it accelerates deactivation to a similar extent as does the CNGA4 subunit. In conclusion, the CNGB1b subunit participates in ligand-gated activation of olfactory CNG channels and, particularly, contributes to rapid termination of odorant signal in an olfactory sensory neuron.

Calcific Aortic Valve Disease: Part 2—Morphomechanical Abnormalities, Gene Reexpression, and Gender Effects on Ventricular Hypertrophy and Its Reversibility

PubMed Central

2016-01-01

In part 1, we considered cytomolecular mechanisms underlying calcific aortic valve disease (CAVD), hemodynamics, and adaptive feedbacks controlling pathological left ventricular hypertrophy provoked by ensuing aortic valvular stenosis (AVS). In part 2, we survey diverse signal transduction pathways that precede cellular/molecular mechanisms controlling hypertrophic gene expression by activation of specific transcription factors that induce sarcomere replication in-parallel. Such signaling pathways represent potential targets for therapeutic intervention and prevention of decompensation/failure. Hypertrophy provoking signals, in the form of dynamic stresses and ligand/effector molecules that bind to specific receptors to initiate the hypertrophy, are transcribed across the sarcolemma by several second messengers. They comprise intricate feedback mechanisms involving gene network cascades, specific signaling molecules encompassing G protein-coupled receptors and mechanotransducers, and myocardial stresses. Future multidisciplinary studies will characterize the adaptive/maladaptive nature of the AVS-induced hypertrophy, its gender- and individual patient-dependent peculiarities, and its response to surgical/medical interventions. They will herald more effective, precision medicine treatments. PMID:27184804

Perception of the plant hormone ethylene: known-knowns and known-unknowns.

PubMed

Light, Kenneth M; Wisniewski, John A; Vinyard, W Andrew; Kieber-Emmons, Matthew T

2016-09-01

The gaseous phytohormone ethylene is implicated in virtually all phases of plant growth and development and thus has a major impact on crop production. This agronomic impact makes understanding ethylene signaling the Philosopher's Stone of the plant biotechnology world in applications including post-harvest transport of foodstuffs, consistency of foodstuff maturity pre-harvest, decorative flower freshness and longevity, and biomass production for biofuel applications. Ethylene is biosynthesized by plants in response to environmental factors and plant life-cycle events, and triggers a signaling cascade that modulates over 1000 genes. The key components in the perception of ethylene are a family of copper dependent receptors, the bioinorganic chemistry of which has been largely ignored by the chemical community. Since identification of these receptors two decades ago, there has been tremendous growth in knowledge in the biological community on the signal transduction pathways and mechanisms of ethylene signaling. In this review, we highlight these advances and key chemical voids in knowledge that are overdue for exploration, and which are required to ultimately regulate and control ethylene signaling.

Auxin-induced nitric oxide, cGMP and gibberellins were involved in the gravitropism

NASA Astrophysics Data System (ADS)

Cai, Weiming; Hu, Liwei; Hu, Xiangyang; Cui, Dayong; Cai, Weiming

Gravitropism is the asymmetric growth or curvature of plant organs in response to gravistimulation. There is a complex signal transduction cascade which involved in the differential growth of plants in response to changes in the gravity vector. The role of auxin in gravitropism has been demonstrated by many experiments, but little is known regarding the molecular details of such effects. In our studies before, mediation of the gravitropic bending of soybean roots and rice leaf sheath bases by nitric oxide, cGMP and gibberellins, are induced by auxin. The asymmetrical distribution of nitric oxide, cGMP and gibberellins resulted from the asymmetrical synthesis of them in bending sites. In soybean roots, inhibitions of NO and cGMP synthesis reduced differential NO and cGMP accumulation respectively, which both of these effects can lead to the reduction of gravitropic bending. Gibberellin-induced OsXET, OsEXPA4 and OsRWC3 were also found involved in the gravitropic bending. These data indicated that auxin-induced nitric oxide, cGMP and gibberellins were involved in the gravitropism. More experiments need to prove the more detailed mechanism of them.

A nonlinear model for ionic polymer metal composites as actuators

NASA Astrophysics Data System (ADS)

Bonomo, C.; Fortuna, L.; Giannone, P.; Graziani, S.; Strazzeri, S.

2007-02-01

This paper introduces a comprehensive nonlinear dynamic model of motion actuators based on ionic polymer metal composites (IPMCs) working in air. Significant quantities ruling the acting properties of IPMC-based actuators are taken into account. The model is organized as follows. As a first step, the dependence of the IPMC absorbed current on the voltage applied across its thickness is taken into account; a nonlinear circuit model is proposed to describe this relationship. In a second step the transduction of the absorbed current into the IPMC mechanical reaction is modelled. The model resulting from the cascade of both the electrical and the electromechanical stages represents a novel contribution in the field of IPMCs, capable of describing the electromechanical behaviour of these materials and predicting relevant quantities in a large range of applied signals. The effect of actuator scaling is also investigated, giving interesting support to the activities involved in the design of actuating devices based on these novel materials. Evidence of the excellent agreement between the estimations obtained by using the proposed model and experimental signals is given.

Cross-talk between oxidative stress and pro-inflammatory cytokines in acute pancreatitis: a key role for protein phosphatases.

PubMed

Escobar, Javier; Pereda, Javier; Arduini, Alessandro; Sandoval, Juan; Sabater, Luis; Aparisi, Luis; López-Rodas, Gerardo; Sastre, Juan

2009-01-01

Acute pancreatitis is an acute inflammatory process localized in the pancreatic gland that frequently involves peripancreatic tissues. It is still under investigation why an episode of acute pancreatitis remains mild affecting only the pancreas or progresses to a severe form leading to multiple organ failure and death. Proinflammatory cytokines and oxidative stress play a pivotal role in the early pathophysiological events of the disease. Cytokines such as interleukin 1beta and tumor necrosis factor alpha initiate and propagate almost all consequences of the systemic inflammatory response syndrome. On the other hand, depletion of pancreatic glutathione is an early hallmark of acute pancreatitis and reactive oxygen species are also associated with the inflammatory process. Changes in thiol homestasis and redox signaling decisively contribute to amplification of the inflammatory cascade through mitogen activated protein kinase (MAP kinase) pathways. This review focuses on the relationship between oxidative stress, pro-inflammatory cytokines and MAP kinase/protein phosphatase pathways as major modulators of the inflammatory response in acute pancreatitis. Redox sensitive signal transduction mediated by inactivation of protein phosphatases, particularly protein tyrosin phosphatases, is highlighted.

Conformational dynamics underlie the activity of the auxin-binding protein, Nt-abp1.

PubMed

David, K; Carnero-Diaz, E; Leblanc, N; Monestiez, M; Grosclaude, J; Perrot-Rechenmann, C

2001-09-14

The auxin-binding protein 1 (ABP1) has been proposed to be involved in the perception of the phytohormone at the plasma membrane. Site-directed mutagenesis was performed on highly conserved residues at the C terminus of ABP1 to investigate their relative importance in protein folding and activation of a functional response at the plasma membrane. Detailed analysis of the dynamic interaction of the wild-type ABP1 and mutated proteins with three distinct monoclonal antibodies recognizing conformation-dependent epitopes was performed by surface plasmon resonance. The influence of auxin on these interactions was also investigated. The Cys(177) as well as Asp(175) and Glu(176) were identified as critical residues for ABP1 folding and action at the plasma membrane. On the contrary, the C-terminal KDEL sequence was demonstrated not to be essential for auxin binding, interaction with the plasma membrane, or activation of the transduction cascade although it does appear to be involved in the stability of ABP1. Taken together, the results confirmed that ABP1 conformational change is the critical step for initiating the signal from the plasma membrane.

Cajal-body formation correlates with differential coilin phosphorylation in primary and transformed cell lines.

PubMed

Hearst, Scoty M; Gilder, Andrew S; Negi, Sandeep S; Davis, Misty D; George, Eric M; Whittom, Angela A; Toyota, Cory G; Husedzinovic, Alma; Gruss, Oliver J; Hebert, Michael D

2009-06-01

Cajal bodies (CBs) are nuclear structures that are thought to have diverse functions, including small nuclear ribonucleoprotein (snRNP) biogenesis. The phosphorylation status of coilin, the CB marker protein, might impact CB formation. We hypothesize that primary cells, which lack CBs, contain different phosphoisoforms of coilin compared with that found in transformed cells, which have CBs. Localization, self-association and fluorescence recovery after photobleaching (FRAP) studies on coilin phosphomutants all suggest this modification impacts the function of coilin and may thus contribute towards CB formation. Two-dimensional gel electrophoresis demonstrates that coilin is hyperphosphorylated in primary cells compared with transformed cells. mRNA levels of the nuclear phosphatase PPM1G are significantly reduced in primary cells and expression of PPM1G in primary cells induces CBs. Additionally, PPM1G can dephosphorylate coilin in vitro. Surprisingly, however, expression of green fluorescent protein alone is sufficient to form CBs in primary cells. Taken together, our data support a model whereby coilin is the target of an uncharacterized signal transduction cascade that responds to the increased transcription and snRNP demands found in transformed cells.

Generalized transduction: new aspects of the events in the water column

NASA Astrophysics Data System (ADS)

Velimirov, B.; Chiura, H. X.; Kogure, K.

2003-04-01

Virus mediated transfer of genetic elements among bacteria in nature has become a major research topic in the last decade. Along with conjugation and transformation, transduction is a well-known mechanism resulting in horizontal gene transfer in procaryotic organisms. In the case of generalized transduction, all regions of the procaryotic chromosome or other genetic elements in the donor cell are transferred with nearly the same frequency to the recipient. The injection of this DNA induces the generation of stable transductants. Both virulent and temperate phages have the capability to induce general transduction.Within the frame of a study on intergeneric phage-mediated gene transfer between marine bacteria and enteric bacteria, namely an auxotrophic mutant of Escherichia coli (AB1157) we used virus like particles (VLPs) from an oligotrophic marine environment (Mediterranean Sea, West coast of Corsica) and obtained gene transfer frequencies ranging between 10-2 to 10-6 per viral particle. Consequently we had to assume that an important fraction of the VLPs obtained via ultrafiltration (Minitan Ultrafiltration System, Millipore, USA. 30 kDA cut-off filter) from surface seawater have the capability to induce general transduction. In the process of this investigation we made a number of new observations which were not compatible with the concept of general transduction. The obtained transductants were able to produce new VLPs, which had again the capability to induce transduction. In an attempt to characterize these particles we show that their appearance in the experiment was neither related to plaque formation nor to cell lysis and we discuss the concept of transduction in the light of new experimental evidence concerning transducing particles. Furthermore, a preliminary numerical model allowing an estimation of the transduction events, taking place in the water column within a year is presented.

Seeing in the dark: vision and visual behaviour in nocturnal bees and wasps.

PubMed

Warrant, Eric J

2008-06-01

In response to the pressures of predation, parasitism and competition for limited resources, several groups of (mainly) tropical bees and wasps have independently evolved a nocturnal lifestyle. Like their day-active (diurnal) relatives, these insects possess apposition compound eyes, a relatively light-insensitive eye design that is best suited to vision in bright light. Despite this, nocturnal bees and wasps are able to forage at night, with many species capable of flying through a dark and complex forest between the nest and a foraging site, a behaviour that relies heavily on vision and is limited by light intensity. In the two best-studied species - the Central American sweat bee Megalopta genalis (Halictidae) and the Indian carpenter bee Xylocopa tranquebarica (Apidae) - learned visual landmarks are used to guide foraging and homing. Their apposition eyes, however, have only around 30 times greater optical sensitivity than the eyes of their closest diurnal relatives, a fact that is apparently inconsistent with their remarkable nocturnal visual abilities. Moreover, signals generated in the photoreceptors, even though amplified by a high transduction gain, are too noisy and slow to transmit significant amounts of information in dim light. How have nocturnal bees and wasps resolved these paradoxes? Even though this question remains to be answered conclusively, a mounting body of theoretical and experimental evidence suggests that the slow and noisy visual signals generated by the photoreceptors are spatially summed by second-order monopolar cells in the lamina, a process that could dramatically improve visual reliability for the coarser and slower features of the visual world at night.

Training-induced recovery of low-level vision followed by mid-level perceptual improvements in developmental object and face agnosia

PubMed Central

Lev, Maria; Gilaie-Dotan, Sharon; Gotthilf-Nezri, Dana; Yehezkel, Oren; Brooks, Joseph L; Perry, Anat; Bentin, Shlomo; Bonneh, Yoram; Polat, Uri

2015-01-01

Long-term deprivation of normal visual inputs can cause perceptual impairments at various levels of visual function, from basic visual acuity deficits, through mid-level deficits such as contour integration and motion coherence, to high-level face and object agnosia. Yet it is unclear whether training during adulthood, at a post-developmental stage of the adult visual system, can overcome such developmental impairments. Here, we visually trained LG, a developmental object and face agnosic individual. Prior to training, at the age of 20, LG's basic and mid-level visual functions such as visual acuity, crowding effects, and contour integration were underdeveloped relative to normal adult vision, corresponding to or poorer than those of 5–6 year olds (Gilaie-Dotan, Perry, Bonneh, Malach & Bentin, 2009). Intensive visual training, based on lateral interactions, was applied for a period of 9 months. LG's directly trained but also untrained visual functions such as visual acuity, crowding, binocular stereopsis and also mid-level contour integration improved significantly and reached near-age-level performance, with long-term (over 4 years) persistence. Moreover, mid-level functions that were tested post-training were found to be normal in LG. Some possible subtle improvement was observed in LG's higher-order visual functions such as object recognition and part integration, while LG's face perception skills have not improved thus far. These results suggest that corrective training at a post-developmental stage, even in the adult visual system, can prove effective, and its enduring effects are the basis for a revival of a developmental cascade that can lead to reduced perceptual impairments. PMID:24698161

Training-induced recovery of low-level vision followed by mid-level perceptual improvements in developmental object and face agnosia.

PubMed

Lev, Maria; Gilaie-Dotan, Sharon; Gotthilf-Nezri, Dana; Yehezkel, Oren; Brooks, Joseph L; Perry, Anat; Bentin, Shlomo; Bonneh, Yoram; Polat, Uri

2015-01-01

Long-term deprivation of normal visual inputs can cause perceptual impairments at various levels of visual function, from basic visual acuity deficits, through mid-level deficits such as contour integration and motion coherence, to high-level face and object agnosia. Yet it is unclear whether training during adulthood, at a post-developmental stage of the adult visual system, can overcome such developmental impairments. Here, we visually trained LG, a developmental object and face agnosic individual. Prior to training, at the age of 20, LG's basic and mid-level visual functions such as visual acuity, crowding effects, and contour integration were underdeveloped relative to normal adult vision, corresponding to or poorer than those of 5-6 year olds (Gilaie-Dotan, Perry, Bonneh, Malach & Bentin, 2009). Intensive visual training, based on lateral interactions, was applied for a period of 9 months. LG's directly trained but also untrained visual functions such as visual acuity, crowding, binocular stereopsis and also mid-level contour integration improved significantly and reached near-age-level performance, with long-term (over 4 years) persistence. Moreover, mid-level functions that were tested post-training were found to be normal in LG. Some possible subtle improvement was observed in LG's higher-order visual functions such as object recognition and part integration, while LG's face perception skills have not improved thus far. These results suggest that corrective training at a post-developmental stage, even in the adult visual system, can prove effective, and its enduring effects are the basis for a revival of a developmental cascade that can lead to reduced perceptual impairments. © 2014 The Authors. Developmental Science Published by John Wiley & Sons Ltd.

Stall flutter experiment in a transonic oscillating linear cascade

NASA Technical Reports Server (NTRS)

Boldman, D. R.; Buggele, A. E.; Michalson, G. M.

1981-01-01

Two dimensional biconvex airfoils were oscillated at reduced frequencies up to 0.5 based on semi-chord and a free stream Mach number of 0.80 to simulate transonic stall flutter in rotors. Steady-state periodicity was confirmed through end-wall pressure measurements, exit flow traverses, and flow visualization. The initial flow visualization results from flutter tests indicated that the oscillating shock on the airfoils lagged the airfoil motion by as much as 80 deg. These initial data exhibited an appreciable amount of scatter; however, a linear fit of the results indicated that the greatest shock phase lag occurred at a positive interblade phase angle. Photographs of the steady-state and unsteady flow fields reveal some of the features of the lambda shock wave on the suction surface of the airfoils.

A comparison of neural network and fuzzy clustering techniques in segmenting magnetic resonance images of the brain

NASA Technical Reports Server (NTRS)

Hall, Lawrence O.; Bensaid, Amine M.; Clarke, Laurence P.; Velthuizen, Robert P.; Silbiger, Martin S.; Bezdek, James C.

1992-01-01

Magnetic resonance (MR) brain section images are segmented and then synthetically colored to give visual representations of the original data with three approaches: the literal and approximate fuzzy c-means unsupervised clustering algorithms and a supervised computational neural network, a dynamic multilayered perception trained with the cascade correlation learning algorithm. Initial clinical results are presented on both normal volunteers and selected patients with brain tumors surrounded by edema. Supervised and unsupervised segmentation techniques provide broadly similar results. Unsupervised fuzzy algorithms were visually observed to show better segmentation when compared with raw image data for volunteer studies. However, for a more complex segmentation problem with tumor/edema or cerebrospinal fluid boundary, where the tissues have similar MR relaxation behavior, inconsistency in rating among experts was observed.

Regional Hydraulic Geometry Curves of the Northern Cascade Mountains, Chelan and King Counties, Washington State, USA

NASA Astrophysics Data System (ADS)

Gasperi, J. T.; McClung, J. M.; Hanson, D. L.

2006-12-01

The USDA-Natural Resources Conservation Service has developed regional hydraulic geometry curves relating drainage area to bankfull top width, mean depth and cross-sectional area for the east and west sides of the northern Cascade Mountains in Chelan and King Counties, Washington. NRCS surveyed 10 channel reaches with drainage areas from 1 to 1000 square miles within the Wenatchee River drainage of Chelan County and 10 channel reaches with drainage areas of 1 to 100 square miles within the Cedar and Green River drainages of King County. Selection criteria for stream reaches required a minimum of 20 years of USGS stream gage discharge records, unregulated flows and safe access. Survey data were collected with a Sokkia Total Station during low flow conditions from August 2004 to September 2005. NRCS measured a channel cross-section at each of the USGS stream gage sites and two or three additional cross-sections up and downstream. The authors also collected samples of bed material for gradation analysis and estimation of Manning's roughness coefficient, n. Bankfull elevations were estimated based on visual identification of field indicators and USGS flood discharges for the 50% exceedance probability event. Field data were evaluated with the Ohio DNR Reference Reach spreadsheet to determine bankfull top width, mean depth and cross-sectional area. We applied a simple linear regression to the data following USGS statistical methods to evaluate the closeness of fit between drainage area and bankfull channel dimensions. The resulting R2 values of 0.83 to 0.93 for the eastern Cascade data of Chelan County and 0.71 to 0.88 for the western Cascade data of King County indicate a close association between drainage area and bankfull channel dimensions for these two sets of data.

Optimization of the transductional efficiency of lentiviral vectors: effect of sera and polycations

PubMed Central

Denning, Warren; Das, Suvendu; Guo, Siqi; Xu, Jun; Kappes, John C.; Hel, Zdenek

2012-01-01

Lentiviral vectors are widely used as effective gene-delivery vehicles. Optimization of the conditions for efficient lentiviral transduction is of a high importance for a variety of research applications. Presence of positively-charged polycations reduces the electrostatic repulsion forces between a negatively-charged cell and an approaching enveloped lentiviral particle resulting in an increase in the transduction efficiency. Although a variety of polycations are commonly used to enhance the transduction with retroviruses, the relative effect of various types of polycations on the efficiency of transduction and on the potential bias in the determination of titer of lentiviral vectors is not fully understood. Here we present data suggesting that DEAE-dextran provides superior results in enhancing lentiviral transduction of most tested cell lines and primary cell cultures. Specific type and source of serum affects the efficiency of transduction of target cell populations. Non-specific binding of enhanced green fluorescent protein (EGFP)-containing membrane aggregates in the presence of DEAE-dextran does not significantly affect the determination of the titer of EGFP-expressing lentiviral vectors. In conclusion, various polycations and types of sera should be tested when optimizing lentiviral transduction of target cell populations. PMID:22407723

Optimization of the transductional efficiency of lentiviral vectors: effect of sera and polycations.

PubMed

Denning, Warren; Das, Suvendu; Guo, Siqi; Xu, Jun; Kappes, John C; Hel, Zdenek

2013-03-01

Lentiviral vectors are widely used as effective gene-delivery vehicles. Optimization of the conditions for efficient lentiviral transduction is of a high importance for a variety of research applications. Presence of positively charged polycations reduces the electrostatic repulsion forces between a negatively charged cell and an approaching enveloped lentiviral particle resulting in an increase in the transduction efficiency. Although a variety of polycations are commonly used to enhance the transduction with retroviruses, the relative effect of various types of polycations on the efficiency of transduction and on the potential bias in the determination of titer of lentiviral vectors is not fully understood. Here, we present data suggesting that DEAE-dextran provides superior results in enhancing lentiviral transduction of most tested cell lines and primary cell cultures. Specific type and source of serum affects the efficiency of transduction of target cell populations. Non-specific binding of enhanced green fluorescent protein (EGFP)-containing membrane aggregates in the presence of DEAE-dextran does not significantly affect the determination of the titer of EGFP-expressing lentiviral vectors. In conclusion, various polycations and types of sera should be tested when optimizing lentiviral transduction of target cell populations.

Enhancing Endosomal Escape of Transduced Proteins by Photochemical Internalisation

PubMed Central

Mellert, Kevin; Lamla, Markus; Scheffzek, Klaus; Wittig, Rainer; Kaufmann, Dieter

2012-01-01

Induced internalisation of functional proteins into cultured cells has become an important aspect in a rising number of in vitro and in vivo assays. The endo-lysosomal entrapment of the transduced proteins remains the major problem in all transduction protocols. In this study we compared the efficiency, cytotoxicity and protein targeting of different commercially available transduction reagents by transducing a well-studied fluorescently labelled protein (Atto488-bovine serum albumin) into cultured human sarcoma cells. The amount of internalised protein and toxicity differed between the different reagents, but the percentage of transduced cells was consistently high. Furthermore, in all protocols the signals of the transduced Atto488-BSA were predominantly punctual consistent with an endosomal localisation. To overcome the endosomal entrapment, the transduction protocols were combined with a photochemical internalisation (PCI) treatment. Using this combination revealed that an endosomal disruption is highly effective in cell penetrating peptide (CPP) mediated transduction, whereas lipid-mediated transductions lead to a lower signal spreading throughout the cytosol. No change in the signal distribution could be achieved in treatments using non-lipid polymers as a transduction reagent. Therefore, the combination of protein transduction protocols based on CPPs with the endosomolytic treatment PCI can facilitate protein transduction experiments in vitro. PMID:23285056

Enhancing endosomal escape of transduced proteins by photochemical internalisation.

PubMed

Mellert, Kevin; Lamla, Markus; Scheffzek, Klaus; Wittig, Rainer; Kaufmann, Dieter

2012-01-01

Induced internalisation of functional proteins into cultured cells has become an important aspect in a rising number of in vitro and in vivo assays. The endo-lysosomal entrapment of the transduced proteins remains the major problem in all transduction protocols. In this study we compared the efficiency, cytotoxicity and protein targeting of different commercially available transduction reagents by transducing a well-studied fluorescently labelled protein (Atto488-bovine serum albumin) into cultured human sarcoma cells. The amount of internalised protein and toxicity differed between the different reagents, but the percentage of transduced cells was consistently high. Furthermore, in all protocols the signals of the transduced Atto488-BSA were predominantly punctual consistent with an endosomal localisation. To overcome the endosomal entrapment, the transduction protocols were combined with a photochemical internalisation (PCI) treatment. Using this combination revealed that an endosomal disruption is highly effective in cell penetrating peptide (CPP) mediated transduction, whereas lipid-mediated transductions lead to a lower signal spreading throughout the cytosol. No change in the signal distribution could be achieved in treatments using non-lipid polymers as a transduction reagent. Therefore, the combination of protein transduction protocols based on CPPs with the endosomolytic treatment PCI can facilitate protein transduction experiments in vitro.

A dual-route cascaded model of reading by deaf adults: evidence for grapheme to viseme conversion.

PubMed

Elliott, Eeva A; Braun, Mario; Kuhlmann, Michael; Jacobs, Arthur M

2012-01-01

There is an ongoing debate whether deaf individuals access phonology when reading, and if so, what impact the ability to access phonology might have on reading achievement. However, the debate so far has been theoretically unspecific on two accounts: (a) the phonological units deaf individuals may have of oral language have not been specified and (b) there seem to be no explicit cognitive models specifying how phonology and other factors operate in reading by deaf individuals. We propose that deaf individuals have representations of the sublexical structure of oral-aural language which are based on mouth shapes and that these sublexical units are activated during reading by deaf individuals. We specify the sublexical units of deaf German readers as 11 "visemes" and incorporate the viseme set into a working model of single-word reading by deaf adults based on the dual-route cascaded model of reading aloud by Coltheart, Rastle, Perry, Langdon, and Ziegler (2001. DRC: A dual route cascaded model of visual word recognition and reading aloud. Psychological Review, 108, 204-256. doi: 10.1037//0033-295x.108.1.204). We assessed the indirect route of this model by investigating the "pseudo-homoviseme" effect using a lexical decision task in deaf German reading adults. We found a main effect of pseudo-homovisemy, suggesting that at least some deaf individuals do automatically access sublexical structure during single-word reading.

Adeno-Associated Virus Type 6 (AAV6) Vectors Mediate Efficient Transduction of Airway Epithelial Cells in Mouse Lungs Compared to That of AAV2 Vectors

PubMed Central

Halbert, Christine L.; Allen, James M.; Miller, A. Dusty

2001-01-01

Although vectors derived from adeno-associated virus type 2 (AAV2) promote gene transfer and expression in many somatic tissues, studies with animal models and cultured cells show that the apical surface of airway epithelia is resistant to transduction by AAV2 vectors. Approaches to increase transduction rates include increasing the amount of vector and perturbing the integrity of the epithelia. In this study, we explored the use of vectors based on AAV6 to increase transduction rates in airways. AAV vectors were made using combinations of rep, cap, and packaged genomes from AAV2 or AAV6. The packaged genomes encoded human placental alkaline phosphatase and contained terminal repeat sequences from AAV2 or AAV6. We found that transduction efficiency was primarily dependent on the source of Cap protein, defined here as the vector pseudotype. The AAV6 and AAV2 pseudotype vectors exhibited different tropisms in tissue-cultured cells, and cell transduction by AAV6 vectors was not inhibited by heparin, nor did they compete for entry in a transduction assay, indicating that AAV6 and AAV2 capsid bind different receptors. In vivo analysis of vectors showed that AAV2 pseudotype vectors gave high transduction rates in alveolar cells but much lower rates in the airway epithelium. In contrast, the AAV6 pseudotype vectors exhibited much more efficient transduction of epithelial cells in large and small airways, showing up to 80% transduction in some airways. These results, combined with our previous results showing lower immunogenicity of AAV6 than of AAV2 vectors, indicate that AAV6 vectors may provide significant advantages over AAV2 for gene therapy of lung diseases like cystic fibrosis. PMID:11413329

Ancestral gene reconstruction and synthesis of ancient rhodopsins in the laboratory.

PubMed

Chang, Belinda S W

2003-08-01

Laboratory synthesis of ancestral proteins offers an intriguing opportunity to study the past directly. The development of Bayesian methods to infer ancestral sequences, combined with advances in models of molecular evolution, and synthetic gene technology make this an increasingly promising approach in evolutionary studies of molecular function. Visual pigments form the first step in the biochemical cascade of events in the retina in all animals known to possess visual capabilities. In vertebrates, the necessity of spanning a dynamic range of light intensities of many orders of magnitude has given rise to two different types of photoreceptors, rods specialized for dim-light conditions, and cones for daylight and color vision. These photoreceptors contain different types of visual pigment genes. Reviewed here are methods of inferring ancestral sequences, chemical synthesis of artificial ancestral genes in the laboratory, and applications to the evolution of vertebrate visual systems and the experimental recreation of an archosaur rod visual pigment. The ancestral archosaurs gave rise to several notable lineages of diapsid reptiles, including the birds and the dinosaurs, and would have existed over 200 MYA. What little is known of their physiology comes from fossil remains, and inference based on the biology of their living descendants. Despite its age, an ancestral archosaur pigment was successfully recreated in the lab, and showed interesting properties of its wavelength sensitivity that may have implications for the visual capabilities of the ancestral archosaurs in dim light.

Membrane penetrating peptides greatly enhance baculovirus transduction efficiency into mammalian cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Hong-Zhang; Institute of Molecular Biology, Academia Sinica, Taipei 115, Taiwan, ROC; Wu, Carol P.

2011-02-11

Research highlights: {yields} Ligation of CTP with GP64 enhances baculovirus transduction into mammalian cells. {yields} Fusion of PTD with VP39 enhances baculovirus transduction into mammalian cells. {yields} CTP and PTD-carrying viruses improve the transduction of co-transduced baculoviruses. {yields} Virus entry and gene expression can be separate events in different cell types. -- Abstract: The baculovirus group of insect viruses is widely used for foreign gene introduction into mammalian cells for gene expression and protein production; however, the efficiency of baculovirus entry into mammalian cells is in general still low. In this study, two recombinant baculoviruses were engineered and their abilitymore » to improve viral entry was examined: (1) cytoplasmic transduction peptide (CTP) was fused with baculovirus envelope protein, GP64, to produce a cytoplasmic membrane penetrating baculovirus (vE-CTP); and (2) the protein transduction domain (PTD) of HIV TAT protein was fused with the baculovirus capsid protein VP39 to form a nuclear membrane penetrating baculovirus (vE-PTD). Transduction experiments showed that both viruses had better transduction efficiency than vE, a control virus that only expresses EGFP in mammalian cells. Interestingly, vE-CTP and vE-PTD were also able to improve the transduction efficiency of a co-transduced baculovirus, resulting in higher levels of gene expression. Our results have described new routes to further enhance the development of baculovirus as a tool for gene delivery into mammalian cells.« less

Disruption of Microtubules Post-Virus Entry Enhances Adeno-Associated Virus Vector Transduction

PubMed Central

Xiao, Ping-Jie; Mitchell, Angela M.; Huang, Lu; Li, Chengwen; Samulski, R. Jude

2016-01-01

Perinuclear retention of viral particles is a poorly understood phenomenon observed during many virus infections. In this study, we investigated whether perinuclear accumulation acts as a barrier to limit recombinant adeno-associated virus (rAAV) transduction. After nocodazole treatment to disrupt microtubules at microtubule-organization center (MT-MTOC) after virus entry, we observed higher rAAV transduction. To elucidate the role of MT-MTOC in rAAV infection and study its underlying mechanisms, we demonstrated that rAAV's perinuclear localization was retained by MT-MTOC with fluorescent analysis, and enhanced rAAV transduction from MT-MTOC disruption was dependent on the rAAV capsid's nuclear import signals. Interestingly, after knocking down RhoA or inhibiting its downstream effectors (ROCK and Actin), MT-MTOC disruption failed to increase rAAV transduction or nuclear entry. These data suggest that enhancement of rAAV transduction is the result of increased trafficking to the nucleus via the RhoA-ROCK-Actin pathway. Ten-fold higher rAAV transduction was also observed by disrupting MT-MTOC in brain, liver, and tumor in vivo. In summary, this study indicates that virus perinuclear accumulation at MT-MTOC is a barrier-limiting parameter for effective rAAV transduction and defines a novel defense mechanism by which host cells restrain viral invasion. PMID:26942476

Polymerase chain reaction-based detection of myc transduction in feline leukemia virus-infected cats.

PubMed

Sumi, Ryosuke; Miyake, Ariko; Endo, Taiji; Ohsato, Yoshiharu; Ngo, Minh Ha; Nishigaki, Kazuo

2018-04-01

Feline lymphomas are associated with the transduction and activation of cellular proto-oncogenes, such as c-myc, by feline leukemia virus (FeLV). We describe a polymerase chain reaction assay for detection of myc transduction usable in clinical diagnosis. The assay targets c-myc exons 2 and 3, which together result in a FeLV-specific fusion gene following c-myc transduction. When this assay was conducted on FeLV-infected feline tissues submitted for clinical diagnosis of tumors, myc transduction was detected in 14% of T-cell lymphoma/leukemias. This newly established system could become a useful diagnostic tool in veterinary medicine.

Tunable assembly of amyloid-forming peptides into nanosheets as a retrovirus carrier.

PubMed

Dai, Bin; Li, Dan; Xi, Wenhui; Luo, Fang; Zhang, Xiang; Zou, Man; Cao, Mi; Hu, Jun; Wang, Wenyuan; Wei, Guanghong; Zhang, Yi; Liu, Cong

2015-03-10

Using and engineering amyloid as nanomaterials are blossoming trends in bionanotechnology. Here, we show our discovery of an amyloid structure, termed "amyloid-like nanosheet," formed by a key amyloid-forming segment of Alzheimer's Aβ. Combining multiple biophysical and computational approaches, we proposed a structural model for the nanosheet that is formed by stacking the amyloid fibril spines perpendicular to the fibril axis. We further used the nanosheet for laboratorial retroviral transduction enhancement and directly visualized the presence of virus on the nanosheet surface by electron microscopy. Furthermore, based on our structural model, we designed nanosheet-forming peptides with different functionalities, elucidating the potential of rational design for amyloid-based materials with novel architecture and function.

Polyploidization without mitosis improves in vivo liver transduction with lentiviral vectors.

PubMed

Pichard, Virginie; Couton, Dominique; Desdouets, Chantal; Ferry, Nicolas

2013-02-01

Lentiviral vectors are efficient gene delivery vehicles for therapeutic and research applications. In contrast to oncoretroviral vectors, they are able to infect most nonproliferating cells. In the liver, induction of cell proliferation dramatically improved hepatocyte transduction using all types of retroviral vectors. However, the precise relationship between hepatocyte division and transduction efficiency has not been determined yet. Here we compared gene transfer efficiency in the liver after in vivo injection of recombinant lentiviral or Moloney murine leukemia viral (MoMuLV) vectors in hepatectomized rats treated or not with retrorsine, an alkaloid that blocks hepatocyte division and induces megalocytosis. Partial hepatectomy alone resulted in a similar increase in hepatocyte transduction using either vector. In retrorsine-treated and partially hepatectomized rats, transduction with MoMuLV vectors dropped dramatically. In contrast, we observed that retrorsine treatment combined with partial hepatectomy increased lentiviral transduction to higher levels than hepatectomy alone. Analysis of nuclear ploidy in single cells showed that a high level of transduction was associated with polyploidization. In conclusion, endoreplication could be exploited to improve the efficiency of liver-directed lentiviral gene therapy.

Polyploidization Without Mitosis Improves In Vivo Liver Transduction With Lentiviral Vectors

PubMed Central

Couton, Dominique; Desdouets, Chantal; Ferry, Nicolas

2013-01-01

Abstract Lentiviral vectors are efficient gene delivery vehicles for therapeutic and research applications. In contrast to oncoretroviral vectors, they are able to infect most nonproliferating cells. In the liver, induction of cell proliferation dramatically improved hepatocyte transduction using all types of retroviral vectors. However, the precise relationship between hepatocyte division and transduction efficiency has not been determined yet. Here we compared gene transfer efficiency in the liver after in vivo injection of recombinant lentiviral or Moloney murine leukemia viral (MoMuLV) vectors in hepatectomized rats treated or not with retrorsine, an alkaloid that blocks hepatocyte division and induces megalocytosis. Partial hepatectomy alone resulted in a similar increase in hepatocyte transduction using either vector. In retrorsine-treated and partially hepatectomized rats, transduction with MoMuLV vectors dropped dramatically. In contrast, we observed that retrorsine treatment combined with partial hepatectomy increased lentiviral transduction to higher levels than hepatectomy alone. Analysis of nuclear ploidy in single cells showed that a high level of transduction was associated with polyploidization. In conclusion, endoreplication could be exploited to improve the efficiency of liver-directed lentiviral gene therapy. PMID:23249390

Enhanced wound healing of tissue-engineered human corneas through altered phosphorylation of the CREB and AKT signal transduction pathways.

PubMed

Couture, Camille; Desjardins, Pascale; Zaniolo, Karine; Germain, Lucie; Guérin, Sylvain L

2018-06-01

The cornea is a transparent organ, highly specialized and unique that is continually subjected to abrasive forces and occasional mechanical or chemical trauma because of its anatomical localization. Upon injury, the extracellular matrix (ECM) rapidly changes to promote wound healing through integrin-dependent activation of specific signal transduction mediators whose contribution is to favor faster closure of the wound by altering the adhesive and migratory properties of the cells surrounding the damaged area. In this study, we exploited the human tissue-engineered cornea (hTECs) as a model to study the signal transduction pathways that participate to corneal wound healing. By exploiting both gene profiling and activated kinases arrays, we could demonstrate the occurrence of important alterations in the level of expression and activation of a few mediators from the PI3K/Akt and CREB pathways in response to the ECM remodeling taking place during wound healing of damaged hTECs. Pharmacological inhibition of CREB with C646 considerably accelerated wound closure compared to controls. This process was considerably accelerated further when both C646 and SC79, an Akt agonist, were added together to wounded hTECs. Therefore, our study demonstrate that proper corneal wound healing requires the activation of Akt together with the inhibition of CREB and that wound healing in vitro can be altered by the use of pharmacological inhibitors (such as C646) or agonists (such as SC79) of these mediators. Corneal wounds account for a large proportion of all visual disabilities in North America. To our knowledge, this is the first time that a tissue-engineered human cornea (hTEC) entirely produced using normal untransformed human cells is used as a biomaterial to study the signal transduction pathways that are critical to corneal wound healing. Through the use of this biomaterial, we demonstrated that human corneal epithelial cells engaged in wound healing reduce phosphorylation of the signal transduction mediator CREB while, in the mean time, they increase that of AKT. By increasing the activation of AKT together with a decrease in CREB activation, we could considerably reduce wound closure time in our punch-damaged hTECs. Considering the increasing interest given to the reconstruction of different types of tissues, we believe these results will have a strong impact on the field of tissue-engineering and biomaterials. Altering the activation status of the Akt and CREB proteins might prove to be a therapeutically interesting avenue and may also find applications in wound healing of other tissues beside the cornea, such as the skin. Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Defining a Novel Role for the Coxsackievirus and Adenovirus Receptor in Human Adenovirus Serotype 5 Transduction In Vitro in the Presence of Mouse Serum

PubMed Central

Lopez-Gordo, Estrella; Doszpoly, Andor; Duffy, Margaret R.; Coughlan, Lynda; Bradshaw, Angela C.; White, Katie M.; Denby, Laura; Nicklin, Stuart A.

2017-01-01

ABSTRACT Human adenoviral serotype 5 (HAdV-5) vectors have predominantly hepatic tropism when delivered intravascularly, resulting in immune activation and toxicity. Coagulation factor X (FX) binding to HAdV-5 mediates liver transduction and provides protection from virion neutralization in mice. FX is dispensable for liver transduction in mice lacking IgM antibodies or complement, suggesting that alternative transduction pathways exist. To identify novel factor(s) mediating HAdV-5 FX-independent entry, we investigated HAdV-5 transduction in vitro in the presence of serum from immunocompetent C57BL/6 or immunocompromised mice lacking IgM antibodies (Rag 2−/− and NOD-scid-gamma [NSG]). Sera from all three mouse strains enhanced HAdV-5 transduction of A549 cells. While inhibition of HAdV-5–FX interaction with FX-binding protein (X-bp) inhibited transduction in the presence of C57BL/6 serum, it had negligible effect on the enhanced transduction observed in the presence of Rag 2−/− or NSG serum. Rag 2−/− serum also enhanced transduction of the FX binding-deficient HAdV-5HVR5*HVR7*E451Q (AdT*). Interestingly, Rag 2−/− serum enhanced HAdV-5 transduction in a FX-independent manner in CHO-CAR and SKOV3-CAR cells (CHO or SKOV3 cells transfected to stably express human coxsackievirus and adenovirus receptor [CAR]). Additionally, blockade of CAR with soluble HAdV-5 fiber knob inhibited mouse serum-enhanced transduction in A549 cells, suggesting a potential role for CAR. Transduction of HAdV-5 KO1 and HAdV-5/F35 (CAR binding deficient) in the presence of Rag 2−/− serum was equivalent to that of HAdV-5, indicating that direct interaction between HAdV-5 and CAR is not required. These data suggest that FX may protect HAdV-5 from neutralization but has minimal contribution to HAdV-5 transduction in the presence of immunocompromised mouse serum. Alternatively, transduction occurs via an unidentified mouse serum protein capable of bridging HAdV-5 to CAR. IMPORTANCE The intravascular administration of HAdV-5 vectors can result in acute liver toxicity, transaminitis, thrombocytopenia, and injury to the vascular endothelium, illustrating challenges yet to overcome for HAdV-5-mediated systemic gene therapy. The finding that CAR and potentially an unidentified factor present in mouse serum might be important mediators of HAdV-5 transduction highlights that a better understanding of the complex biology defining the interplay between adenovirus immune recognition and cellular uptake mechanisms is still required. These findings are important to inform future optimization and development of HAdV-5-based adenoviral vectors for gene therapy. PMID:28381574

Thyrotropin-induced hydrogen peroxide production in FRTL-5 thyroid cells is mediated not by adenosine 3',5'-monophosphate, but by Ca2+ signaling followed by phospholipase-A2 activation and potentiated by an adenosine derivative.

PubMed

Kimura, T; Okajima, F; Sho, K; Kobayashi, I; Kondo, Y

1995-01-01

The production of hydrogen peroxide (H2O2) as an essential process for iodide organification is a key reaction in TSH-induced thyroid hormone synthesis. Here we characterize the signal transduction pathway involved in TSH-induced H2O2 production in FRTL-5 thyroid cells. At higher than 1 nM TSH, N6-(L-2-phenylisopropyl)adenosine (PIA), an adenosine receptor agonist having, by itself, no influence on H2O2 generation, potentiated this TSH action, whereas the TSH increase and PIA addition reduced cAMP accumulation. RO 20-1724, a phosphodiesterase inhibitor, amplified the TSH-induced cAMP accumulation, but did not change H2O2 generation in the whole range of TSH used. Ca(2+)-mobilizing agonists, GTP and ATP, also induced H2O2 production without stimulating cAMP accumulation. Chelation of intracellular Ca2+ markedly inhibited the TSH action, but intracellular Ca2+ increases by either thapsigargin or ionomycin mimicking it. All of the findings show the participation of Ca2+, but not cAMP, in the action of TSH. Desensitization of protein kinase-C (PKC) did not influence the receptor-mediated H2O2 production, suggesting the reduced importance of PKC activation compared to Ca2+ signaling to the reaction. A rise in intracellular Ca2+ independent of receptor activation also induced H2O2 production as well as arachidonate release, and both were potentiated by PIA. In addition, inhibitors of phospholipase-A2 and the arachidonate metabolic pathway depressed H2O2 generation, suggesting the participation of an arachidonate cascade in the Ca(2+)-dependent H2O2 production. Lipoxygenase inhibitors depressed the Ca2+ action without influencing arachidonate release, suggesting the involvement of a lipoxygenase product(s) of arachidonate in the Ca(2+)-signaling mechanism. In conclusion, in FRTL-5 cells, TSH-induced H2O2 production is mediated not by cAMP, but by the phospholipase-C/Ca2+ cascade, possibly followed by the Ca(2+)-dependent phospholipase-A2/arachidonate cascade. PIA amplifies TSH-induced H2O2 production at the steps of phospholipase-C and phospholipase-A2 activation in a pertussis toxin-sensitive manner.

Mutation in rod PDE6 linked to congenital stationary night blindness impairs the enzyme inhibition by its gamma-subunit.

PubMed

Muradov, Khakim G; Granovsky, Alexey E; Artemyev, Nikolai O

2003-03-25

Photoreceptor cGMP phosphodiesterase (PDE6) is the effector enzyme in the vertebrate visual transduction cascade. The activity of rod PDE6 catalytic alpha- and beta-subunits is blocked in the dark by two inhibitory Pgamma-subunits. The inhibition is released upon light-stimulation of photoreceptor cells. Mutation H258N in PDE6beta has been linked to congenital stationary night blindness (CSNB) in a large Danish family (Rambusch pedigree) (Gal, A., Orth, U., Baehr, W., Schwinger, E., and Rosenberg, T. (1994) Nat. Genet. 7, 64-67.) We have analyzed the consequences of this mutation for PDE6 function using a Pgamma-sensitive PDE6alpha'/PDE5 chimera, Chi16. Biochemical analysis of the H257N mutant, an equivalent of PDE6betaH258N, demonstrates that this substitution does not alter the ability of chimeric PDE to dimerize or the enzyme's catalytic properties. The sensitivity of H257N to a competitive inhibitor zaprinast was also unaffected. However, the mutant displayed a significant impairment in the inhibitory interaction with Pgamma, which was apparent from a approximately 20-fold increase in the K(i) value (46 nM) and incomplete maximal inhibition. The inhibitory defect of H257N is not due to perturbation of noncatalytic cGMP binding to the PDE6alpha' GAF domains. The noncatalytic cGMP-binding characteristics of the H257N mutant were similar to those of the parent PDE6alpha'/PDE5 chimera. Since rod PDE6 in the Rambusch CSNB is a catalytic heterodimer of the wild-type PDE6alpha and mutant PDE6beta, Chi16 and H257N were coexpressed, and a heterodimeric PDE, Chi16/H257N, was isolated. It displayed two Pgamma inhibitory sites with the K(i) values of 5 and 57 nM. Our results support the hypothesis that mutation H258N in PDE6beta causes CSNB through incomplete inhibition of PDE6 activity by Pgamma, which leads to desensitization of rod photoreceptors.

Complex computation in the retina

NASA Astrophysics Data System (ADS)

Deshmukh, Nikhil Rajiv

Elucidating the general principles of computation in neural circuits is a difficult problem requiring both a tractable model circuit as well as sophisticated measurement tools. This thesis advances our understanding of complex computation in the salamander retina and its underlying circuitry and furthers the development of advanced tools to enable detailed study of neural circuits. The retina provides an ideal model system for neural circuits in general because it is capable of producing complex representations of the visual scene, and both its inputs and outputs are accessible to the experimenter. Chapter 2 describes the biophysical mechanisms that give rise to the omitted stimulus response in retinal ganglion cells described in Schwartz et al., (2007) and Schwartz and Berry, (2008). The extra response to omitted flashes is generated at the input to bipolar cells, and is separable from the characteristic latency shift of the OSR apparent in ganglion cells, which must occur downstream in the circuit. Chapter 3 characterizes the nonlinearities at the first synapse of the ON pathway in response to high contrast flashes and develops a phenomenological model that captures the effect of synaptic activation and intracellular signaling dynamics on flash responses. This work is the first attempt to model the dynamics of the poorly characterized mGluR6 transduction cascade unique to ON bipolar cells, and explains the second lobe of the biphasic flash response. Complementary to the study of neural circuits, recent advances in wafer-scale photolithography have made possible new devices to measure the electrical and mechanical properties of neurons. Chapter 4 reports a novel piezoelectric sensor that facilitates the simultaneous measurement of electrical and mechanical signals in neural tissue. This technology could reveal the relationship between the electrical activity of neurons and their local mechanical environment, which is critical to the study of mechanoreceptors, neural development, and traumatic brain injury. Chapter 5 describes advances in the development, fabrication, and testing of a prototype silicon micropipette for patch clamp physiology. Nanoscale photolithography addresses some of the limitations of traditional glass patch electrodes, such as the rapid dialysis of the cell with internal solution, and provides a platform for integration of microfluidics and electronics into the device, which can enable novel experimental methodology.

Proteomic Identification of Differentially Expressed Proteins during Alfalfa (Medicago sativa L.) Flower Development.

PubMed

Chen, Lingling; Chen, Quanzhu; Zhu, Yanqiao; Hou, Longyu; Mao, Peisheng

2016-01-01

Flower development, pollination, and fertilization are important stages in the sexual reproduction process of plants; they are also critical steps in the control of seed formation and development. During alfalfa ( Medicago sativa L.) seed production, some distinct phenomena such as a low seed setting ratio, serious flower falling, and seed abortion commonly occur. However, the causes of these phenomena are complicated and largely unknown. An understanding of the mechanisms that regulate alfalfa flowering is important in order to increase seed yield. Hence, proteomic technology was used to analyze changes in protein expression during the stages of alfalfa flower development. Flower samples were collected at pre-pollination (S1), pollination (S2), and the post-pollination senescence period (S3). Twenty-four differentially expressed proteins were successfully identified, including 17 down-regulated in pollinated flowers, one up-regulated in pollinated and senesced flowers, and six up-regulated in senesced flowers. The largest proportions of the identified proteins were involved in metabolism, signal transduction, defense response, oxidation reduction, cell death, and programmed cell death (PCD). Their expression profiles demonstrated that energy metabolism, carbohydrate metabolism, and amino acid metabolism provided the nutrient foundation for pollination in alfalfa. Furthermore, there were three proteins involved in multiple metabolic pathways: dual specificity kinase splA-like protein (kinase splALs), carbonic anhydrase, and NADPH: quinone oxidoreductase-like protein. Expression patterns of these proteins indicated that MAPK cascades regulated multiple processes, such as signal transduction, stress response, and cell death. PCD also played an important role in the alfalfa flower developmental process, and regulated both pollination and flower senescence. The current study sheds some light on protein expression profiles during alfalfa flower development and contributes to the understanding of the basic molecular mechanisms during the alfalfa flowering process. These results may offer insight into potential strategies for improving seed yield, quality, and stress tolerance in alfalfa.

Conventional protein kinase C-α (PKC-α) and PKC-β negatively regulate RIG-I antiviral signal transduction.

PubMed

Maharaj, Natalya P; Wies, Effi; Stoll, Andrej; Gack, Michaela U

2012-02-01

Retinoic acid-inducible gene I (RIG-I) is a key sensor for viral RNA in the cytosol, and it initiates a signaling cascade that leads to the establishment of an interferon (IFN)-mediated antiviral state. Because of its integral role in immune signaling, RIG-I activity must be precisely controlled. Recent studies have shown that RIG-I CARD-dependent signaling function is regulated by the dynamic balance between phosphorylation and TRIM25-induced K₆₃-linked ubiquitination. While ubiquitination of RIG-I is critical for RIG-I's ability to induce an antiviral IFN response, phosphorylation of RIG-I at S₈ or T₁₇₀ suppresses RIG-I signal-transducing activity under normal conditions. Here, we not only further define the roles of S₈ and T₁₇₀ phosphorylation for controlling RIG-I activity but also identify conventional protein kinase C-α (PKC-α) and PKC-β as important negative regulators of the RIG-I signaling pathway. Mutational analysis indicated that while the phosphorylation of S₈ or T₁₇₀ potently inhibits RIG-I downstream signaling, the dephosphorylation of RIG-I at both residues is necessary for optimal TRIM25 binding and ubiquitination-mediated RIG-I activation. Furthermore, exogenous expression, gene silencing, and specific inhibitor treatment demonstrated that PKC-α/β are the primary kinases responsible for RIG-I S₈ and T₁₇₀ phosphorylation. Coimmunoprecipitation showed that PKC-α/β interact with RIG-I under normal conditions, leading to its phosphorylation, which suppresses TRIM25 binding, RIG-I CARD ubiquitination, and thereby RIG-I-mediated IFN induction. PKC-α/β double-knockdown cells exhibited markedly decreased S₈/T₁₇₀ phosphorylation levels of RIG-I and resistance to infection by vesicular stomatitis virus. Thus, these findings demonstrate that PKC-α/β-induced RIG-I phosphorylation is a critical regulatory mechanism for controlling RIG-I antiviral signal transduction under normal conditions.

Conventional Protein Kinase C-α (PKC-α) and PKC-β Negatively Regulate RIG-I Antiviral Signal Transduction

PubMed Central

Maharaj, Natalya P.; Wies, Effi; Stoll, Andrej

2012-01-01

Retinoic acid-inducible gene I (RIG-I) is a key sensor for viral RNA in the cytosol, and it initiates a signaling cascade that leads to the establishment of an interferon (IFN)-mediated antiviral state. Because of its integral role in immune signaling, RIG-I activity must be precisely controlled. Recent studies have shown that RIG-I CARD-dependent signaling function is regulated by the dynamic balance between phosphorylation and TRIM25-induced K63-linked ubiquitination. While ubiquitination of RIG-I is critical for RIG-I's ability to induce an antiviral IFN response, phosphorylation of RIG-I at S8 or T170 suppresses RIG-I signal-transducing activity under normal conditions. Here, we not only further define the roles of S8 and T170 phosphorylation for controlling RIG-I activity but also identify conventional protein kinase C-α (PKC-α) and PKC-β as important negative regulators of the RIG-I signaling pathway. Mutational analysis indicated that while the phosphorylation of S8 or T170 potently inhibits RIG-I downstream signaling, the dephosphorylation of RIG-I at both residues is necessary for optimal TRIM25 binding and ubiquitination-mediated RIG-I activation. Furthermore, exogenous expression, gene silencing, and specific inhibitor treatment demonstrated that PKC-α/β are the primary kinases responsible for RIG-I S8 and T170 phosphorylation. Coimmunoprecipitation showed that PKC-α/β interact with RIG-I under normal conditions, leading to its phosphorylation, which suppresses TRIM25 binding, RIG-I CARD ubiquitination, and thereby RIG-I-mediated IFN induction. PKC-α/β double-knockdown cells exhibited markedly decreased S8/T170 phosphorylation levels of RIG-I and resistance to infection by vesicular stomatitis virus. Thus, these findings demonstrate that PKC-α/β-induced RIG-I phosphorylation is a critical regulatory mechanism for controlling RIG-I antiviral signal transduction under normal conditions. PMID:22114345

Fast voltage-sensitive dye imaging of excitatory and inhibitory synaptic transmission in the rat granular retrosplenial cortex.

PubMed

Nixima, Ken'ichi; Okanoya, Kazuo; Ichinohe, Noritaka; Kurotani, Tohru

2017-09-01

Rodent granular retrosplenial cortex (GRS) has dense connections between the anterior thalamic nuclei (ATN) and hippocampal formation. GRS superficial pyramidal neurons exhibit distinctive late spiking (LS) firing property and form patchy clusters with prominent apical dendritic bundles. The aim of this study was to investigate spatiotemporal dynamics of signal transduction in the GRS induced by ATN afferent stimulation by using fast voltage-sensitive dye imaging in rat brain slices. In coronal slices, layer 1a stimulation, which presumably activated thalamic fibers, evoked propagation of excitatory synaptic signals from layers 2-4 to layers 5-6 in a direction perpendicular to the layer axis, followed by transverse signal propagation within each layer. In the presence of ionotropic glutamate receptor antagonists, inhibitory responses were observed in superficial layers, induced by direct activation of inhibitory interneurons in layer 1. In horizontal slices, excitatory signals in deep layers propagated transversely mainly from posterior to anterior via superficial layers. Cortical inhibitory responses upon layer 1a stimulation in horizontal slices were weaker than those in the coronal slices. Observed differences between coronal and horizontal planes suggest anisotropy of the intracortical circuitry. In conclusion, ATN inputs are processed differently in coronal and horizontal planes of the GRS and then conveyed to other cortical areas. In both planes, GRS superficial layers play an important role in signal propagation, which suggests that superficial neuronal cascade is crucial in the integration of multiple information sources. NEW & NOTEWORTHY Superficial neurons in the rat granular retrosplenial cortex (GRS) show distinctive late-spiking (LS) firing property. However, little is known about spatiotemporal dynamics of signal transduction in the GRS. We demonstrated LS neuron network relaying thalamic inputs to deep layers and anisotropic distribution of inhibition between coronal and horizontal planes. Since deep layers of the GRS receive inputs from the subiculum, GRS circuits may work as an integrator of multiple sources such as sensory and memory information. Copyright © 2017 the American Physiological Society.

Discrete diffusion models to study the effects of Mg2+ concentration on the PhoPQ signal transduction system

PubMed Central

2010-01-01

Background The challenge today is to develop a modeling and simulation paradigm that integrates structural, molecular and genetic data for a quantitative understanding of physiology and behavior of biological processes at multiple scales. This modeling method requires techniques that maintain a reasonable accuracy of the biological process and also reduces the computational overhead. This objective motivates the use of new methods that can transform the problem from energy and affinity based modeling to information theory based modeling. To achieve this, we transform all dynamics within the cell into a random event time, which is specified through an information domain measure like probability distribution. This allows us to use the “in silico” stochastic event based modeling approach to find the molecular dynamics of the system. Results In this paper, we present the discrete event simulation concept using the example of the signal transduction cascade triggered by extra-cellular Mg2+ concentration in the two component PhoPQ regulatory system of Salmonella Typhimurium. We also present a model to compute the information domain measure of the molecular transport process by estimating the statistical parameters of inter-arrival time between molecules/ions coming to a cell receptor as external signal. This model transforms the diffusion process into the information theory measure of stochastic event completion time to get the distribution of the Mg2+ departure events. Using these molecular transport models, we next study the in-silico effects of this external trigger on the PhoPQ system. Conclusions Our results illustrate the accuracy of the proposed diffusion models in explaining the molecular/ionic transport processes inside the cell. Also, the proposed simulation framework can incorporate the stochasticity in cellular environments to a certain degree of accuracy. We expect that this scalable simulation platform will be able to model more complex biological systems with reasonable accuracy to understand their temporal dynamics. PMID:21143785

Cell cycle control, checkpoint mechanisms, and genotoxic stress.

PubMed Central

Shackelford, R E; Kaufmann, W K; Paules, R S

1999-01-01

The ability of cells to maintain genomic integrity is vital for cell survival and proliferation. Lack of fidelity in DNA replication and maintenance can result in deleterious mutations leading to cell death or, in multicellular organisms, cancer. The purpose of this review is to discuss the known signal transduction pathways that regulate cell cycle progression and the mechanisms cells employ to insure DNA stability in the face of genotoxic stress. In particular, we focus on mammalian cell cycle checkpoint functions, their role in maintaining DNA stability during the cell cycle following exposure to genotoxic agents, and the gene products that act in checkpoint function signal transduction cascades. Key transitions in the cell cycle are regulated by the activities of various protein kinase complexes composed of cyclin and cyclin-dependent kinase (Cdk) molecules. Surveillance control mechanisms that check to ensure proper completion of early events and cellular integrity before initiation of subsequent events in cell cycle progression are referred to as cell cycle checkpoints and can generate a transient delay that provides the cell more time to repair damage before progressing to the next phase of the cycle. A variety of cellular responses are elicited that function in checkpoint signaling to inhibit cyclin/Cdk activities. These responses include the p53-dependent and p53-independent induction of Cdk inhibitors and the p53-independent inhibitory phosphorylation of Cdk molecules themselves. Eliciting proper G1, S, and G2 checkpoint responses to double-strand DNA breaks requires the function of the Ataxia telangiectasia mutated gene product. Several human heritable cancer-prone syndromes known to alter DNA stability have been found to have defects in checkpoint surveillance pathways. Exposures to several common sources of genotoxic stress, including oxidative stress, ionizing radiation, UV radiation, and the genotoxic compound benzo[a]pyrene, elicit cell cycle checkpoint responses that show both similarities and differences in their molecular signaling. Images Figure 3 PMID:10229703

A mass spectrometry-based proteomic analysis of Homer2-interacting proteins in the mouse brain.

PubMed

Goulding, Scott P; Szumlinski, Karen K; Contet, Candice; MacCoss, Michael J; Wu, Christine C

2017-08-23

In the brain, the Homer protein family modulates excitatory signal transduction and receptor plasticity through interactions with other proteins in dendritic spines. Homer proteins are implicated in a variety of psychiatric disorders such as schizophrenia and addiction. Since long Homers serve as scaffolding proteins, identifying their interacting partners is an important first step in understanding their biological function and could help to guide the design of new therapeutic strategies. The present study set out to document Homer2-interacting proteins in the mouse brain using a co-immunoprecipitation-based mass spectrometry approach where Homer2 knockout samples were used to filter out non-specific interactors. We found that in the mouse brain, Homer2 interacts with a limited subset of its previously reported interacting partners (3 out of 31). Importantly, we detected an additional 15 novel Homer2-interacting proteins, most of which are part of the N-methyl-D-aspartate receptor signaling pathway. These results corroborate the central role Homer2 plays in glutamatergic transmission and expand the network of proteins potentially contributing to the behavioral abnormalities associated with altered Homer2 expression. Long Homer proteins are scaffolding proteins that regulate signal transduction in neurons. Identifying their interacting partners is key to understanding their function. We used co-immunoprecipitation in combination with mass spectrometry to establish the first comprehensive list of Homer2-interacting partners in the mouse brain. The specificity of interactions was evaluated using Homer2 knockout brain tissue as a negative control. The set of proteins that we identified minimally overlaps with previously reported interacting partners of Homer2; however, we identified novel interactors that are part of a signaling cascade activated by glutamatergic transmission, which improves our mechanistic understanding of the role of Homer2 in behavior. Copyright © 2017 Elsevier B.V. All rights reserved.

Cationic Polyamidoamine Dendrimers as Modulators of EGFR Signaling In Vitro and In Vivo

PubMed Central

Akhtar, Saghir; Al-Zaid, Bashayer; El-Hashim, Ahmed Z.; Chandrasekhar, Bindu; Attur, Sreeja; Yousif, Mariam H. M.; Benter, Ibrahim F.

2015-01-01

Cationic polyamidoamine (PAMAM) dendrimers are branch-like spherical polymers being investigated for a variety of applications in nanomedicine including nucleic acid drug delivery. Emerging evidence suggests they exhibit intrinsic biological and toxicological effects but little is known of their interactions with signal transduction pathways. We previously showed that the activated (fragmented) generation (G) 6 PAMAM dendrimer, Superfect (SF), stimulated epidermal growth factor receptor (EGFR) tyrosine kinase signaling—an important signaling cascade that regulates cell growth, survival and apoptosis- in cultured human embryonic kidney (HEK 293) cells. Here, we firstly studied the in vitro effects of Polyfect (PF), a non-activated (intact) G6 PAMAM dendrimer, on EGFR tyrosine kinase signaling via extracellular-regulated kinase 1/2 (ERK1/2) and p38 mitogen-activated protein kinase (MAPK) in cultured HEK 293 cells and then compared the in vivo effects of a single administration (10mg/kg i.p) of PF or SF on EGFR signaling in the kidneys of normal and diabetic male Wistar rats. Polyfect exhibited a dose- and time-dependent inhibition of EGFR, ERK1/2 and p38 MAPK phosphorylation in HEK-293 cells similar to AG1478, a selective EGFR inhibitor. Administration of dendrimers to non-diabetic or diabetic animals for 24h showed that PF inhibited whereas SF stimulated EGFR phosphorylation in the kidneys of both sets of animals. PF-mediated inhibition of EGFR phosphorylation as well as SF or PF-mediated apoptosis in HEK 293 cells could be significantly reversed by co-treatment with antioxidants such as tempol implying that both these effects involved an oxidative stress-dependent mechanism. These results show for the first time that SF and PF PAMAM dendrimers can differentially modulate the important EGFR signal transduction pathway in vivo and may represent a novel class of EGFR modulators. These findings could have important clinical implications for the use of PAMAM dendrimers in nanomedicine. PMID:26167903

Discrete diffusion models to study the effects of Mg2+ concentration on the PhoPQ signal transduction system.

PubMed

Ghosh, Preetam; Ghosh, Samik; Basu, Kalyan; Das, Sajal K; Zhang, Chaoyang

2010-12-01

The challenge today is to develop a modeling and simulation paradigm that integrates structural, molecular and genetic data for a quantitative understanding of physiology and behavior of biological processes at multiple scales. This modeling method requires techniques that maintain a reasonable accuracy of the biological process and also reduces the computational overhead. This objective motivates the use of new methods that can transform the problem from energy and affinity based modeling to information theory based modeling. To achieve this, we transform all dynamics within the cell into a random event time, which is specified through an information domain measure like probability distribution. This allows us to use the "in silico" stochastic event based modeling approach to find the molecular dynamics of the system. In this paper, we present the discrete event simulation concept using the example of the signal transduction cascade triggered by extra-cellular Mg2+ concentration in the two component PhoPQ regulatory system of Salmonella Typhimurium. We also present a model to compute the information domain measure of the molecular transport process by estimating the statistical parameters of inter-arrival time between molecules/ions coming to a cell receptor as external signal. This model transforms the diffusion process into the information theory measure of stochastic event completion time to get the distribution of the Mg2+ departure events. Using these molecular transport models, we next study the in-silico effects of this external trigger on the PhoPQ system. Our results illustrate the accuracy of the proposed diffusion models in explaining the molecular/ionic transport processes inside the cell. Also, the proposed simulation framework can incorporate the stochasticity in cellular environments to a certain degree of accuracy. We expect that this scalable simulation platform will be able to model more complex biological systems with reasonable accuracy to understand their temporal dynamics.

Proteomic Identification of Differentially Expressed Proteins during Alfalfa (Medicago sativa L.) Flower Development

PubMed Central

Chen, Lingling; Chen, Quanzhu; Zhu, Yanqiao; Hou, Longyu; Mao, Peisheng

2016-01-01

Flower development, pollination, and fertilization are important stages in the sexual reproduction process of plants; they are also critical steps in the control of seed formation and development. During alfalfa (Medicago sativa L.) seed production, some distinct phenomena such as a low seed setting ratio, serious flower falling, and seed abortion commonly occur. However, the causes of these phenomena are complicated and largely unknown. An understanding of the mechanisms that regulate alfalfa flowering is important in order to increase seed yield. Hence, proteomic technology was used to analyze changes in protein expression during the stages of alfalfa flower development. Flower samples were collected at pre-pollination (S1), pollination (S2), and the post-pollination senescence period (S3). Twenty-four differentially expressed proteins were successfully identified, including 17 down-regulated in pollinated flowers, one up-regulated in pollinated and senesced flowers, and six up-regulated in senesced flowers. The largest proportions of the identified proteins were involved in metabolism, signal transduction, defense response, oxidation reduction, cell death, and programmed cell death (PCD). Their expression profiles demonstrated that energy metabolism, carbohydrate metabolism, and amino acid metabolism provided the nutrient foundation for pollination in alfalfa. Furthermore, there were three proteins involved in multiple metabolic pathways: dual specificity kinase splA-like protein (kinase splALs), carbonic anhydrase, and NADPH: quinone oxidoreductase-like protein. Expression patterns of these proteins indicated that MAPK cascades regulated multiple processes, such as signal transduction, stress response, and cell death. PCD also played an important role in the alfalfa flower developmental process, and regulated both pollination and flower senescence. The current study sheds some light on protein expression profiles during alfalfa flower development and contributes to the understanding of the basic molecular mechanisms during the alfalfa flowering process. These results may offer insight into potential strategies for improving seed yield, quality, and stress tolerance in alfalfa. PMID:27757120

Toll-like receptor 4 mediates inflammatory signaling by bacterial lipopolysaccharide in human hepatic stellate cells.

PubMed

Paik, Yong-Han; Schwabe, Robert F; Bataller, Ramón; Russo, Maria P; Jobin, Christian; Brenner, David A

2003-05-01

Bacterial lipopolysaccharide (LPS) stimulates Kupffer cells and participates in the pathogenesis of alcohol-induced liver injury. However, it is unknown whether LPS directly affects hepatic stellate cells (HSCs), the main fibrogenic cell type in the injured liver. This study characterizes LPS-induced signal transduction and proinflammatory gene expression in activated human HSCs. Culture-activated HSCs and HSCs isolated from patients with hepatitis C virus-induced cirrhosis express LPS-associated signaling molecules, including CD14, toll-like receptor (TLR) 4, and MD2. Stimulation of culture-activated HSCs with LPS results in a rapid and marked activation of NF-kappaB, as assessed by in vitro kinase assays for IkappaB kinase (IKK), IkappaBalpha steady-state levels, p65 nuclear translocation, NF-kappaB-dependent luciferase reporter gene assays, and electrophoretic mobility shift assays. Lipid A induces NF-kappaB activation in a similar manner. Both LPS- and lipid A-induced NF-kappaB activation is blocked by preincubation with either anti-TLR4 blocking antibody (HTA125) or Polymyxin B. Lipid A induces NF-kappaB activation in HSCs from TLR4-sufficient (C3H/OuJ) mice but not from TLR4-deficient (C3H/HeJ) mice. LPS also activates c-Jun N-terminal kinase (JNK), as assessed by in vitro kinase assays. LPS up-regulates IL-8 and MCP-1 gene expression and secretion. LPS-induced IL-8 secretion is completely inhibited by the IkappaB super repressor (Ad5IkappaB) and partially inhibited by a specific JNK inhibitor, SP600125. LPS also up-regulates cell surface expression of ICAM-1 and VCAM-1. In conclusion, human activated HSCs utilize components of TLR4 signal transduction cascade to stimulate NF-kappaB and JNK and up-regulate chemokines and adhesion molecules. Thus, HSCs are a potential mediator of LPS-induced liver injury.

Dynamical Modeling of the Moth Pheromone-Sensitive Olfactory Receptor Neuron within Its Sensillar Environment

PubMed Central

Gu, Yuqiao; Rospars, Jean-Pierre

2011-01-01

In insects, olfactory receptor neurons (ORNs), surrounded with auxiliary cells and protected by a cuticular wall, form small discrete sensory organs – the sensilla. The moth pheromone-sensitive sensillum is a well studied example of hair-like sensillum that is favorable to both experimental and modeling investigations. The model presented takes into account both the molecular processes of ORNs, i.e. the biochemical reactions and ionic currents giving rise to the receptor potential, and the cellular organization and compartmentalization of the organ represented by an electrical circuit. The number of isopotential compartments needed to describe the long dendrite bearing pheromone receptors was determined. The transduction parameters that must be modified when the number of compartments is increased were identified. This model reproduces the amplitude and time course of the experimentally recorded receptor potential. A first complete version of the model was analyzed in response to pheromone pulses of various strengths. It provided a quantitative description of the spatial and temporal evolution of the pheromone-dependent conductances, currents and potentials along the outer dendrite and served to determine the contribution of the various steps in the cascade to its global sensitivity. A second simplified version of the model, utilizing a single depolarizing conductance and leak conductances for repolarizing the ORN, was derived from the first version. It served to analyze the effects on the sensory properties of varying the electrical parameters and the size of the main sensillum parts. The consequences of the results obtained on the still uncertain mechanisms of olfactory transduction in moth ORNs – involvement or not of G-proteins, role of chloride and potassium currents – are discussed as well as the optimality of the sensillum organization, the dependence of biochemical parameters on the neuron spatial extension and the respective contributions of the biochemical and electrical parameters to the overall neuron response. PMID:21399691

Analysis of cellular signal transduction from an information theoretic approach.

PubMed

Uda, Shinsuke; Kuroda, Shinya

2016-03-01

Signal transduction processes the information of various cellular functions, including cell proliferation, differentiation, and death. The information for controlling cell fate is transmitted by concentrations of cellular signaling molecules. However, how much information is transmitted in signaling pathways has thus far not been investigated. Shannon's information theory paves the way to quantitatively analyze information transmission in signaling pathways. The theory has recently been applied to signal transduction, and mutual information of signal transduction has been determined to be a measure of information transmission. We review this work and provide an overview of how signal transduction transmits informational input and exerts biological output. Copyright © 2015 Elsevier Ltd. All rights reserved.

Microfluidic Transduction Harnesses Mass Transport Principles to Enhance Gene Transfer Efficiency.

PubMed

Tran, Reginald; Myers, David R; Denning, Gabriela; Shields, Jordan E; Lytle, Allison M; Alrowais, Hommood; Qiu, Yongzhi; Sakurai, Yumiko; Li, William C; Brand, Oliver; Le Doux, Joseph M; Spencer, H Trent; Doering, Christopher B; Lam, Wilbur A

2017-10-04

Ex vivo gene therapy using lentiviral vectors (LVs) is a proven approach to treat and potentially cure many hematologic disorders and malignancies but remains stymied by cumbersome, cost-prohibitive, and scale-limited production processes that cannot meet the demands of current clinical protocols for widespread clinical utilization. However, limitations in LV manufacture coupled with inefficient transduction protocols requiring significant excess amounts of vector currently limit widespread implementation. Herein, we describe a microfluidic, mass transport-based approach that overcomes the diffusion limitations of current transduction platforms to enhance LV gene transfer kinetics and efficiency. This novel ex vivo LV transduction platform is flexible in design, easy to use, scalable, and compatible with standard cell transduction reagents and LV preparations. Using hematopoietic cell lines, primary human T cells, primary hematopoietic stem and progenitor cells (HSPCs) of both murine (Sca-1 + ) and human (CD34 + ) origin, microfluidic transduction using clinically processed LVs occurs up to 5-fold faster and requires as little as one-twentieth of LV. As an in vivo validation of the microfluidic-based transduction technology, HSPC gene therapy was performed in hemophilia A mice using limiting amounts of LV. Compared to the standard static well-based transduction protocols, only animals transplanted with microfluidic-transduced cells displayed clotting levels restored to normal. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Flow and heat transfer experiments in the turbine airfoil/endwall region

NASA Astrophysics Data System (ADS)

Chung, Jin Taek

An experimental investigation of the three-dimensional flow and heat transfer near the junction between the endwall and suction wall of a gas turbine was performed. A large-scale, two-half-blade facility which simulates a turbine cascade was introduced. The simulator consists of two large half-blade sections, one wall simulating the pressure surface and the other wall simulating the suction surface. The advantage of this configuration is that the features of the secondary flow are large, because of the relatively large test section, and the flow is easily accessible with probes. Qualification of this simulator was by comparison to a multi-blade cascade flow. Various flow visualization techniques--oil and lampblack, ink and oil of wintergeeen, a single tuft probe, and a tuft grid--were employed to confirm that the important features of the cascade flow were replicated in this simulator. The triangular region on the suction surface, which was affected by the passage vortex, and the endwall secondary crossflow were observed by shear stress visualization and the liquid crystal measurement techniques. In order to investigate the effects of the turbulence level on the secondary flow in a turbine passage, a turbulence generator, designed to reproduce the characteristics of a combustor exit flow, was built. The generator was designed not only to generate a high turbulence level but to produce three main features of a combustor exit flow. The generator produced a turbulence intensity level of about 10 percent and an integral length scale of 5 centimeters. It was observed that the endwall secondary flow, including the passage vortex, is not significantly influenced by freestream turbulence levels up to 10 percent. A flow management technique using a boundary layer fence designed to reduce some harmful effects of secondary flow in the endwall region of a turbine passage was introduced. The boundary layer fence is effective in changing the passage of the vortex and reducing the influence of the vortex near the suction wall. The fence was even more effective in reducing secondary flows for high levels of freestream turbulence (approximately 10 percent).

Delivery and evaluation of recombinant adeno-associated viral vectors in the equine distal extremity for the treatment of laminitis.

PubMed

Mason, J B; Gurda, B L; Van Wettere, A; Engiles, J B; Wilson, J M; Richardson, D W

2017-01-01

Our long-term aim is to develop a gene therapy approach for the prevention of laminitis in the contralateral foot of horses with major musculoskeletal injuries and non-weightbearing lameness. The goal of this study was to develop a practical method to efficiently deliver therapeutic proteins deep within the equine foot. Randomised in vivo experiment. We used recombinant adeno-associated viral vectors (rAAVs) to deliver marker genes using regional limb perfusion through the palmar digital artery of the horse. Vector serotypes rAAV2/1, 2/8 and 2/9 all successfully transduced equine foot tissues and displayed similar levels and patterns of transduction. The regional distribution of transduction within the foot decreased with decreasing vector dose. The highest transduction values were seen in the sole and coronary regions and the lowest transduction values were detected in the dorsal hoof-wall region. The use of a surfactant-enriched vector diluent increased regional distribution of the vector and improved the transduction in the hoof-wall region. The hoof-wall region of the foot, which exhibited the lowest levels of transduction using saline as the vector diluent, displayed a dramatic increase in transduction when surfactant was included in the vector diluent (9- to 81-fold increase). In transduced tissues, no significant difference was observed between promoters (chicken β-actin vs. cytomegalovirus) for gene expression. All horses tested for vector-neutralising antibodies were positive for serotype-specific neutralising antibodies to rAAV2/5. The current experiments demonstrate that transgenes can be successfully delivered to the equine distal extremity using rAAV vectors and that serotypes 2/8, 2/9 and 2/1 can successfully transduce tissues of the equine foot. When the vector was diluted with surfactant-containing saline, the level of transduction increased dramatically. The increased level of transduction due to the addition of surfactant also improved the distribution pattern of transduction. © 2015 EVJ Ltd.

Fyn kinase genetic ablation causes structural abnormalities in mature retina and defective Müller cell function.

PubMed

Chavez-Solano, Marbella; Ibarra-Sanchez, Alfredo; Treviño, Mario; Gonzalez-Espinosa, Claudia; Lamas, Monica

2016-04-01

Fyn kinase is widely expressed in neuronal and glial cells of the brain, where it exerts multiple functional roles that affect fundamental physiological processes. The aim of our study was to investigate the, so far unknown, functional role of Fyn in the retina. We report that Fyn is expressed, in vivo, in a subpopulation of Müller glia. We used a mouse model of Fyn genetic ablation and Müller-enriched primary cultures to demonstrate that Fyn deficiency induces morphological alterations in the mature retina, a reduction in the thickness of the outer and inner nuclear layers and alterations in postnatal Müller cell physiology. These include shortening of Müller cell processes, a decrease in cell proliferation, inactivation of the Akt signal transduction pathway, a reduced number of focal adhesions points and decreased adhesion of these cells to the ECM. As abnormalities in Müller cell physiology have been previously associated to a compromised retinal function we evaluated behavioral responses to visual stimulation. Our results associate Fyn deficiency with impaired visual optokinetic responses under scotopic and photopic light conditions. Our study reveals novel roles for Fyn kinase in retinal morphology and Müller cell physiology and suggests that Fyn is required for optimal visual processing. Copyright © 2016 Elsevier Inc. All rights reserved.

Genetically fluorescent melanoma bone and organ metastasis models.

PubMed

Yang, M; Jiang, P; An, Z; Baranov, E; Li, L; Hasegawa, S; Al-Tuwaijri, M; Chishima, T; Shimada, H; Moossa, A R; Hoffman, R M

1999-11-01

We report here the establishment and metastatic properties of bright, highly stable, green fluorescent protein (GFP) expression transductants of the B16 mouse malignant melanoma cell line and the LOX human melanoma line. The highly fluorescent malignant melanoma cell lines allowed the visualization of skeletal and multiorgan metastases after i.v. injection of B16 cells in C57BL/6 mice and intradermal injection of LOX cells in nude mice. The melanoma cell lines were transduced with the pLEIN expression retroviral vector containing the GFP and neomycin resistance genes. Stable B16F0 and LOX clones expressing high levels of GFP were selected stepwise in vitro in levels of G418 of up to 800 microg/ml. Extensive bone and bone marrow metastases of B16F0 were visualized by GFP expression when the animals were sacrificed 3 weeks after cell implantation. Metastases for both cell lines were visualized in many organs, including the brain, lung, pleural membrane, liver, kidney, adrenal gland, lymph nodes, skeleton, muscle, and skin by GFP fluorescence. This is the first observation of experimental skeletal metastases of melanoma, which was made possible by GFP expression. These models should facilitate future studies of the mechanism and therapy of bone and multiorgan metastasis of melanoma.

Object segmentation controls image reconstruction from natural scenes

PubMed Central

2017-01-01

The structure of the physical world projects images onto our eyes. However, those images are often poorly representative of environmental structure: well-defined boundaries within the eye may correspond to irrelevant features of the physical world, while critical features of the physical world may be nearly invisible at the retinal projection. The challenge for the visual cortex is to sort these two types of features according to their utility in ultimately reconstructing percepts and interpreting the constituents of the scene. We describe a novel paradigm that enabled us to selectively evaluate the relative role played by these two feature classes in signal reconstruction from corrupted images. Our measurements demonstrate that this process is quickly dominated by the inferred structure of the environment, and only minimally controlled by variations of raw image content. The inferential mechanism is spatially global and its impact on early visual cortex is fast. Furthermore, it retunes local visual processing for more efficient feature extraction without altering the intrinsic transduction noise. The basic properties of this process can be partially captured by a combination of small-scale circuit models and large-scale network architectures. Taken together, our results challenge compartmentalized notions of bottom-up/top-down perception and suggest instead that these two modes are best viewed as an integrated perceptual mechanism. PMID:28827801

Modeling the infection dynamics of bacteriophages in enteric Escherichia coli: estimating the contribution of transduction to antimicrobial gene spread.

PubMed

Volkova, Victoriya V; Lu, Zhao; Besser, Thomas; Gröhn, Yrjö T

2014-07-01

Animal-associated bacterial communities are infected by bacteriophages, although the dynamics of these infections are poorly understood. Transduction by bacteriophages may contribute to transfer of antimicrobial resistance genes, but the relative importance of transduction among other gene transfer mechanisms is unknown. We therefore developed a candidate deterministic mathematical model of the infection dynamics of enteric coliphages in commensal Escherichia coli in the large intestine of cattle. We assumed the phages were associated with the intestine and were predominantly temperate. Model simulations demonstrated how, given the bacterial ecology and infection dynamics, most (>90%) commensal enteric E. coli bacteria may become lysogens of enteric coliphages during intestinal transit. Using the model and the most liberal assumptions about transduction efficiency and resistance gene frequency, we approximated the upper numerical limits ("worst-case scenario") of gene transfer through specialized and generalized transduction in E. coli by enteric coliphages when the transduced genetic segment is picked at random. The estimates were consistent with a relatively small contribution of transduction to lateral gene spread; for example, generalized transduction delivered the chromosomal resistance gene to up to 8 E. coli bacteria/hour within the population of 1.47 × 10(8) E. coli bacteria/liter luminal contents. In comparison, the plasmidic blaCMY-2 gene carried by ~2% of enteric E. coli was transferred by conjugation at a rate at least 1.4 × 10(3) times greater than our generalized transduction estimate. The estimated numbers of transductants varied nonlinearly depending on the ecology of bacteria available for phages to infect, that is, on the assumed rates of turnover and replication of enteric E. coli. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Modeling the Infection Dynamics of Bacteriophages in Enteric Escherichia coli: Estimating the Contribution of Transduction to Antimicrobial Gene Spread

PubMed Central

Lu, Zhao; Besser, Thomas; Gröhn, Yrjö T.

2014-01-01

Animal-associated bacterial communities are infected by bacteriophages, although the dynamics of these infections are poorly understood. Transduction by bacteriophages may contribute to transfer of antimicrobial resistance genes, but the relative importance of transduction among other gene transfer mechanisms is unknown. We therefore developed a candidate deterministic mathematical model of the infection dynamics of enteric coliphages in commensal Escherichia coli in the large intestine of cattle. We assumed the phages were associated with the intestine and were predominantly temperate. Model simulations demonstrated how, given the bacterial ecology and infection dynamics, most (>90%) commensal enteric E. coli bacteria may become lysogens of enteric coliphages during intestinal transit. Using the model and the most liberal assumptions about transduction efficiency and resistance gene frequency, we approximated the upper numerical limits (“worst-case scenario”) of gene transfer through specialized and generalized transduction in E. coli by enteric coliphages when the transduced genetic segment is picked at random. The estimates were consistent with a relatively small contribution of transduction to lateral gene spread; for example, generalized transduction delivered the chromosomal resistance gene to up to 8 E. coli bacteria/hour within the population of 1.47 × 108 E. coli bacteria/liter luminal contents. In comparison, the plasmidic blaCMY-2 gene carried by ∼2% of enteric E. coli was transferred by conjugation at a rate at least 1.4 × 103 times greater than our generalized transduction estimate. The estimated numbers of transductants varied nonlinearly depending on the ecology of bacteria available for phages to infect, that is, on the assumed rates of turnover and replication of enteric E. coli. PMID:24814786

Light-Mediated Kinetic Control Reveals the Temporal Effect of the Raf/MEK/ERK Pathway in PC12 Cell Neurite Outgrowth

PubMed Central

Zhang, Kai; Duan, Liting; Ong, Qunxiang; Lin, Ziliang; Varman, Pooja Mahendra; Sung, Kijung; Cui, Bianxiao

2014-01-01

It has been proposed that differential activation kinetics allows cells to use a common set of signaling pathways to specify distinct cellular outcomes. For example, nerve growth factor (NGF) and epidermal growth factor (EGF) induce different activation kinetics of the Raf/MEK/ERK signaling pathway and result in differentiation and proliferation, respectively. However, a direct and quantitative linkage between the temporal profile of Raf/MEK/ERK activation and the cellular outputs has not been established due to a lack of means to precisely perturb its signaling kinetics. Here, we construct a light-gated protein-protein interaction system to regulate the activation pattern of the Raf/MEK/ERK signaling pathway. Light-induced activation of the Raf/MEK/ERK cascade leads to significant neurite outgrowth in rat PC12 pheochromocytoma cell lines in the absence of growth factors. Compared with NGF stimulation, light stimulation induces longer but fewer neurites. Intermittent on/off illumination reveals that cells achieve maximum neurite outgrowth if the off-time duration per cycle is shorter than 45 min. Overall, light-mediated kinetic control enables precise dissection of the temporal dimension within the intracellular signal transduction network. PMID:24667437

PaASK1, a mitogen-activated protein kinase kinase kinase that controls cell degeneration and cell differentiation in Podospora anserina.

PubMed Central

Kicka, Sébastien; Silar, Philippe

2004-01-01

MAPKKK are kinases involved in cell signaling. In fungi, these kinases are known to regulate development, pathogenicity, and the sensing of external conditions. We show here that Podospora anserina strains mutated in PaASK1, a MAPKKK of the MEK family, are impaired in the development of crippled growth, a cell degeneration process caused by C, a nonconventional infectious element. They also display defects in mycelium pigmentation, differentiation of aerial hyphae, and making of fruiting bodies, three hallmarks of cell differentiation during stationary phase in P. anserina. Overexpression of PaASK1 results in exacerbation of crippled growth. PaASK1 is a large protein of 1832 amino acids with several domains, including a region rich in proline and a 60-amino-acid-long polyglutamine stretch. Deletion analysis reveals that the polyglutamine stretch is dispensable for PaASK1 activity, whereas the region that contains the prolines is essential but insufficient to promote full activity. We discuss a model based on the hysteresis of a signal transduction cascade to account for the role of PaASK1 in both cell degeneration and stationary-phase cell differentiation. PMID:15082544