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Sample records for pathway components involved

  1. Biosynthesis of Unusual Moth Pheromone Components Involves Two Different Pathways in the Navel Orangeworm, Amyelois transitella

    PubMed Central

    Wang, Hong-Lei; Zhao, Cheng-Hua; Millar, Jocelyn G.; Cardé, Ring T.

    2010-01-01

    The sex pheromone of the navel orangeworm, Amyelois transitella (Walker) (Lepidoptera: Pyralidae), consists of two different types of components, one type including (11Z,13Z)-11,13-hexadecadienal (11Z,13Z-16:Ald) with a terminal functional group containing oxygen, similar to the majority of moth pheromones reported, and another type including the unusual long-chain pentaenes, (3Z,6Z,9Z,12Z,15Z)-3,6,9,12,15-tricosapentaene (3Z,6Z,9Z,12Z,15Z-23:H) and (3Z,6Z,9Z,12Z,15Z)- 3,6,9,12,15-pentacosapentaene (3Z,6Z,9Z,12Z,15Z-25:H). After decapitation of females, the titer of 11Z,13Z-16:Ald in the pheromone gland decreased significantly, whereas the titer of the pentaenes remained unchanged. Injection of a pheromone biosynthesis activating peptide (PBAN) into the abdomens of decapitated females restored the titer of 11Z,13Z-16:Ald and even increased it above that in intact females, whereas the titer of the pentaenes in the pheromone gland was not affected by PBAN injection. In addition to common fatty acids, two likely precursors of 11Z,13Z-16:Ald, i.e., (Z)-11-hexadecenoic and (11Z,13Z)-11,13-hexadecadienoic acid, as well as traces of (Z)-6-hexadecenoic acid, were found in gland extracts. In addition, pheromone gland lipids contained (5Z,8Z,11Z,14Z,17Z)-5,8,11,14,17-icosapentaenoic acid, which also was found in extracts of the rest of the abdomen. Deuterium-labeled fatty acids, (16,16,16-D3)-hexadecanoic acid and (Z)-[13,13,14,14,15,15,16,16,16-D9]-11-hexadecenoic acid, were incorporated into 11Z,13Z-16:Ald after topical application to the sex pheromone gland coupled with abdominal injection of PBAN. Deuterium label was incorporated into the C23 and C25 pentaenes after injection of (9Z,12Z,15Z)- [17,17,18,18,18-D5]-9,12,15-octadecatrienoic acid into 1–2 d old female pupae. These labeling results, in conjunction with the composition of fatty acid intermediates found in pheromone gland extracts, support different pathways leading to the two pheromone components. 11Z,13Z-16

  2. Vestibular pathways involved in cognition

    PubMed Central

    Hitier, Martin; Besnard, Stephane; Smith, Paul F.

    2014-01-01

    Recent discoveries have emphasized the role of the vestibular system in cognitive processes such as memory, spatial navigation and bodily self-consciousness. A precise understanding of the vestibular pathways involved is essential to understand the consequences of vestibular diseases for cognition, as well as develop therapeutic strategies to facilitate recovery. The knowledge of the “vestibular cortical projection areas”, defined as the cortical areas activated by vestibular stimulation, has dramatically increased over the last several years from both anatomical and functional points of view. Four major pathways have been hypothesized to transmit vestibular information to the vestibular cortex: (1) the vestibulo-thalamo-cortical pathway, which probably transmits spatial information about the environment via the parietal, entorhinal and perirhinal cortices to the hippocampus and is associated with spatial representation and self-versus object motion distinctions; (2) the pathway from the dorsal tegmental nucleus via the lateral mammillary nucleus, the anterodorsal nucleus of the thalamus to the entorhinal cortex, which transmits information for estimations of head direction; (3) the pathway via the nucleus reticularis pontis oralis, the supramammillary nucleus and the medial septum to the hippocampus, which transmits information supporting hippocampal theta rhythm and memory; and (4) a possible pathway via the cerebellum, and the ventral lateral nucleus of the thalamus (perhaps to the parietal cortex), which transmits information for spatial learning. Finally a new pathway is hypothesized via the basal ganglia, potentially involved in spatial learning and spatial memory. From these pathways, progressively emerges the anatomical network of vestibular cognition. PMID:25100954

  3. Activation of eNOS in endothelial cells exposed to ionizing radiation involves components of the DNA damage response pathway

    SciTech Connect

    Nagane, Masaki; Yasui, Hironobu; Sakai, Yuri; Yamamori, Tohru; Niwa, Koichi; Hattori, Yuichi; Kondo, Takashi; Inanami, Osamu

    2015-01-02

    Highlights: • eNOS activity is increased in BAECs exposed to X-rays. • ATM is involved in this increased eNOS activity. • HSP90 modulates the radiation-induced activation of ATM and eNOS. - Abstract: In this study, the involvement of ataxia telangiectasia mutated (ATM) kinase and heat shock protein 90 (HSP90) in endothelial nitric oxide synthase (eNOS) activation was investigated in X-irradiated bovine aortic endothelial cells. The activity of nitric oxide synthase (NOS) and the phosphorylation of serine 1179 of eNOS (eNOS-Ser1179) were significantly increased in irradiated cells. The radiation-induced increases in NOS activity and eNOS-Ser1179 phosphorylation levels were significantly reduced by treatment with either an ATM inhibitor (Ku-60019) or an HSP90 inhibitor (geldanamycin). Geldanamycin was furthermore found to suppress the radiation-induced phosphorylation of ATM-Ser1181. Our results indicate that the radiation-induced eNOS activation in bovine aortic endothelial cells is regulated by ATM and HSP90.

  4. Proteomic mapping of stimulus-specific signaling pathways involved in THP-1 cells exposed to Porphyromonas gingivalis or its purified components.

    PubMed

    Saba, Julian A; McComb, Mark E; Potts, Donna L; Costello, Catherine E; Amar, Salomon

    2007-06-01

    Periodontitis is an inflammatory disease initiated by host-parasite interactions which contributes to connective tissue destruction and alveolar bone resorption. Porphyromonas gingivalis (P.g.), a black-pigmented Gram-negative anaerobic bacterium, is a major pathogen in the development and progression of periodontitis. To characterize the role that P. gingivalis and its cell surface components play in disease processes, we investigated the differential expression of proteins induced by live P.g., P.g. LPS, and P.g. FimA, using two-dimensional gel electrophoresis in combination with mass spectrometry. We have tested whether, at the level of protein expression, unique signaling pathways are differentially induced by the bacterial components P.g. LPS and P.g. FimA, as compared to live P.g. We found that P.g. LPS stimulation of THP-1 up-regulated the expression of a set of proteins compared to control: deoxyribonuclease, actin, carbonic anhydrase 2, alpha enolase, adenylyl cyclase-associated protein (CAP1), protein disulfide isomerase (PDI), glucose regulated protein (grp78), and 70-kDa heat shock protein (HSP70), whereas FimA treatment did not result in statistically significant changes to protein levels versus the control. Live P.g. stimulation resulted in 12 differentially expressed proteins: CAP1, tubulin beta-2 chain, ATP synthase beta chain, tubulin alpha-6 chain, PDI, vimentin, 60-kDa heat shock protein, and nucleolin were found to be up-regulated, while carbonic anhydrase II, beta-actin, and HSP70 were down-regulated relative to control. These differential changes by the bacteria and its components are interpreted as preferential signal pathway activation in host immune/inflammatory responses to P.g. infection. PMID:17477557

  5. Immunoregulatory Pathways Involved in Inflammatory Bowel Disease.

    PubMed

    Fonseca-Camarillo, Gabriela; Yamamoto-Furusho, Jesús K

    2015-09-01

    Inflammatory bowel diseases (IBD) include ulcerative colitis and Crohn's disease. The immune response in ulcerative colitis is different from the Crohn's disease. Accumulating evidence suggests that IBD results from an inappropriate inflammatory response to intestinal microbes in a genetically susceptible host. Several immunoregulatory abnormalities have been reported in patients with IBD, including the ratio of proinflammatory (tumor necrosis factor alpha, IL-6, IL-1-β) to immunoregulatory cytokines (IL-10, TGF-β, IL-35) and selective activation of T-helper (Th) lymphocyte subsets (Th1, Th2, Th9, Th17, and regulatory T cells). The purpose of this review is to show the immunoregulatory pathways (regulatory cells and cytokines) involved in IBD published in recent years. PMID:26111210

  6. Studying lipids involved in the endosomal pathway.

    PubMed

    Bissig, Christin; Johnson, Shem; Gruenberg, Jean

    2012-01-01

    Endosomes along the degradation pathway exhibit a multivesicular appearance and differ in their lipid compositions. Association of proteins to specific membrane lipids and presumably also lipid-lipid interactions contribute to the formation of functional membrane platforms that regulate endosome biogenesis and function. This chapter provides a brief review of the functions of endosomal lipids in the degradation pathway, a discussion of techniques that allow studying lipid-based mechanisms and a selection of step-by-step protocols for in vivo and in vitro methods commonly used to study lipid roles in endocytosis. The techniques described here have been used to elucidate the function of the late endosomal lipid lysobisphosphatidic acid and allow the monitoring of lipid distribution, levels and dynamics, as well as the characterization of lipid-binding partners. PMID:22325596

  7. Temporal and evolutionary dynamics of two-component signaling pathways.

    PubMed

    Salazar, Michael E; Laub, Michael T

    2015-04-01

    Bacteria sense and respond to numerous environmental signals through two-component signaling pathways. Typically, a given stimulus will activate a sensor histidine kinase to autophosphorylate and then phosphotransfer to a cognate response regulator, which can mount an appropriate response. Although these signaling pathways often appear to be simple switches, they can also orchestrate surprisingly sophisticated and complex responses. These temporal dynamics arise from several key regulatory features, including the bifunctionality of histidine kinases as well as positive and negative feedback loops. Two-component signaling pathways are also dynamic on evolutionary time-scales, expanding dramatically in many species through gene duplication and divergence. Here, we review recent work probing the temporal and evolutionary dynamics of two-component signaling systems. PMID:25589045

  8. Temporal and Evolutionary Dynamics of Two-Component Signaling Pathways

    PubMed Central

    Salazar, Michael E.; Laub, Michael T.

    2015-01-01

    Bacteria sense and respond to numerous environmental signals through two-component signaling pathways. Typically, a given stimulus will activate a sensor histidine kinase to autophosphorylate and then phosphotransfer to a cognate response regulator, which can mount an appropriate response. Although these signaling pathways often appear to be simple switches, they can also orchestrate surprisingly sophisticated and complex responses. These temporal dynamics arise from several key regulatory features, including the bifunctionality of histidine kinases as well as positive and negative feedback loops. Two-component signaling pathways are also dynamic on evolutionary time-scales, expanding dramatically in many species through gene duplication and divergence. Here, we review recent work probing the temporal and evolutionary dynamics of two-component signaling systems. PMID:25589045

  9. Evolutionary conservation of plant gibberellin signalling pathway components

    PubMed Central

    Vandenbussche, Filip; Fierro, Ana C; Wiedemann, Gertrud; Reski, Ralf; Van Der Straeten, Dominique

    2007-01-01

    Background: Gibberellins (GA) are plant hormones that can regulate germination, elongation growth, and sex determination. They ubiquitously occur in seed plants. The discovery of gibberellin receptors, together with advances in understanding the function of key components of GA signalling in Arabidopsis and rice, reveal a fairly short GA signal transduction route. The pathway essentially consists of GID1 gibberellin receptors that interact with F-box proteins, which in turn regulate degradation of downstream DELLA proteins, suppressors of GA-controlled responses. Results: Arabidopsis sequences of the gibberellin signalling compounds were used to screen databases from a variety of plants, including protists, for homologues, providing indications for the degree of conservation of the pathway. The pathway as such appears completely absent in protists, the moss Physcomitrella patens shares only a limited homology with the Arabidopsis proteins, thus lacking essential characteristics of the classical GA signalling pathway, while the lycophyte Selaginella moellendorffii contains a possible ortholog for each component. The occurrence of classical GA responses can as yet not be linked with the presence of homologues of the signalling pathway. Alignments and display in neighbour joining trees of the GA signalling components confirm the close relationship of gymnosperms, monocotyledonous and dicotyledonous plants, as suggested from previous studies. Conclusion: Homologues of the GA-signalling pathway were mainly found in vascular plants. The GA signalling system may have its evolutionary molecular onset in Physcomitrella patens, where GAs at higher concentrations affect gravitropism and elongation growth. PMID:18047669

  10. The coupling of pathways and processes through shared components

    PubMed Central

    2011-01-01

    Background The coupling of pathways and processes through shared components is being increasingly recognised as a common theme which occurs in many cell signalling contexts, in which it plays highly non-trivial roles. Results In this paper we develop a basic modelling and systems framework in a general setting for understanding the coupling of processes and pathways through shared components. Our modelling framework starts with the interaction of two components with a common third component and includes production and degradation of all these components. We analyze the signal processing in our model to elucidate different aspects of the coupling. We show how different kinds of responses, including "ultrasensitive" and adaptive responses, may occur in this setting. We then build on the basic model structure and examine the effects of additional control regulation, switch-like signal processing, and spatial signalling. In the process, we identify a way in which allosteric regulation may contribute to signalling specificity, and how competitive effects may allow an enzyme to robustly coordinate and time the activation of parallel pathways. Conclusions We have developed and analyzed a common systems platform for examining the effects of coupling of processes through shared components. This can be the basis for subsequent expansion and understanding the many biologically observed variations on this common theme. PMID:21714894

  11. Dissecting Abscisic Acid Signaling Pathways Involved in Cuticle Formation.

    PubMed

    Cui, Fuqiang; Brosché, Mikael; Lehtonen, Mikko T; Amiryousefi, Ali; Xu, Enjun; Punkkinen, Matleena; Valkonen, Jari P T; Fujii, Hiroaki; Overmyer, Kirk

    2016-06-01

    The cuticle is the outer physical barrier of aerial plant surfaces and an important interaction point between plants and the environment. Many environmental stresses affect cuticle formation, yet the regulatory pathways involved remain undefined. We used a genetics and gene expression analysis in Arabidopsis thaliana to define an abscisic acid (ABA) signaling loop that positively regulates cuticle formation via the core ABA signaling pathway, including the PYR/PYL receptors, PP2C phosphatase, and SNF1-Related Protein Kinase (SnRK) 2.2/SnRK2.3/SnRK2.6. Downstream of the SnRK2 kinases, cuticle formation was not regulated by the ABA-responsive element-binding transcription factors but rather by DEWAX, MYB16, MYB94, and MYB96. Additionally, low air humidity increased cuticle formation independent of the core ABA pathway and cell death/reactive oxygen species signaling attenuated expression of cuticle-biosynthesis genes. In Physcomitrella patens, exogenous ABA suppressed expression of cuticle-related genes, whose Arabidopsis orthologs were ABA-induced. Hence, the mechanisms regulating cuticle formation are conserved but sophisticated in land plants. Signaling specifically related to cuticle deficiency was identified to play a major role in the adaptation of ABA signaling pathway mutants to increased humidity and in modulating their immunity to Botrytis cinerea in Arabidopsis. These results define a cuticle-specific downstream branch in the ABA signaling pathway that regulates responses to the external environment. PMID:27060495

  12. Engineering key components in a synthetic eukaryotic signal transduction pathway

    PubMed Central

    Antunes, Mauricio S; Morey, Kevin J; Tewari-Singh, Neera; Bowen, Tessa A; Smith, J Jeff; Webb, Colleen T; Hellinga, Homme W; Medford, June I

    2009-01-01

    Signal transduction underlies how living organisms detect and respond to stimuli. A goal of synthetic biology is to rewire natural signal transduction systems. Bacteria, yeast, and plants sense environmental aspects through conserved histidine kinase (HK) signal transduction systems. HK protein components are typically comprised of multiple, relatively modular, and conserved domains. Phosphate transfer between these components may exhibit considerable cross talk between the otherwise apparently linear pathways, thereby establishing networks that integrate multiple signals. We show that sequence conservation and cross talk can extend across kingdoms and can be exploited to produce a synthetic plant signal transduction system. In response to HK cross talk, heterologously expressed bacterial response regulators, PhoB and OmpR, translocate to the nucleus on HK activation. Using this discovery, combined with modification of PhoB (PhoB-VP64), we produced a key component of a eukaryotic synthetic signal transduction pathway. In response to exogenous cytokinin, PhoB-VP64 translocates to the nucleus, binds a synthetic PlantPho promoter, and activates gene expression. These results show that conserved-signaling components can be used across kingdoms and adapted to produce synthetic eukaryotic signal transduction pathways. PMID:19455134

  13. Characterisation of ethylene pathway components in non-climacteric capsicum

    PubMed Central

    2013-01-01

    Background Climacteric fruit exhibit high ethylene and respiration levels during ripening but these levels are limited in non-climacteric fruit. Even though capsicum is in the same family as the well-characterised climacteric tomato (Solanaceae), it is non-climacteric and does not ripen normally in response to ethylene or if harvested when mature green. However, ripening progresses normally in capsicum fruit when they are harvested during or after what is called the ‘Breaker stage’. Whether ethylene, and components of the ethylene pathway such as 1-aminocyclopropane 1-carboxylate (ACC) oxidase (ACO), ACC synthase (ACS) and the ethylene receptor (ETR), contribute to non-climacteric ripening in capsicum has not been studied in detail. To elucidate the behaviour of ethylene pathway components in capsicum during ripening, further analysis is therefore needed. The effects of ethylene or inhibitors of ethylene perception, such as 1-methylcyclopropene, on capsicum fruit ripening and the ethylene pathway components may also shed some light on the role of ethylene in non-climacteric ripening. Results The expression of several isoforms of ACO, ACS and ETR were limited during capsicum ripening except one ACO isoform (CaACO4). ACS activity and ACC content were also low in capsicum despite the increase in ACO activity during the onset of ripening. Ethylene did not stimulate capsicum ripening but 1-methylcyclopropene treatment delayed the ripening of Breaker-harvested fruit. Some of the ACO, ACS and ETR isoforms were also differentially expressed upon treatment with ethylene or 1-methylcyclopropene. Conclusions ACS activity may be the rate limiting step in the ethylene pathway of capsicum which restricts ACC content. The differential expression of several ethylene pathway components during ripening and upon ethylene or 1-methylclopropene treatment suggests that the ethylene pathway may be regulated differently in non-climacteric capsicum compared to the climacteric tomato

  14. Kavain Involvement in LPS-Induced Signaling Pathways.

    PubMed

    Tang, Xiaoren; Amar, Salomon

    2016-10-01

    Kavain, a compound extracted from the Kava plant, Piper methysticum, is found to be involved in TNF-α expression in human and mouse cells via regulation of transcriptional factors such as NF-kB and LITAF. LITAF is known to activate the transcription of more than 20 cytokines that are involved in a variety of cellular processes and is associated with many inflammatory diseases, including angiogenesis, cancer, arthritis, and more. The modulation of LITAF is expected to positively affect cytokine-mediated diseases. Thus, intensive efforts have been deployed in search of LITAF inhibitors. In this work, we found that, in vitro, Kavain reduced LPS- induced TNF-α secretion in mouse macrophages, mouse bone marrow macrophages (BMM), and human peripheral blood mononuclear cells (HPBMC). We also found that Kavain treatment in RAW264.7 cells deactivated MyD88 and Akt, inhibited LITAF, and reduced the production of TNF-α, IL-27, and MIG in response to LPS. Similarly, it had a significant in vivo anti-inflammatory effect on wild-type (WT) mice that developed Collagen Antibody Induced Arthritis (CAIA). Overall, MyD88 was found to be an important mediator of the LPS-induced inflammatory response that can be distinguished from the NF-κB pathway. We also found that MyD88 is involved in the pathway linking LPS/LITAF to TNF-α. Therefore, given that Kavain modulates LPS-induced signaling pathways leading to cytokine expression, therapeutic interventions involving Kavain in inflammatory diseases are warranted. J. Cell. Biochem. 117: 2272-2280, 2016. © 2016 Wiley Periodicals, Inc. PMID:26917453

  15. Signal transduction pathways involved in mechanotransduction in bone cells

    SciTech Connect

    Liedert, Astrid . E-mail: astrid.liedert@uni-ulm.de; Kaspar, Daniela; Blakytny, Robert; Claes, Lutz; Ignatius, Anita

    2006-10-13

    Several in vivo and in vitro studies with different loading regimens showed that mechanical stimuli have an influence on proliferation and differentiation of bone cells. Prerequisite for this influence is the transduction of mechanical signals into the cell, a phenomenon that is termed mechanotransduction, which is essential for the maintenance of skeletal homeostasis in adults. Mechanoreceptors, such as the integrins, cadherins, and stretch-activated Ca{sup 2+} channels, together with various signal transduction pathways, are involved in the mechanotransduction process that ultimately regulates gene expression in the nucleus. Mechanotransduction itself is considered to be regulated by hormones, the extracellular matrix of the osteoblastic cells and the mode of the mechanical stimulus.

  16. Signaling Pathways Involved in Lunar Dust Induced Cytotoxicity

    NASA Technical Reports Server (NTRS)

    Zhang, Ye; Lam, Chiu-Wing; Scully, Robert R.; Williams, Kyle; Zalesak, Selina; Wu, Honglu; James, John T.

    2014-01-01

    The Moon's surface is covered by a layer of fine, reactive dust. Lunar dust contain about 1-2% of very fine dust (< 3 micron), that is respirable. The habitable area of any lunar landing vehicle and outpost would inevitably be contaminated with lunar dust that could pose a health risk. The purpose of the study is to evaluate the toxicity of Apollo moon dust in rodents to assess the health risk of dust exposures to humans. One of the particular interests in the study is to evaluate dust-induced changes of the expression of fibrosis-related genes, and to identify specific signaling pathways involved in lunar dust-induced toxicity. F344 rats were exposed for 4 weeks (6h/d; 5d/wk) in nose-only inhalation chambers to concentrations of 0 (control air), 2.1, 6.1, 21, and 61 mg/m(exp 3) of lunar dust. Five rats per group were euthanized 1 day, 1 week, 1 month, and 3 months after the last inhalation exposure. The total RNAs were isolated from the blood or lung tissue after being lavaged, using the Qigen RNeasy kit. The Rat Fibrosis RT2 Profile PCR Array was used to profile the expression of 84 genes relevant to fibrosis. The genes with significant expression changes are identified and the gene expression data were further analyzed using IPA pathway analysis tool to determine the signaling pathways with significant changes.

  17. Analysis of PI3K pathway components in human cancers

    PubMed Central

    DARAGMEH, JAMILA; BARRIAH, WASEIM; SAAD, BASHAR; ZAID, HILAL

    2016-01-01

    Recent advances in genomics, proteomics, cell biology and biochemistry of tumors have revealed new pathways that are aberrantly activated in numerous cancer types. However, the enormous amount of data available in this field may mislead scientists in focused research. As cancer cell growth and progression is often dependent upon the phosphoinositide 3-kinase (PI3K)/AKT pathway, there has been extensive research into the proteins implicated in the PI3K pathway. Using data available in the Human Protein Atlas database, the current study investigated the expression of 25 key proteins that are known to be involved with PI3K pathway activation in a distinct group of 20 cancer types. These proteins are AKTIP, ARP1, BAD, GSK3A, GSK3B, MERTK-1, PIK3CA, PRR5, PSTPIP2, PTEN, FOX1, RHEB, RPS6KB1, TSC1, TP53, BCL2, CCND1, WFIKKN2, CREBBP, caspase-9, PTK2, EGFR, FAS, CDKN1A and XIAP. The analysis revealed pronounced expression of specific proteins in distinct cancer tissues, which may have the potential to serve as targets for treatments and provide insights into the molecular basis of cancer. PMID:27073576

  18. Anatomical Pathways Involved in Generating and Sensing Rhythmic Whisker Movements

    PubMed Central

    Bosman, Laurens W. J.; Houweling, Arthur R.; Owens, Cullen B.; Tanke, Nouk; Shevchouk, Olesya T.; Rahmati, Negah; Teunissen, Wouter H. T.; Ju, Chiheng; Gong, Wei; Koekkoek, Sebastiaan K. E.; De Zeeuw, Chris I.

    2011-01-01

    The rodent whisker system is widely used as a model system for investigating sensorimotor integration, neural mechanisms of complex cognitive tasks, neural development, and robotics. The whisker pathways to the barrel cortex have received considerable attention. However, many subcortical structures are paramount to the whisker system. They contribute to important processes, like filtering out salient features, integration with other senses, and adaptation of the whisker system to the general behavioral state of the animal. We present here an overview of the brain regions and their connections involved in the whisker system. We do not only describe the anatomy and functional roles of the cerebral cortex, but also those of subcortical structures like the striatum, superior colliculus, cerebellum, pontomedullary reticular formation, zona incerta, and anterior pretectal nucleus as well as those of level setting systems like the cholinergic, histaminergic, serotonergic, and noradrenergic pathways. We conclude by discussing how these brain regions may affect each other and how they together may control the precise timing of whisker movements and coordinate whisker perception. PMID:22065951

  19. Cerulenin-mediated apoptosis is involved in adenine metabolic pathway

    SciTech Connect

    Chung, Kyung-Sook; Sun, Nam-Kyu; Lee, Seung-Hee; Lee, Hyun-Jee; Choi, Shin-Jung; Kim, Sun-Kyung; Song, Ju-Hyun; Jang, Young-Joo; Song, Kyung-Bin; Yoo, Hyang-Sook; Simon, Julian . E-mail: jsimon@fhcrc.org; Won, Misun . E-mail: misun@kribb.re.kr

    2006-10-27

    Cerulenin, a fatty acid synthase (FAS) inhibitor, induces apoptosis of variety of tumor cells. To elucidate mode of action by cerulenin, we employed the proteomics approach using Schizosaccharomyces pombe. The differential protein expression profile of S. pombe revealed that cerulenin modulated the expressions of proteins involved in stresses and metabolism, including both ade10 and adk1 proteins. The nutrient supplementation assay demonstrated that cerulenin affected enzymatic steps transferring a phosphoribosyl group. This result suggests that cerulenin accumulates AMP and p-ribosyl-s-amino-imidazole carboxamide (AICAR) and reduces other necessary nucleotides, which induces feedback inhibition of enzymes and the transcriptional regulation of related genes in de novo and salvage adenine metabolic pathway. Furthermore, the deregulation of adenine nucleotide synthesis may interfere ribonucleotide reductase and cause defects in cell cycle progression and chromosome segregation. In conclusion, cerulenin induces apoptosis through deregulation of adenine nucleotide biosynthesis resulting in nuclear division defects in S. pombe.

  20. Ter94/VCP Is a Novel Component Involved in BMP Signaling

    PubMed Central

    Zeng, Zhao; de Gorter, David J. J.; Kowalski, Maria; ten Dijke, Peter; Shimmi, Osamu

    2014-01-01

    Bone morphogenetic proteins (BMPs), a subgroup of the transforming growth factor (TGF)-β family, transduce their signal through multiple components downstream of their receptors. Even though the components involved in the BMP signaling pathway have been intensely studied, many molecules mediating BMP signaling remain to be addressed. To identify novel components that participate in BMP signaling, RNA interference (RNAi)-based screening was established by detecting phosphorylated Mad (pMad) in Drosophila S2 cells. Ter94, a member of the family of AAA ATPases, was identified as a novel mediator of BMP signaling, which is required for the phosphorylation of Mad in Drosophila S2 cells. Moreover, the mammalian orthlog of Ter94 valosin-containing protein (VCP) plays a critical role in the BMP-Smad1/5/8 signaling pathway in mammalian cells. Genetic evidence suggests that Ter94 is involved in the dorsal-ventral patterning of the Drosophila early embryo through regulating decapentaplegic (Dpp)/BMP signals. Taken together, our data suggest that Ter94/VCP appears to be an evolutionarily conserved component that regulates BMP-Smad1/5/8 signaling. PMID:25469707

  1. Solution structure of ligands involved in purine salvage pathway.

    PubMed

    Karnawat, Vishakha; Puranik, Mrinalini

    2015-12-01

    Analogues of intermediates involved in the purine salvage pathway can be exploited as potential drug molecules against enzymes of protozoan parasites. To develop such analogues we need knowledge of the solution structures, predominant tautomer at physiological pH and protonation-state of the corresponding natural ligand. In this regard, we have employed ultraviolet resonance Raman spectroscopy (UVRR) in combination with density functional theory (DFT) to study the solution structures of two relatively unexplored intermediates, 6-phosphoryl IMP (6-pIMP) and succinyl adenosine-5'-monophosphate (sAMP), of purine salvage pathway. These molecules are intermediates in a two step enzymatic process that converts inosine-5'-monpophosphate (IMP) to adenosine-5'-monophosphate (AMP). Experimental data on the molecular structure of these ligands is lacking. We report UVRR spectra of these two ligands, obtained at an excitation wavelength of 260 nm. Using isotope induced shifts and DFT calculations we assigned observed spectra to computed normal modes. We find that sAMP exists as neutral species at physiological pH and the predominant tautomer in solution bears proton at N10 position of purine ring. Though transient in solution, 6-pIMP is captured in the enzyme-bound form. This work provides the structural information of these ligands in solution state at physiological pH. We further compare these structures with the structures of AMP and IMP. Despite the presence of similar purine rings in AMP and sAMP, their UVRR spectra are found to be very different. Similarly, though the purine ring in 6-pIMP resembles that of IMP, UVRR spectra of the two molecules are distinct. These differences in the vibrational spectra provide direct information on the effects of exocyclic groups on the skeletal structures of these molecules. Our results identify key bands in the vibrational spectra of these ligands which may serve as markers of hydrogen bonding interactions upon binding to the active

  2. Four Pathways Involving Innate Immunity in the Pathogenesis of Preeclampsia

    PubMed Central

    Bounds, Kelsey R.; Newell-Rogers, M. Karen; Mitchell, Brett M.

    2015-01-01

    The maternal innate immune system plays an important role both in normal pregnancy as well as hypertensive disorders of pregnancy including preeclampsia (PE). We propose four pathways that involve excessive innate immunity that lead to most forms of PE. Pre-existing endothelial dysfunction plus pregnancy leads to an excessive innate immune response resulting in widespread inflammation, placental and renal dysfunction, vasoconstriction, and PE. Placental dysfunction due to shallow trophoblast invasion, inadequate spiral artery remodeling, and/or low placental perfusion initiates an innate immune response leading to excessive inflammation, endothelial and renal dysfunction, and PE. A heightened innate immune system due to pre-existing or acquired infections plus the presence of a paternally derived placenta and semi-allogeneic fetus cause an excessive innate immune response which manifests as PE. Lastly, an abnormal and excessive maternal immune response to pregnancy leads to widespread inflammation, organ dysfunction, and PE. We discuss the potential role of innate immunity in each of these scenarios, as well as the overlap, and how targeting the innate immune system might lead to therapies for the treatment of PE. PMID:26664892

  3. Involvement of the Kynurenine Pathway in Human Glioma Pathophysiology

    PubMed Central

    Adams, Seray; Teo, Charles; McDonald, Kerrie L.; Zinger, Anna; Bustamante, Sonia; Lim, Chai K.; Sundaram, Gayathri; Braidy, Nady; Brew, Bruce J.; Guillemin, Gilles J.

    2014-01-01

    The kynurenine pathway (KP) is the principal route of L-tryptophan (TRP) catabolism leading to the production of kynurenine (KYN), the neuroprotectants, kynurenic acid (KYNA) and picolinic acid (PIC), the excitotoxin, quinolinic acid (QUIN) and the essential pyridine nucleotide, nicotinamide adenine dinucleotide (NAD+). The enzymes indoleamine 2,3-dioxygenase-1 (IDO-1), indoleamine 2,3-dioxygenase-2 (IDO-2) and tryptophan 2,3-dioxygenase (TDO-2) initiate the first step of the KP. IDO-1 and TDO-2 induction in tumors are crucial mechanisms implicated to play pivotal roles in suppressing anti-tumor immunity. Here, we report the first comprehensive characterisation of the KP in 1) cultured human glioma cells and 2) plasma from patients with glioblastoma (GBM). Our data revealed that interferon-gamma (IFN-γ) stimulation significantly potentiated the expression of the KP enzymes, IDO-1 IDO-2, kynureninase (KYNU), kynurenine hydroxylase (KMO) and significantly down-regulated 2-amino-3-carboxymuconate semialdehyde decarboxylase (ACMSD) and kynurenine aminotransferase-I (KAT-I) expression in cultured human glioma cells. This significantly increased KP activity but significantly lowered the KYNA/KYN neuroprotective ratio in human cultured glioma cells. KP activation (KYN/TRP) was significantly higher, whereas the concentrations of the neuroreactive KP metabolites TRP, KYNA, QUIN and PIC and the KYNA/KYN ratio were significantly lower in GBM patient plasma (n = 18) compared to controls. These results provide further evidence for the involvement of the KP in glioma pathophysiology and highlight a potential role of KP products as novel and highly attractive therapeutic targets to evaluate for the treatment of brain tumors, aimed at restoring anti-tumor immunity and reducing the capacity for malignant cells to produce NAD+, which is necessary for energy production and DNA repair. PMID:25415278

  4. Involvement of the kynurenine pathway in human glioma pathophysiology.

    PubMed

    Adams, Seray; Teo, Charles; McDonald, Kerrie L; Zinger, Anna; Bustamante, Sonia; Lim, Chai K; Sundaram, Gayathri; Braidy, Nady; Brew, Bruce J; Guillemin, Gilles J

    2014-01-01

    The kynurenine pathway (KP) is the principal route of L-tryptophan (TRP) catabolism leading to the production of kynurenine (KYN), the neuroprotectants, kynurenic acid (KYNA) and picolinic acid (PIC), the excitotoxin, quinolinic acid (QUIN) and the essential pyridine nucleotide, nicotinamide adenine dinucleotide (NAD(+)). The enzymes indoleamine 2,3-dioxygenase-1 (IDO-1), indoleamine 2,3-dioxygenase-2 (IDO-2) and tryptophan 2,3-dioxygenase (TDO-2) initiate the first step of the KP. IDO-1 and TDO-2 induction in tumors are crucial mechanisms implicated to play pivotal roles in suppressing anti-tumor immunity. Here, we report the first comprehensive characterisation of the KP in 1) cultured human glioma cells and 2) plasma from patients with glioblastoma (GBM). Our data revealed that interferon-gamma (IFN-γ) stimulation significantly potentiated the expression of the KP enzymes, IDO-1 IDO-2, kynureninase (KYNU), kynurenine hydroxylase (KMO) and significantly down-regulated 2-amino-3-carboxymuconate semialdehyde decarboxylase (ACMSD) and kynurenine aminotransferase-I (KAT-I) expression in cultured human glioma cells. This significantly increased KP activity but significantly lowered the KYNA/KYN neuroprotective ratio in human cultured glioma cells. KP activation (KYN/TRP) was significantly higher, whereas the concentrations of the neuroreactive KP metabolites TRP, KYNA, QUIN and PIC and the KYNA/KYN ratio were significantly lower in GBM patient plasma (n = 18) compared to controls. These results provide further evidence for the involvement of the KP in glioma pathophysiology and highlight a potential role of KP products as novel and highly attractive therapeutic targets to evaluate for the treatment of brain tumors, aimed at restoring anti-tumor immunity and reducing the capacity for malignant cells to produce NAD(+), which is necessary for energy production and DNA repair. PMID:25415278

  5. Pathway Network Analyses for Autism Reveal Multisystem Involvement, Major Overlaps with Other Diseases and Convergence upon MAPK and Calcium Signaling

    PubMed Central

    Wen, Ya; Alshikho, Mohamad J.; Herbert, Martha R.

    2016-01-01

    We used established databases in standard ways to systematically characterize gene ontologies, pathways and functional linkages in the large set of genes now associated with autism spectrum disorders (ASDs). These conditions are particularly challenging—they lack clear pathognomonic biological markers, they involve great heterogeneity across multiple levels (genes, systemic biological and brain characteristics, and nuances of behavioral manifestations)—and yet everyone with this diagnosis meets the same defining behavioral criteria. Using the human gene list from Simons Foundation Autism Research Initiative (SFARI) we performed gene set enrichment analysis with the Kyoto Encyclopedia of Genes and Genomes (KEGG) Pathway Database, and then derived a pathway network from pathway-pathway functional interactions again in reference to KEGG. Through identifying the GO (Gene Ontology) groups in which SFARI genes were enriched, mapping the coherence between pathways and GO groups, and ranking the relative strengths of representation of pathway network components, we 1) identified 10 disease-associated and 30 function-associated pathways 2) revealed calcium signaling pathway and neuroactive ligand-receptor interaction as the most enriched, statistically significant pathways from the enrichment analysis, 3) showed calcium signaling pathways and MAPK signaling pathway to be interactive hubs with other pathways and also to be involved with pervasively present biological processes, 4) found convergent indications that the process “calcium-PRC (protein kinase C)-Ras-Raf-MAPK/ERK” is likely a major contributor to ASD pathophysiology, and 5) noted that perturbations associated with KEGG’s category of environmental information processing were common. These findings support the idea that ASD-associated genes may contribute not only to core features of ASD themselves but also to vulnerability to other chronic and systemic problems potentially including cancer, metabolic

  6. Pathway Network Analyses for Autism Reveal Multisystem Involvement, Major Overlaps with Other Diseases and Convergence upon MAPK and Calcium Signaling.

    PubMed

    Wen, Ya; Alshikho, Mohamad J; Herbert, Martha R

    2016-01-01

    We used established databases in standard ways to systematically characterize gene ontologies, pathways and functional linkages in the large set of genes now associated with autism spectrum disorders (ASDs). These conditions are particularly challenging--they lack clear pathognomonic biological markers, they involve great heterogeneity across multiple levels (genes, systemic biological and brain characteristics, and nuances of behavioral manifestations)-and yet everyone with this diagnosis meets the same defining behavioral criteria. Using the human gene list from Simons Foundation Autism Research Initiative (SFARI) we performed gene set enrichment analysis with the Kyoto Encyclopedia of Genes and Genomes (KEGG) Pathway Database, and then derived a pathway network from pathway-pathway functional interactions again in reference to KEGG. Through identifying the GO (Gene Ontology) groups in which SFARI genes were enriched, mapping the coherence between pathways and GO groups, and ranking the relative strengths of representation of pathway network components, we 1) identified 10 disease-associated and 30 function-associated pathways 2) revealed calcium signaling pathway and neuroactive ligand-receptor interaction as the most enriched, statistically significant pathways from the enrichment analysis, 3) showed calcium signaling pathways and MAPK signaling pathway to be interactive hubs with other pathways and also to be involved with pervasively present biological processes, 4) found convergent indications that the process "calcium-PRC (protein kinase C)-Ras-Raf-MAPK/ERK" is likely a major contributor to ASD pathophysiology, and 5) noted that perturbations associated with KEGG's category of environmental information processing were common. These findings support the idea that ASD-associated genes may contribute not only to core features of ASD themselves but also to vulnerability to other chronic and systemic problems potentially including cancer, metabolic conditions

  7. Bacitracin and nisin resistance in Staphylococcus aureus: a novel pathway involving the BraS/BraR two-component system (SA2417/SA2418) and both the BraD/BraE and VraD/VraE ABC transporters.

    PubMed

    Hiron, Aurélia; Falord, Mélanie; Valle, Jaione; Débarbouillé, Michel; Msadek, Tarek

    2011-08-01

    Two-component systems (TCSs) are key regulatory pathways allowing bacteria to adapt their genetic expression to environmental changes. Bacitracin, a cyclic dodecylpeptide antibiotic, binds to undecaprenyl pyrophosphate, the lipid carrier for cell wall precursors, effectively inhibiting peptidoglycan biosynthesis. We have identified a novel and previously uncharacterized TCS in the major human pathogen Staphylococcus aureus that we show to be essential for bacitracin and nisin resistance: the BraS/BraR system (Bacitracin resistance associated; SA2417/SA2418). The braRS genes are located immediately upstream from genes encoding an ABC transporter, accordingly designated BraDE. We have shown that the BraSR/BraDE module is a key bacitracin and nisin resistance determinant in S. aureus. In the presence of low antibiotic concentrations, BraSR activate transcription of two operons encoding ABC transporters: braDE and vraDE. We identified a highly conserved imperfect palindromic sequence upstream from the braDE and vraDE promoter sequences, essential for their transcriptional activation by BraSR, suggesting it is the likely BraR binding site. We demonstrated that the two ABC transporters play distinct and original roles in antibiotic resistance: BraDE is involved in bacitracin sensing and signalling through BraSR, whereas VraDE acts specifically as a detoxification module and is sufficient to confer bacitracin and nisin resistance when produced on its own. We show that these processes require functional BraD and VraD nucleotide-binding domain proteins, and that the large extracellular loop of VraE confers its specificity in bacitracin resistance. This is the first example of a TCS associated with two ABC transporters playing separate roles in signal transduction and antibiotic resistance. PMID:21696458

  8. Proteomic Analysis Reveals a Novel Mutator S (MutS) Partner Involved in Mismatch Repair Pathway.

    PubMed

    Chen, Zhen; Tran, Mykim; Tang, Mengfan; Wang, Wenqi; Gong, Zihua; Chen, Junjie

    2016-04-01

    The mismatch repair (MMR) family is a highly conserved group of proteins that function in correcting base-base and insertion-deletion mismatches generated during DNA replication. Disruption of this process results in characteristic microsatellite instability (MSI), repair defects, and susceptibility to cancer. However, a significant fraction of MSI-positive cancers express MMR genes at normal levels and do not carry detectable mutation in known MMR genes, suggesting that additional factors and/or mechanisms may exist to explain these MSI phenotypes in patients. To systematically investigate the MMR pathway, we conducted a proteomic analysis and identified MMR-associated protein complexes using tandem-affinity purification coupled with mass spectrometry (TAP-MS) method. The mass spectrometry data have been deposited to the ProteomeXchange with identifier PXD003014 and DOI 10.6019/PXD003014. We identified 230 high-confidence candidate interaction proteins (HCIPs). We subsequently focused on MSH2, an essential component of the MMR pathway and uncovered a novel MSH2-binding partner, WDHD1. We further demonstrated that WDHD1 forms a stable complex with MSH2 and MSH3 or MSH6,i.e.the MutS complexes. The specific MSH2/WDHD1 interaction is mediated by the second lever domain of MSH2 and Ala(1123)site of WDHD1. Moreover, we showed that, just like MSH2-deficient cells, depletion of WDHD1 also led to 6-thioguanine (6-TG) resistance, indicating that WDHD1 likely contributes to the MMR pathway. Taken together, our study uncovers new components involved in the MMR pathway, which provides candidate genes that may be responsible for the development of MSI-positive cancers. PMID:27037360

  9. Involvement of DNA Damage Response Pathways in Hepatocellular Carcinoma

    PubMed Central

    Yang, Sheau-Fang; Wei, Ren-Jie; Shiue, Yow-Ling; Wang, Shen-Nien

    2014-01-01

    Hepatocellular carcinoma (HCC) has been known as one of the most lethal human malignancies, due to the difficulty of early detection, chemoresistance, and radioresistance, and is characterized by active angiogenesis and metastasis, which account for rapid recurrence and poor survival. Its development has been closely associated with multiple risk factors, including hepatitis B and C virus infection, alcohol consumption, obesity, and diet contamination. Genetic alterations and genomic instability, probably resulted from unrepaired DNA lesions, are increasingly recognized as a common feature of human HCC. Dysregulation of DNA damage repair and signaling to cell cycle checkpoints, known as the DNA damage response (DDR), is associated with a predisposition to cancer and affects responses to DNA-damaging anticancer therapy. It has been demonstrated that various HCC-associated risk factors are able to promote DNA damages, formation of DNA adducts, and chromosomal aberrations. Hence, alterations in the DDR pathways may accumulate these lesions to trigger hepatocarcinogenesis and also to facilitate advanced HCC progression. This review collects some of the most known information about the link between HCC-associated risk factors and DDR pathways in HCC. Hopefully, the review will remind the researchers and clinicians of further characterizing and validating the roles of these DDR pathways in HCC. PMID:24877058

  10. Probing a Coral Genome for Components of the Photoprotective Scytonemin Biosynthetic Pathway and the 2-Aminoethylphosphonate Pathway

    PubMed Central

    Shoguchi, Eiichi; Tanaka, Makiko; Takeuchi, Takeshi; Shinzato, Chuya; Satoh, Nori

    2013-01-01

    Genome sequences of the reef-building coral, Acropora digitifera, have been decoded. Acropora inhabits an environment with intense ultraviolet exposure and hosts the photosynthetic endosymbiont, Symbiodinium. Acropora homologs of all four genes necessary for biosynthesis of the photoprotective cyanobacterial compound, shinorine, are present. Among metazoans, these genes are found only in anthozoans. To gain further evolutionary insights into biosynthesis of photoprotective compounds and associated coral proteins, we surveyed the Acropora genome for 18 clustered genes involved in cyanobacterial synthesis of the anti-UV compound, scytonemin, even though it had not previously been detected in corals. We identified candidates for only 6 of the 18 genes, including tyrP, scyA, and scyB. Therefore, it does not appear that Acropora digitifera can synthesize scytonemin independently. On the other hand, molecular phylogenetic analysis showed that one tyrosinase gene is an ortholog of vertebrate tyrosinase genes and that the coral homologs, scyA and scyB, are similar to bacterial metabolic genes, phosphonopyruvate (ppyr) decarboxylase and glutamate dehydrogenase (GDH), respectively. Further genomic searches for ppyr gene-related biosynthetic components indicate that the coral possesses a metabolic pathway similar to the bacterial 2-aminoethylphosphonate (AEP) biosynthetic pathway. The results suggest that de novo synthesis of carbon-phosphorus compounds is performed in corals. PMID:23434798

  11. The Notch pathway mediates the angiotensin II-induced synthesis of extracellular matrix components in podocytes.

    PubMed

    Yao, Min; Wang, Xiaomei; Wang, Xiaomeng; Zhang, Tao; Chi, Yanqing; Gao, Feng

    2015-07-01

    The Notch pathway is known to contribute to the development of glomerular disease. Angiotensin II (Ang II), an important member of the renin-angiotensin system, stimulates the accumulation of extracellular matrix components in glomerular disease; however, the exact mechanisms involved remain to be elucidated. In the present study, we aimed to investigate the effects of the Notch pathway on the synthesis of extracellular matrix components in Ang II-stimulated podocytes. Mouse podocytes were stimulated with Ang II (10-6 mol/l). The activation of the Notch pathway was inhibited by a vector carrying short hairpin RNA (shRNA) targeting Notch1 (sh-Notch1) or by γ-secretase inhibitor (GSI). The protein levels of Notch1, Notch intracellular domain 1 (NICD1), hairy and enhancer of split-1 (Hes1), matrix metalloproteinase (MMP)-2, MMP-9, transforming growth factor-β1 (TGF-β1), type IV collagen and laminin were determined by western blot analysis. The Notch1, Hes1, MMP-2, MMP-9, TGF-β1, type IV collagen and laminin mRNA levels were detected by RT-PCR. The MMP-2 and MMP-9 activity was measured using a cell active fluorescence assay kit. The levels of TGF-β1, type IV collagen and laminin were determined in the culture medium of the podocytes by enzyme-linked immunosorbent assay (ELISA). Our results revealed that Ang II upregulated Notch1, NICD1, Hes1, TGF-β1, type IV collagen and laminin expression and downregulated MMP-2 and MMP-9 expression in the cultured podocytes. The inhibition of the Notch pathway by sh-Notch1 or GSI increased MMP-2 and MMP-9 expression, decreased the TGF-β1 level and suppressed type IV collagen and laminin expression. The inhibition of the Notch pathway by sh-Notch1 or GSI also increased MMP-2 and MMP-9 activity, and decreased TGF-β1 levels, type IV collagen levels and laminin secretion. These findings indicate that the Notch pathway potentially mediates the Ang II-induced synthesis of extracellular matrix components in podocytes through the

  12. BIODEGRADATION OF TRICHLOROETHYLENE AND INVOLVEMENT OF AN AROMATIC BIODEGRADATIVE PATHWAY

    EPA Science Inventory

    Biodegradation of trichloroethylene (TCE) by the bacterial isolate strain G4 resulted in complete dechlorination of the compound as indicated by the production of inorganic chloride. A component of the water from which strain G4 was isolated that was required for TCE degradation ...

  13. Descending motor pathways and the spinal motor system - Limbic and non-limbic components

    NASA Technical Reports Server (NTRS)

    Holstege, Gert

    1991-01-01

    Research on descending motor pathways to caudal brainstem and spinal cord in the spinal motor system is reviewed. Particular attention is given to somatic and autonomic motoneurons in the spinal cord and brainstem, local projections to motoneurons, bulbospinal interneurons projecting to motoneurons, descending pathways of somatic motor control systems, and descending pathways involved in limbic motor control systems.

  14. Chronic alcohol exposure affects the cell components involved in membrane traffic in neuronal dendrites.

    PubMed

    Romero, Ana M; Renau-Piqueras, Jaime; Marín, M Pilar; Esteban-Pretel, Guillermo

    2015-01-01

    The specific traffic of the membrane components in neurons is a major requirement to establish and maintain neuronal domains-the axonal and the somatodendritic domains-and their polarized morphology. Unlike axons, dendrites contain membranous organelles, which are involved in the secretory pathway, including the endoplasmic reticulum, the Golgi apparatus and post-Golgi apparatus carriers, the cytoskeleton, and plasma membrane. A variety of molecules and factors are also involved in this process. Previous studies have shown that chronic alcohol exposure negatively affects several of these cell components, such as the Golgi apparatus or cytoskeleton in neurons. Yet very little information is available on the possible effects of this exposure on the remaining cell elements involved in intracellular trafficking in neurons, particularly in dendrites. By qualitative and quantitative electron microscopy, immunofluorescence and immunoblotting, we herein show that chronic exposure to moderate levels (30 mM) of ethanol in cultured neurons reduces the volume and surface density of the rough endoplasmic reticulum, and increases the levels of GRP78, a chaperone involved in endoplasmic reticulum stress. Ethanol also significantly diminishes the proportion of neurons that show an extension of Golgi into dendrites and dendritic Golgi outposts, a structure present exclusively in longer, thicker apical dendrites. Both Golgi apparatus types were also fragmented into a large number of cells. We also investigated the effect of alcohol on the levels of microtubule-based motor proteins KIF5, KIF17, KIFC2, dynein, and myosin IIb, responsible for transporting different cargoes in dendrites. Of these, alcohol differently affects several of them by lowering dynein and raising KIF5, KIFC2, and myosin IIb. These results, together with other previously published ones, suggest that practically all the protein trafficking steps in dendrites are altered to a greater or lesser extent by chronic

  15. Trichoderma biocontrol: signal transduction pathways involved in host sensing and mycoparasitism.

    PubMed

    Zeilinger, Susanne; Omann, Markus

    2007-01-01

    Fungi of the genus Trichoderma are used as biocontrol agents against several plant pathogenic fungi like Rhizoctonia spp., Pythium spp., Botrytis cinerea and Fusarium spp. which cause both soil-borne and leaf- or flower-borne diseases of agricultural plants. Plant disease control by Trichoderma is based on complex interactions between Trichoderma, the plant pathogen and the plant. Until now, two main components of biocontrol have been identified: direct activity of Trichoderma against the plant pathogen by mycoparasitism and induced systemic resistance in plants. As the mycoparasitic interaction is host-specific and not merely a contact response, it is likely that signals from the host fungus are recognised by Trichoderma and provoke transcription of mycoparasitism-related genes. In the last few years examination of signalling pathways underlying Trichoderma biocontrol started and it was shown that heterotrimeric G-proteins and mitogen-activated protein (MAP) kinases affected biocontrol-relevant processes such as the production of hydrolytic enzymes and antifungal metabolites and the formation of infection structures. MAPK signalling was also found to be involved in induction of plant systemic resistance in Trichoderma virens and in the hyperosmotic stress response in Trichoderma harzianum. Analyses of the function of components of the cAMP pathway during Trichoderma biocontrol revealed that mycoparasitism-associated coiling and chitinase production as well as secondary metabolism are affected by the internal cAMP level; in addition, a cross talk between regulation of light responses and the cAMP signalling pathway was found in Trichoderma atroviride. PMID:19936091

  16. Pathways involved in trifluoperazine-, dibucaine- and praziquantel-induced hemolysis.

    PubMed

    Malheiros, S V; Meirelles, N C; de Paula, E

    2000-01-17

    This work elucidates differences in the hemolytic pathway developed by the antipsychotic trifluoperazine (TFP), the local anesthetic dibucaine (DBC) and the antihelminthic praziquantel (PZQ). Their partition coefficients (P) were measured at pH 7.4 between n-octanol, microsomes, liposomes, erythrocyte ghosts and n-octanol/water. The effective drug:lipid molar ratios for the onset of membrane solubilization (ReSAT) and complete hemolysis (ReSOL) were calculated from the experimental P values and compared with a classical surface-active compound treatment Lichtenberg, D. Biochim. Biophys. Acta 821 (1985) 470-478[. The contribution of charged/uncharged forms of TFP and DBC for the hemolytic activity was also analyzed. In all cases the hemolytic phenomena could be related to the monomeric drug insertion into the membrane. Only for TFP at isosmotic condition lysis occurs at concentrations beyond the CMC of the drug, indicating that micellization facilitates TFP hemolytic effect, while DBC and PZQ reach a real membrane saturation at their monomeric form. PMID:10672415

  17. Multiple MONOPTEROS-dependent pathways are involved in leaf initiation.

    PubMed

    Schuetz, Mathias; Berleth, Thomas; Mattsson, Jim

    2008-10-01

    Initiation of leaves at the flanks of the shoot apical meristem occurs at sites of auxin accumulation and pronounced expression of auxin-inducible PIN-FORMED1 (PIN) genes, suggesting a feedback loop to progressively focus auxin in concrete spots. Because PIN expression is regulated by auxin response factor activity, including MONOPTEROS (MP), it appeared possible that MP affects leaf formation as a positive regulator of PIN genes and auxin transport. Here, we analyze a novel, completely leafless phenotype arising from simultaneous interference with both auxin signaling and auxin transport. We show that mp pin1 double mutants, as well as mp mutants treated with auxin-efflux inhibitors, display synergistic abnormalities not seen in wild type regardless of how strongly auxin transport was reduced. The synergism of abnormalities indicates that the role of MP in shoot meristem organization is not limited to auxin transport regulation. In the mp mutant background, auxin transport inhibition completely abolishes leaf formation. Instead of forming leaves, the abnormal shoot meristems dramatically increase in size, harboring correspondingly enlarged expression domains of CLAVATA3 and SHOOTMERISTEMLESS, molecular markers for the central stem cell zone and the complete meristem, respectively. The observed synergism under conditions of auxin efflux inhibition was further supported by an unrestricted PIN1 expression in mp meristems, as compared to a partial restriction in wild-type meristems. Auxin transport-inhibited mp meristems also lacked detectable auxin maxima. We conclude that MP promotes the focusing of auxin and leaf initiation in part through pathways not affected by auxin efflux inhibitors. PMID:18685044

  18. CD39/Adenosine Pathway Is Involved in AIDS Progression

    PubMed Central

    Limou, Sophie; Younas, Mehwish; Kök, Ayrin; Huë, Sophie; Seddiki, Nabila; Hulin, Anne; Delaneau, Olivier; Schuitemaker, Hanneke; Herbeck, Joshua T.; Mullins, James I.; Muhtarova, Maria; Bensussan, Armand; Zagury, Jean-François; Lelievre, Jean-Daniel; Lévy, Yves

    2011-01-01

    HIV-1 infection is characterized by a chronic activation of the immune system and suppressed function of T lymphocytes. Regulatory CD4+ CD25high FoxP3+CD127low T cells (Treg) play a key role in both conditions. Here, we show that HIV-1 positive patients have a significant increase of Treg-associated expression of CD39/ENTPD1, an ectoenzyme which in concert with CD73 generates adenosine. We show in vitro that the CD39/adenosine axis is involved in Treg suppression in HIV infection. Treg inhibitory effects are relieved by CD39 down modulation and are reproduced by an adenosine-agonist in accordance with a higher expression of the adenosine A2A receptor on patients' T cells. Notably, the expansion of the Treg CD39+ correlates with the level of immune activation and lower CD4+ counts in HIV-1 infected patients. Finally, in a genetic association study performed in three different cohorts, we identified a CD39 gene polymorphism that was associated with down-modulated CD39 expression and a slower progression to AIDS. PMID:21750674

  19. Lysine Acetylation: Elucidating the Components of an Emerging Global Signaling Pathway in Trypanosomes

    PubMed Central

    Alonso, Victoria Lucia; Serra, Esteban Carlos

    2012-01-01

    In the past ten years the number of acetylated proteins reported in literature grew exponentially. Several authors have proposed that acetylation might be a key component in most eukaryotic signaling pathways, as important as phosphorylation. The enzymes involved in this process are starting to emerge; acetyltransferases and deacetylases are found inside and outside the nuclear compartment and have different regulatory functions. In trypanosomatids several of these enzymes have been described and are postulated to be novel antiparasitic targets for the rational design of drugs. In this paper we overview the most important known acetylated proteins and the advances made in the identification of new acetylated proteins using high-resolution mass spectrometry. Also, we summarize what is known so far about the acetyltransferases and deacetylases in eukaryotes, focusing on trypanosomes and their potential use as chemotherapeutic targets. PMID:23093844

  20. The Budding Yeast Ubiquitin Protease Ubp7 Is a Novel Component Involved in S Phase Progression.

    PubMed

    Böhm, Stefanie; Szakal, Barnabas; Herken, Benjamin W; Sullivan, Meghan R; Mihalevic, Michael J; Kabbinavar, Faiz F; Branzei, Dana; Clark, Nathan L; Bernstein, Kara A

    2016-02-26

    DNA damage must be repaired in an accurate and timely fashion to preserve genome stability. Cellular mechanisms preventing genome instability are crucial to human health because genome instability is considered a hallmark of cancer. Collectively referred to as the DNA damage response, conserved pathways ensure proper DNA damage recognition and repair. The function of numerous DNA damage response components is fine-tuned by posttranslational modifications, including ubiquitination. This not only involves the enzyme cascade responsible for conjugating ubiquitin to substrates but also requires enzymes that mediate directed removal of ubiquitin. Deubiquitinases remove ubiquitin from substrates to prevent degradation or to mediate signaling functions. The Saccharomyces cerevisiae deubiquitinase Ubp7 has been characterized previously as an endocytic factor. However, here we identify Ubp7 as a novel factor affecting S phase progression after hydroxyurea treatment and demonstrate an evolutionary and genetic interaction of Ubp7 with DNA damage repair pathways of homologous recombination and nucleotide excision repair. We find that deletion of UBP7 sensitizes cells to hydroxyurea and cisplatin and demonstrate that factors that stabilize replication forks are critical under these conditions. Furthermore, ubp7Δ cells exhibit an S phase progression defect upon checkpoint activation by hydroxyurea treatment. ubp7Δ mutants are epistatic to factors involved in histone maintenance and modification, and we find that a subset of Ubp7 is chromatin-associated. In summary, our results suggest that Ubp7 contributes to S phase progression by affecting the chromatin state at replication forks, and we propose histone H2B ubiquitination as a potential substrate of Ubp7. PMID:26740628

  1. Phytohormones Signaling Pathways and ROS Involvement in Seed Germination

    PubMed Central

    Oracz, Krystyna; Karpiński, Stanisław

    2016-01-01

    Phytohormones and reactive oxygen species (ROS) are major determinants of the regulation of development and stress responses in plants. During life cycle of these organisms, signaling networks of plant growth regulators and ROS interact in order to render an appropriate developmental and environmental response. In plant’s photosynthetic (e.g., leaves) and non-photosynthetic (e.g., seeds) tissues, enhanced and suboptimal ROS production is usually associated with stress, which in extreme cases can be lethal to cells, a whole organ or even an organism. However, controlled production of ROS is appreciated for cellular signaling. Despite the current progress that has been made in plant biology and increasing number of findings that have revealed roles of ROS and hormonal signaling in germination, some questions still arise, e.g., what are the downstream protein targets modified by ROS enabling stimulus-specific cellular responses of the seed? Or which molecular regulators allow ROS/phytohormones interactions and what is their function in seed life? In this particular review the role of some transcription factors, kinases and phosphatases is discussed, especially those which usually known to be involved in ROS and hormonal signal transduction under stress in plants, may also play a role in the regulation of processes occurring in seeds. The summarized recent findings regarding particular ROS- and phytohormones-related regulatory proteins, as well as their integration, allowed to propose a novel, possible model of action of LESION SIMULATING DISEASE 1, ENHANCED DISEASE SUSCEPTIBILITY 1, and PHYTOALEXIN DEFICIENT 4 functioning during seeds life. PMID:27379144

  2. Cinnamon polyphenols regulate multiple metabolic pathways involved in intestinal lipid metabolism of primary small intestinal enterocytes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Increasing evidence suggests that dietary factors may affect the expression of multiple genes and signaling pathways including those that regulate intestinal lipoprotein metabolism. The small intestine is actively involved in the regulation of dietary lipid absorption, intracellular transport and me...

  3. Transplantation of prokaryotic two-component signaling pathways into mammalian cells

    PubMed Central

    Hansen, Jonathan; Mailand, Erik; Swaminathan, Krishna Kumar; Schreiber, Joerg; Angelici, Bartolomeo; Benenson, Yaakov

    2014-01-01

    Signaling pathway engineering is a promising route toward synthetic biological circuits. Histidine–aspartate phosphorelays are thought to have evolved in prokaryotes where they form the basis for two-component signaling. Tyrosine-serine–threonine phosphorelays, exemplified by MAP kinase cascades, are predominant in eukaryotes. Recently, a prokaryotic two-component pathway was implemented in a plant species to sense environmental trinitrotoluene. We reasoned that “transplantation” of two-component pathways into mammalian host could provide an orthogonal and diverse toolkit for a variety of signal processing tasks. Here we report that two-component pathways could be partially reconstituted in mammalian cell culture and used for programmable control of gene expression. To enable this reconstitution, coding sequences of histidine kinase (HK) and response regulator (RR) components were codon-optimized for human cells, whereas the RRs were fused with a transactivation domain. Responsive promoters were furnished by fusing DNA binding sites in front of a minimal promoter. We found that coexpression of HKs and their cognate RRs in cultured mammalian cells is necessary and sufficient to strongly induce gene expression even in the absence of pathways’ chemical triggers in the medium. Both loss-of-function and constitutive mutants behaved as expected. We further used the two-component signaling pathways to implement two-input logical AND, NOR, and OR gene regulation. Thus, two-component systems can be applied in different capacities in mammalian cells and their components can be used for large-scale synthetic gene circuits. PMID:25331891

  4. The IgE-dependent pathway in allergic transfusion reactions: involvement of donor blood allergens other than plasma proteins.

    PubMed

    Matsuyama, Nobuki; Yasui, Kazuta; Amakishi, Etsuko; Hayashi, Tomoya; Kuroishi, Ayumu; Ishii, Hiroyuki; Matsukura, Harumichi; Tani, Yoshihiko; Furuta, Rika A; Hirayama, Fumiya

    2015-07-01

    On transfusion, several plasma proteins can cause anaphylaxis in patients deficient in the corresponding plasma proteins. However, little is known about other allergens, which are encountered much more infrequently. Although it has been speculated that an allergen-independent pathway underlying allergic transfusion reactions (ATRs) is elicited by biological response modifiers accumulated in blood components during storage, the exact mechanisms remain unresolved. Furthermore, it is difficult even to determine whether ATRs are induced via allergen-dependent or allergen-independent pathways. To distinguish these two pathways in ATR cases, we established a basophil activation test, in which the basophil-activating ability of supernatants of residual transfused blood of ATR cases to whole blood basophils was assessed in the presence or absence of dasatinib, an inhibitor of IgE-mediated basophil activation. Three of 37 supernatants from the platelet concentrates with ATRs activated panel blood basophils in the absence, but not in the presence, of dasatinib. The basophil activation was inhibited by treatment of anti-fish collagen I MoAb in one case, suggesting that the involvement of fish allergens may have been present in donor plasma. We concluded that unknown non-plasma proteins, some of which had epitopes similar to fish antigens, in blood component may be involved in ATRs via an allergen/IgE-dependent pathway. PMID:25840771

  5. Disentangling metabolic pathways involved in copper resistance in Candida fukuyamaensis RCL-3 indigenous yeast.

    PubMed

    Irazusta, Verónica; Michel, Lucas; de Figueroa, Lucía I C

    2016-07-01

    Candida fukuyamaensis RCL-3 yeast strain isolated from a copper filter plant is able to lower copper concentration in culture medium. In the present study, effect of copper in proteins expression and mechanisms involved in copper resistance were explored using comparative proteomics. Mono-dimensional gel electrophoresis revealed differential band expressions between cells grown with or without copper. 2-DE analysis of C. fukuyamaensis RCL-3 revealed that copper exposure produced at least an over-expression of 40 proteins. Sixteen proteins were identified and grouped in four categories according to their functions: glycolysis and ATP production, synthesis of proteins, oxidative stress response, and processing and transport of proteins. Integral membrane proteins and membrane-associated proteins were analyzed, showing nine protein bands over-expressed in Cu-supplemented medium. Four proteins were identified, namely nucleoporin pom152, elongation factor 2, copper chaperone Sod1 Ccs1, and eiosome component Lsp1. The proteomic analysis performed allowed the identification of different metabolic pathways and certain proteins involved in metal input and storage related to cell ability to bioremediate copper. These proteins and mechanisms could be used for future applications of C. fukuyamaensis RCL-3 in biotechnological processes such as remediation of heavy metals. PMID:26568043

  6. Systems analysis of gene ontology and biological pathways involved in post-myocardial infarction responses

    PubMed Central

    2015-01-01

    Background Pathway analysis has been widely used to gain insight into essential mechanisms of the response to myocardial infarction (MI). Currently, there exist multiple pathway databases that organize molecular datasets and manually curate pathway maps for biological interpretation at varying forms of organization. However, inconsistencies among different databases in pathway descriptions, frequently due to conflicting results in the literature, can generate incorrect interpretations. Furthermore, although pathway analysis software provides detailed images of interactions among molecules, it does not exhibit how pathways interact with one another or with other biological processes under specific conditions. Methods We propose a novel method to standardize descriptions of enriched pathways for a set of genes/proteins using Gene Ontology terms. We used this method to examine the relationships among pathways and biological processes for a set of condition-specific genes/proteins, represented as a functional biological pathway-process network. We applied this algorithm to a set of 613 MI-specific proteins we previously identified. Results A total of 96 pathways from Biocarta, KEGG, and Reactome, and 448 Gene Ontology Biological Processes were enriched with these 613 proteins. The pathways were represented as Boolean functions of biological processes, delivering an interactive scheme to organize enriched information with an emphasis on involvement of biological processes in pathways. We extracted a network focusing on MI to demonstrate that tyrosine phosphorylation of Signal Transducer and Activator of Transcription (STAT) protein, positive regulation of collagen metabolic process, coagulation, and positive/negative regulation of blood coagulation have immediate impacts on the MI response. Conclusions Our method organized biological processes and pathways in an unbiased approach to provide an intuitive way to identify biological properties of pathways under specific

  7. Synthesis of classical pathway complement components by chondrocytes.

    PubMed Central

    Bradley, K; North, J; Saunders, D; Schwaeble, W; Jeziorska, M; Woolley, D E; Whaley, K

    1996-01-01

    Using immunohistochemical studies, C1q, C1s, C4 and C2 were detected in chondrocytes in normal human articular cartilage and macroscopically normal articular cartilage from the inferior surfaces of hip joints of patients with osteoarthritis. Using reverse-transcribed polymerase chain reaction (RT-PCR), mRNA for C1q, C1s, C4 and C2 was also detected in RNA extracted from articular cartilage. C1r, C3, C1-inhibitor, C4-binding protein and factor I were not detected by either technique. Articular chondrocytes cultured in vitro synthesized C1r, C1s, C4, C2, C3 and C1-inhibitor but not C1q, C4-binding protein or factor I, as assessed by enzyme-linked immunosorbent assay (ELISA) and Northern blot analysis. Thus cultured articular chondrocytes have a complement profile that is similar to that of cultured human fibroblasts rather than that of articular chondrocytes in vivo. Complement synthesis in cultured chondrocytes was modulated by the cytokines interleukin-1 beta (IL-1 beta), tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma), showing that cytokines can probably regulate complement synthesis in intact cartilage. The possible roles of local synthesis of complement components by chondrocytes in matrix turnover and the regulation chondrocyte function are discussed. Images Figure 1 Figure 2 Figure 4 PMID:8881771

  8. A second component of the SltA-dependent cation tolerance pathway in Aspergillus nidulans.

    PubMed

    Mellado, Laura; Calcagno-Pizarelli, Ana Maria; Lockington, Robin A; Cortese, Marc S; Kelly, Joan M; Arst, Herbert N; Espeso, Eduardo A

    2015-09-01

    The transcriptional response to alkali metal cation stress is mediated by the zinc finger transcription factor SltA in Aspergillus nidulans and probably in other fungi of the pezizomycotina subphylum. A second component of this pathway has been identified and characterized. SltB is a 1272 amino acid protein with at least two putative functional domains, a pseudo-kinase and a serine-endoprotease, involved in signaling to the transcription factor SltA. Absence of SltB activity results in nearly identical phenotypes to those observed for a null sltA mutant. Hypersensitivity to a variety of monovalent and divalent cations, and to medium alkalinization are among the phenotypes exhibited by a null sltB mutant. Calcium homeostasis is an exception and this cation improves growth of sltΔ mutants. Moreover, loss of kinase HalA in conjunction with loss-of-function sltA or sltB mutations leads to pronounced calcium auxotrophy. sltA sltB double null mutants display a cation stress sensitive phenotype indistinguishable from that of single slt mutants showing the close functional relationship between these two proteins. This functional relationship is reinforced by the fact that numerous mutations in both slt loci can be isolated as suppressors of poor colonial growth resulting from certain null vps (vacuolar protein sorting) mutations. In addition to allowing identification of sltB, our sltB missense mutations enabled prediction of functional regions in the SltB protein. Although the relationship between the Slt and Vps pathways remains enigmatic, absence of SltB, like that of SltA, leads to vacuolar hypertrophy. Importantly, the phenotypes of selected sltA and sltB mutations demonstrate that suppression of null vps mutations is not dependent on the inability to tolerate cation stress. Thus a specific role for both SltA and SltB in the VPS pathway seems likely. Finally, it is noteworthy that SltA and SltB have a similar, limited phylogenetic distribution, being restricted to

  9. The molecular components of the extracellular protein-degradation pathways of the ectomycorrhizal fungus Paxillus involutus

    PubMed Central

    Shah, Firoz; Rineau, Francois; Canbäck, Björn; Johansson, Tomas; Tunlid, Anders

    2013-01-01

    Proteins contribute to a major part of the organic nitrogen (N) in forest soils. This N is mobilized and becomes available to trees as a result of the depolymerizing activities of symbiotic ectomycorrhizal fungi. The mechanisms by which these fungi depolymerize proteins and assimilate the released N are poorly characterized. Biochemical analysis and transcriptome profiling were performed to examine the proteolytic machinery and the uptake system of the ectomycorrhizal basidiomycete Paxillus involutus during the assimilation of organic N from various protein sources and extracts of organic matter. All substrates induced secretion of peptidase activity with an acidic pH optimum, mostly contributed by aspartic peptidases. The peptidase activity was transiently repressed by ammonium. Transcriptional analysis revealed a large number of extracellular endo- and exopeptidases. The expression levels of these peptidases were regulated in parallel with transporters and enzymes involved in the assimilation and metabolism of the released peptides and amino acids. For the first time the molecular components of the protein degradation pathways of an ectomycorrhizal fungus are described. The data suggest that the transcripts encoding these components are regulated in response to the chemical properties and the availability of the protein substrates. PMID:23902518

  10. Two glycosyltransferases involved in anthocyanin modification delineated by transcriptome independent component analysis in Arabidopsis thaliana

    PubMed Central

    Yonekura-Sakakibara, Keiko; Fukushima, Atsushi; Nakabayashi, Ryo; Hanada, Kousuke; Matsuda, Fumio; Sugawara, Satoko; Inoue, Eri; Kuromori, Takashi; Ito, Takuya; Shinozaki, Kazuo; Wangwattana, Bunyapa; Yamazaki, Mami; Saito, Kazuki

    2012-01-01

    To identify candidate genes involved in Arabidopsis flavonoid biosynthesis, we applied transcriptome coexpression analysis and independent component analyses with 1388 microarray data from publicly available databases. Two glycosyltransferases, UGT79B1 and UGT84A2 were found to cluster with anthocyanin biosynthetic genes. Anthocyanin was drastically reduced in ugt79b1 knockout mutants. Recombinant UGT79B1 protein converted cyanidin 3-O-glucoside to cyanidin 3-O-xylosyl(1→2)glucoside. UGT79B1 recognized 3-O-glucosylated anthocyanidins/flavonols and uridine diphosphate (UDP)-xylose, but not 3,5-O-diglucosylated anthocyanidins, indicating that UGT79B1 encodes anthocyanin 3-O-glucoside: 2′′-O-xylosyltransferase. UGT84A2 is known to encode sinapic acid: UDP-glucosyltransferase. In ugt84a2 knockout mutants, a major sinapoylated anthocyanin was drastically reduced. A comparison of anthocyanin profiles in ugt84a knockout mutants indicated that UGT84A2 plays a major role in sinapoylation of anthocyanin, and that other UGT84As contribute the production of 1-O-sinapoylglucose to a lesser extent. These data suggest major routes from cyanidin 3-O-glucoside to the most highly modified cyanidin in the potential intricate anthocyanin modification pathways in Arabidopsis. PMID:21899608

  11. Evolution of JAK-STAT Pathway Components: Mechanisms and Role in Immune System Development

    PubMed Central

    Liongue, Clifford; O'Sullivan, Lynda A.; Trengove, Monique C.; Ward, Alister C.

    2012-01-01

    Background Lying downstream of a myriad of cytokine receptors, the Janus kinase (JAK) – Signal transducer and activator of transcription (STAT) pathway is pivotal for the development and function of the immune system, with additional important roles in other biological systems. To gain further insight into immune system evolution, we have performed a comprehensive bioinformatic analysis of the JAK-STAT pathway components, including the key negative regulators of this pathway, the SH2-domain containing tyrosine phosphatase (SHP), Protein inhibitors against Stats (PIAS), and Suppressor of cytokine signaling (SOCS) proteins across a diverse range of organisms. Results Our analysis has demonstrated significant expansion of JAK-STAT pathway components co-incident with the emergence of adaptive immunity, with whole genome duplication being the principal mechanism for generating this additional diversity. In contrast, expansion of upstream cytokine receptors appears to be a pivotal driver for the differential diversification of specific pathway components. Conclusion Diversification of JAK-STAT pathway components during early vertebrate development occurred concurrently with a major expansion of upstream cytokine receptors and two rounds of whole genome duplications. This produced an intricate cell-cell communication system that has made a significant contribution to the evolution of the immune system, particularly the emergence of adaptive immunity. PMID:22412924

  12. Stenotrophomonas maltophilia PhoP, a Two-Component Response Regulator, Involved in Antimicrobial Susceptibilities.

    PubMed

    Liu, Ming-Che; Tsai, Yi-Lin; Huang, Yi-Wei; Chen, Hsing-Yu; Hsueh, Po-Ren; Lai, Szu-Yu; Chen, Li-Chia; Chou, Yi-Hwa; Lin, Wen-Yuan; Liaw, Shwu-Jen

    2016-01-01

    Stenotrophomonas maltophilia, a gram-negative bacterium, has increasingly emerged as an important nosocomial pathogen. It is well-known for resistance to a variety of antimicrobial agents including cationic antimicrobial polypeptides (CAPs). Resistance to polymyxin B, a kind of CAPs, is known to be controlled by the two-component system PhoPQ. To unravel the role of PhoPQ in polymyxin B resistance of S. maltophilia, a phoP mutant was constructed. We found MICs of polymyxin B, chloramphenicol, ampicillin, gentamicin, kanamycin, streptomycin and spectinomycin decreased 2-64 fold in the phoP mutant. Complementation of the phoP mutant by the wild-type phoP gene restored all of the MICs to the wild type levels. Expression of PhoP was shown to be autoregulated and responsive to Mg2+ levels. The polymyxin B and gentamicin killing tests indicated that pretreatment of low Mg2+ can protect the wild-type S. maltophilia from killing but not phoP mutant. Interestingly, we found phoP mutant had a decrease in expression of SmeZ, an efflux transporter protein for aminoglycosides in S. maltophilia. Moreover, phoP mutant showed increased permeability in the cell membrane relative to the wild-type. In summary, we demonstrated the two-component regulator PhoP of S. maltophilia is involved in antimicrobial susceptibilities and low Mg2+ serves as a signal for triggering the pathway. Both the alteration in membrane permeability and downregulation of SmeZ efflux transporter in the phoP mutant contributed to the increased drug susceptibilities of S. maltophilia, in particular for aminoglycosides. This is the first report to describe the role of the Mg2+-sensing PhoP signaling pathway of S. maltophilia in regulation of the SmeZ efflux transporter and in antimicrobial susceptibilities. This study suggests PhoPQ TCS may serve as a target for development of antimicrobial agents against multidrug-resistant S. maltophilia. PMID:27159404

  13. Stenotrophomonas maltophilia PhoP, a Two-Component Response Regulator, Involved in Antimicrobial Susceptibilities

    PubMed Central

    Huang, Yi-Wei; Chen, Hsing-Yu; Hsueh, Po-Ren; Lai, Szu-Yu; Chen, Li-Chia; Chou, Yi-Hwa; Lin, Wen-Yuan; Liaw, Shwu-Jen

    2016-01-01

    Stenotrophomonas maltophilia, a gram-negative bacterium, has increasingly emerged as an important nosocomial pathogen. It is well-known for resistance to a variety of antimicrobial agents including cationic antimicrobial polypeptides (CAPs). Resistance to polymyxin B, a kind of CAPs, is known to be controlled by the two-component system PhoPQ. To unravel the role of PhoPQ in polymyxin B resistance of S. maltophilia, a phoP mutant was constructed. We found MICs of polymyxin B, chloramphenicol, ampicillin, gentamicin, kanamycin, streptomycin and spectinomycin decreased 2–64 fold in the phoP mutant. Complementation of the phoP mutant by the wild-type phoP gene restored all of the MICs to the wild type levels. Expression of PhoP was shown to be autoregulated and responsive to Mg2+ levels. The polymyxin B and gentamicin killing tests indicated that pretreatment of low Mg2+ can protect the wild-type S. maltophilia from killing but not phoP mutant. Interestingly, we found phoP mutant had a decrease in expression of SmeZ, an efflux transporter protein for aminoglycosides in S. maltophilia. Moreover, phoP mutant showed increased permeability in the cell membrane relative to the wild-type. In summary, we demonstrated the two-component regulator PhoP of S. maltophilia is involved in antimicrobial susceptibilities and low Mg2+ serves as a signal for triggering the pathway. Both the alteration in membrane permeability and downregulation of SmeZ efflux transporter in the phoP mutant contributed to the increased drug susceptibilities of S. maltophilia, in particular for aminoglycosides. This is the first report to describe the role of the Mg2+-sensing PhoP signaling pathway of S. maltophilia in regulation of the SmeZ efflux transporter and in antimicrobial susceptibilities. This study suggests PhoPQ TCS may serve as a target for development of antimicrobial agents against multidrug-resistant S. maltophilia. PMID:27159404

  14. Regulation of Arabidopsis MAPKKK18 by ABI1 and SnRK2, components of the ABA signaling pathway.

    PubMed

    Tajdel, Małgorzata; Mituła, Filip; Ludwików, Agnieszka

    2016-04-01

    The plant hormone abscisic acid (ABA), a key regulator in many crucial developmental and physiological processes, recruits diverse components into precisely regulated signaling network. We recently discovered that MAPKKK18, an ABA-activated kinase, is regulated by the protein phosphatase type 2C (PP2C) ABI1 and the kinase SnRK2.6, both components of the ABA core signaling pathway. ABI1 acts to inhibit MAPKKK18 kinase activity, but also affects MAPKKK18 protein turnover via the ubiquitin-proteasome pathway. SnRK2.6 kinase also seems to be important for the regulation of MAPKKK18 function. In this review we summarize the mechanisms that are exclusively involved in MAPKKK18 kinase regulation and that ensure specificity in its activation. PMID:26852793

  15. Current perspectives of molecular pathways involved in chronic inflammation-mediated breast cancer.

    PubMed

    Suman, Shankar; Sharma, Pradeep Kumar; Rai, Girish; Mishra, Sanjay; Arora, Deepika; Gupta, Prachi; Shukla, Yogeshwer

    2016-04-01

    Inflammation has multifaceted role in cancer progression including initiation, promotion and invasion by affecting the immune surveillance and associated signaling pathways. Inflammation facilitates the over-expression of cytokines, chemokines and growth factors involved in progression of different cancers including breast cancer progression. Deregulation of biological processes such as oxidative stress, angiogenesis, and autophagy elicit favorable immune response towards chronic inflammation. Apart from the role in carcinogenesis, chronic inflammation also favors the emergence of drug resistance clones by inducing the growth of breast cancer stem-like cells. Immunomodulation mediated by cytokines, chemokines and several other growth factors present in the tumor microenvironment regulate chronic inflammatory response and alter crosstalk among various signaling pathways such as NF-κB, Nrf-2, JAK-STAT, Akt and MAPKs involved in the progression of breast cancer. In this review, we focused on cellular and molecular processes involved in chronic inflammation, crosstalk among different signaling pathways and their association in breast cancer pathogenesis. PMID:26522220

  16. Involvement of matrix metalloproteinases (MMPs) and inflammasome pathway in molecular mechanisms of fibrosis.

    PubMed

    Robert, Sacha; Gicquel, Thomas; Victoni, Tatiana; Valença, Samuel; Barreto, Emiliano; Bailly-Maître, Béatrice; Boichot, Elisabeth; Lagente, Vincent

    2016-08-01

    Fibrosis is a basic connective tissue lesion defined by the increase in the fibrillar extracellular matrix (ECM) components in tissue or organ. Matrix metalloproteinases (MMPs) are a major group of proteases known to regulate the turn-over of ECM and so they are suggested to be important in tissue remodelling observed during fibrogenic process associated with chronic inflammation. Tissue remodelling is the result of an imbalance in the equilibrium of the normal processes of synthesis and degradation of ECM components markedly controlled by the MMPs/TIMP imbalance. We previously showed an association of the differences in collagen deposition in the lungs of bleomycin-treated mice with a reduced molar pro-MMP-9/TIMP-1 ratio. Using the carbon tetrachloride (CCl4) preclinical model of liver fibrosis in mice, we observed a significant increase in collagen deposition with increased expression and release of tissue inhibitors of metalloproteinase (TIMP)-1 both at 24 h and 3 weeks later. This suggests an early altered regulation of matrix turnover involved in the development of fibrosis. We also demonstrated an activation of NLRP3-inflammasome pathway associated with the IL-1R/MyD88 signalling in the development of experimental fibrosis both in lung and liver. This was also associated with an increased expression of purinergic receptors mainly P2X7 Finally, these observations emphasize those effective therapies for these disorders must be given early in the natural history of the disease, prior to the development of tissue remodelling and fibrosis. PMID:27247426

  17. Involvement of matrix metalloproteinases (MMPs) and inflammasome pathway in molecular mechanisms of fibrosis

    PubMed Central

    Robert, Sacha; Gicquel, Thomas; Victoni, Tatiana; Valença, Samuel; Barreto, Emiliano; Bailly-Maître, Béatrice; Boichot, Elisabeth; Lagente, Vincent

    2016-01-01

    Fibrosis is a basic connective tissue lesion defined by the increase in the fibrillar extracellular matrix (ECM) components in tissue or organ. Matrix metalloproteinases (MMPs) are a major group of proteases known to regulate the turn-over of ECM and so they are suggested to be important in tissue remodelling observed during fibrogenic process associated with chronic inflammation. Tissue remodelling is the result of an imbalance in the equilibrium of the normal processes of synthesis and degradation of ECM components markedly controlled by the MMPs/TIMP imbalance. We previously showed an association of the differences in collagen deposition in the lungs of bleomycin-treated mice with a reduced molar pro-MMP-9/TIMP-1 ratio. Using the carbon tetrachloride (CCl4) preclinical model of liver fibrosis in mice, we observed a significant increase in collagen deposition with increased expression and release of tissue inhibitors of metalloproteinase (TIMP)-1 both at 24 h and 3 weeks later. This suggests an early altered regulation of matrix turnover involved in the development of fibrosis. We also demonstrated an activation of NLRP3-inflammasome pathway associated with the IL-1R/MyD88 signalling in the development of experimental fibrosis both in lung and liver. This was also associated with an increased expression of purinergic receptors mainly P2X7. Finally, these observations emphasize those effective therapies for these disorders must be given early in the natural history of the disease, prior to the development of tissue remodelling and fibrosis. PMID:27247426

  18. PATHWAY-BASED ANALYSIS FOR GENOME-WIDE ASSOCIATION STUDIES USING SUPERVISED PRINCIPAL COMPONENTS

    PubMed Central

    Chen, Xi; Wang, Lily; Hu, Bo; Guo, Mingsheng; Barnard, John; Zhu, Xiaofeng

    2012-01-01

    Many complex diseases are influenced by genetic variations in multiple genes, each with only a small marginal effect on disease susceptibility. Pathway analysis, which identifies biological pathways associated with disease outcome, has become increasingly popular for genome-wide association studies (GWAS). In addition to combining weak signals from a number of SNPs in the same pathway, results from pathway analysis also shed light on the biological processes underlying disease. We propose a new pathway-based analysis method for GWAS, the supervised principal component analysis (SPCA) model. In the proposed SPCA model, a selected subset of SNPs most associated with disease outcome is used to estimate the latent variable for a pathway. The estimated latent variable for each pathway is an optimal linear combination of a selected subset of SNPs; therefore, the proposed SPCA model provides the ability to borrow strength across the SNPs in a pathway. In addition to identifying pathways associated with disease outcome, SPCA also carries out additional within-category selection to identify the most important SNPs within each gene set. The proposed model operates in a well-established statistical framework and can handle design information such as covariate adjustment and matching information in GWAS. We compare the proposed method with currently available methods using data with realistic linkage disequilibrium structures and we illustrate the SPCA method using the Wellcome Trust Case-Control Consortium Crohn Disease (CD) dataset. PMID:20842628

  19. UNCOVERING GENETIC COMPONENTS INVOLVED IN EARLY REGULATORY IMMUNE RESPONSE DURING PRRSV INFECTION

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Our goal is to identify the most significant pathways involved in early immune responses during porcine reproductive and respiratory syndrome virus (PRRSV) infection as compared to protective vaccination. For this experiment PRRSV-naïve animals were divided into four groups: (1) pigs infected with A...

  20. Uniform curation protocol of metazoan signaling pathways to predict novel signaling components.

    PubMed

    Pálfy, Máté; Farkas, Illés J; Vellai, Tibor; Korcsmáros, Tamás

    2013-01-01

    A relatively large number of signaling databases available today have strongly contributed to our understanding of signaling pathway properties. However, pathway comparisons both within and across databases are currently severely hampered by the large variety of data sources and the different levels of detail of their information content (on proteins and interactions). In this chapter, we present a protocol for a uniform curation method of signaling pathways, which intends to overcome this insufficiency. This uniformly curated database called SignaLink ( http://signalink.org ) allows us to systematically transfer pathway annotations between different species, based on orthology, and thereby to predict novel signaling pathway components. Thus, this method enables the compilation of a comprehensive signaling map of a given species and identification of new potential drug targets in humans. We strongly believe that the strict curation protocol we have established to compile a signaling pathway database can also be applied for the compilation of other (e.g., metabolic) databases. Similarly, the detailed guide to the orthology-based prediction of novel signaling components across species may also be utilized for predicting components of other biological processes. PMID:23715991

  1. Genome-Wide Pathway Association Studies of Multiple Correlated Quantitative Phenotypes Using Principle Component Analyses

    PubMed Central

    Zhang, Feng; Guo, Xiong; Wu, Shixun; Han, Jing; Liu, Yongjun; Shen, Hui; Deng, Hong-Wen

    2012-01-01

    Genome-wide pathway association studies provide novel insight into the biological mechanism underlying complex diseases. Current pathway association studies primarily focus on single important disease phenotype, which is sometimes insufficient to characterize the clinical manifestations of complex diseases. We present a multi-phenotypes pathway association study(MPPAS) approach using principle component analysis(PCA). In our approach, PCA is first applied to multiple correlated quantitative phenotypes for extracting a set of orthogonal phenotypic components. The extracted phenotypic components are then used for pathway association analysis instead of original quantitative phenotypes. Four statistics were proposed for PCA-based MPPAS in this study. Simulations using the real data from the HapMap project were conducted to evaluate the power and type I error rates of PCA-based MPPAS under various scenarios considering sample sizes, additive and interactive genetic effects. A real genome-wide association study data set of bone mineral density (BMD) at hip and spine were also analyzed by PCA-based MPPAS. Simulation studies illustrated the performance of PCA-based MPPAS for identifying the causal pathways underlying complex diseases. Genome-wide MPPAS of BMD detected associations between BMD and KENNY_CTNNB1_TARGETS_UP as well as LONGEVITYPATHWAY pathways in this study. We aim to provide a applicable MPPAS approach, which may help to gain deep understanding the potential biological mechanism of association results for complex diseases. PMID:23285279

  2. The Heparan and Heparin Metabolism Pathway is Involved in Regulation of Fatty Acid Composition

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Six genes involved in the heparan sulfate and heparin metabolism pathway, DSEL (dermatan sulfate epimerase-like), EXTL1 (exostoses (multiple)-like 1), HS6ST1 (heparan sulfate 6-O-sulfotransferase 1), HS6ST3 (heparan sulfate 6-O-sulfotransferase 3), NDST3 (N-deacetylase/N-sulfotransferase (heparan gl...

  3. TEC protein tyrosine kinase is involved in the Erk signaling pathway induced by HGF

    SciTech Connect

    Li, Feifei; Jiang, Yinan; Zheng, Qiping; Yang, Xiaoming; Wang, Siying

    2011-01-07

    Research highlights: {yields} TEC is rapidly tyrosine-phosphorylated and activated by HGF-stimulation in vivo or after partial hepatectomy in mice. {yields} TEC enhances the activity of Elk and serum response element (SRE) in HGF signaling pathway in hepatocyte. {yields} TEC promotes hepatocyte proliferation through the Erk-MAPK pathway. -- Abstract: Background/aims: TEC, a member of the TEC family of non-receptor type protein tyrosine kinases, has recently been suggested to play a role in hepatocyte proliferation and liver regeneration. This study aims to investigate the putative mechanisms of TEC kinase regulation of hepatocyte differentiation, i.e. to explore which signaling pathway TEC is involved in, and how TEC is activated in hepatocyte after hepatectomy and hepatocyte growth factor (HGF) stimulation. Methods: We performed immunoprecipitation (IP) and immunoblotting (IB) to examine TEC tyrosine phosphorylation after partial hepatectomy in mice and HGF stimulation in WB F-344 hepatic cells. The TEC kinase activity was determined by in vitro kinase assay. Reporter gene assay, antisense oligonucleotide and TEC dominant negative mutant (TEC{sup KM}) were used to examine the possible signaling pathways in which TEC is involved. The cell proliferation rate was evaluated by {sup 3}H-TdR incorporation. Results: TEC phosphorylation and kinase activity were increased in 1 h after hepatectomy or HGF treatment. TEC enhanced the activity of Elk and serum response element (SRE). Inhibition of MEK1 suppressed TEC phosphorylation. Blocking TEC activity dramatically decreased the activation of Erk. Reduced TEC kinase activity also suppressed the proliferation of WB F-344 cells. These results suggest TEC is involved in the Ras-MAPK pathway and acts between MEK1 and Erk. Conclusions: TEC promotes hepatocyte proliferation and regeneration and is involved in HGF-induced Erk signaling pathway.

  4. Components of the dorsal-ventral pathway also contribute to anterior-posterior patterning in honeybee embryos (Apis mellifera)

    PubMed Central

    2014-01-01

    Background A key early step in embryogenesis is the establishment of the major body axes; the dorsal-ventral (DV) and anterior-posterior (AP) axes. Determination of these axes in some insects requires the function of different sets of signalling pathways for each axis. Patterning across the DV axis requires interaction between the Toll and Dpp/TGF-β pathways, whereas patterning across the AP axis requires gradients of bicoid/orthodenticle proteins and the actions of a hierarchy of gene transcription factors. We examined the expression and function of Toll and Dpp signalling during honeybee embryogenesis to assess to the role of these genes in DV patterning. Results Pathway components that are required for dorsal specification in Drosophila are expressed in an AP-restricted pattern in the honeybee embryo, including Dpp and its receptor Tkv. Components of the Toll pathway are expressed in a more conserved pattern along the ventral axis of the embryo. Late-stage embryos from RNA interference (RNAi) knockdown of Toll and Dpp pathways had both DV and AP patterning defects, confirmed by staining with Am-sna, Am-zen, Am-eve, and Am-twi at earlier stages. We also identified two orthologues of dorsal in the honeybee genome, with one being expressed during embryogenesis and having a minor role in axis patterning, as determined by RNAi and the other expressed during oogenesis. Conclusions We found that early acting pathways (Toll and Dpp) are involved not only in DV patterning but also AP patterning in honeybee embryogenesis. Changes to the expression patterns and function of these genes may reflect evolutionary changes in the placement of the extra-embryonic membranes during embryogenesis with respect to the AP and DV axes. PMID:24620747

  5. Relevant Networks involving the p53 Signalling Pathway in Renal Cell Carcinoma

    PubMed Central

    Villaamil, V. Medina; Gallego, G. Aparicio; Caínzos, I. Santamarina; Ruvira, L. Valbuena; Valladares-Ayerbes, M.; Aparicio, L. M. Antón

    2011-01-01

    Introduction: Renal cell carcinoma is the most common type of kidney cancer. A better understanding of the critical pathways and interactions associated with alterations in renal function and renal tumour properties is required. Our final goal is to combine the knowledge provided by a regulatory network with experimental observations provided by the dataset. Methods: In this study, a systems biology approach was used, integrating immunohistochemistry protein expression profiles and protein interaction information with the STRING and MeV bioinformatics tools. A group consisting of 80 patients with renal cell carcinoma was studied. The expression of selected markers was assessed using tissue microarray technology on immunohistochemically stained slides. The immunohistochemical data of the molecular factors studied were analysed using a parametric statistical test, Pearson’s correlation coefficient test. Results: Bioinformatics analysis of tumour samples resulted in 2 protein networks. The first network consists of proteins involved in the angiogenesis pathway and the apoptosis suppressor, BCL2, and includes both positive and negative correlations. The second network shows a negative interaction between the p53 tumour suppressor protein and the glucose transporter type 4. Conclusion: The comprehensive pathway network will help us to realise the cooperative behaviours among pathways. Regulation of metabolic pathways is an important role of p53. The pathway involving the tumour suppressor gene p53 could regulate tumour angiogenesis. Further investigation of the proteins that interact with this pathway in this type of tumour may provide new strategies for cancer therapies to specifically inhibit the molecules that play crucial roles in tumour progression. PMID:23675247

  6. Signaling pathways involved in PDGF-evoked cellular responses in human RPE cells

    SciTech Connect

    Hollborn, Margrit . E-mail: hollbm@medizin.uni-leipzig.de; Bringmann, Andreas; Faude, Frank; Wiedemann, Peter; Kohen, Leon

    2006-06-09

    We examined whether PDGF may directly stimulate the expression of VEGF by retinal pigment epithelial (RPE) cells in vitro, and the involvement of three signal transduction pathways in the regulation of PDGF-evoked cell proliferation, migration, and production of VEGF-A was investigated. PDGF stimulated the gene and protein expression of VEGF-A by RPE cells, and increased cell proliferation and chemotaxis. PDGF activated all signaling pathways investigated, as determined by increased phosphorylation levels of ERK1/2, p38, and Akt proteins. The three signaling pathways were involved in the mediation of PDGF-evoked cell proliferation, while p38 and PI3K mediated cell migration, and PI3K mediated secretion of VEGF-A. In addition to VEGF-A, the cells expressed mRNAs for various members of the VEGF family and for their receptors, including VEGF-B, -C, -D, flt-1, and KDR. The data indicate that PDGF selectively stimulates the expression of VEGF-A in RPE cells. PDGF evokes at least three signal transduction pathways which are differentially involved in various cellular responses.

  7. Boolean Modeling of Cellular and Molecular Pathways Involved in Influenza Infection

    PubMed Central

    Anderson, Christopher S.; DeDiego, Marta L.; Topham, David J.; Thakar, Juilee

    2016-01-01

    Systems virology integrates host-directed approaches with molecular profiling to understand viral pathogenesis. Self-contained statistical approaches that combine expression profiles of genes with the available databases defining the genes involved in the pathways (gene-sets) have allowed characterization of predictive gene-signatures associated with outcome of the influenza virus (IV) infection. However, such enrichment techniques do not take into account interactions among pathways that are responsible for the IV infection pathogenesis. We investigate dendritic cell response to seasonal H1N1 influenza A/New Caledonia/20/1999 (NC) infection and infer the Boolean logic rules underlying the interaction network of ligand induced signaling pathways and transcription factors. The model reveals several novel regulatory modes and provides insights into mechanism of cross talk between NFκB and IRF mediated signaling. Additionally, the logic rule underlying the regulation of IL2 pathway that was predicted by the Boolean model was experimentally validated. Thus, the model developed in this paper integrates pathway analysis tools with the dynamic modeling approaches to reveal the regulation between signaling pathways and transcription factors using genome-wide transcriptional profiles measured upon influenza infection. PMID:26981147

  8. Conserved features and evolutionary shifts of the EDA signaling pathway involved in vertebrate skin appendage development.

    PubMed

    Pantalacci, Sophie; Chaumot, Arnaud; Benoît, Gérard; Sadier, Alexa; Delsuc, Frédéric; Douzery, Emmanuel J P; Laudet, Vincent

    2008-05-01

    It is widely accepted that evolutionary changes in conserved developmental signaling pathways play an important role in morphological evolution. However, few in silico studies were interested in tracking such changes in a signaling pathway. The Ectodysplasin (EDA) pathway provides an opportunity to fill this gap because it is involved in vertebrate skin appendage development such as scales, teeth, hair, and feathers that take an obvious part in the adaptation of species to their environment. We benefited from the large amount of genomic data now available to explore the evolution of the upstream genes of the EDA pathway. In mammals, these genes are eda (encoding 2 ligands, EDA-A1 and EDA-A2), edar (EDA-A1 receptor), edaradd (EDA receptor [EDAR] adapter), xedar (EDA-A2 receptor), and troy (a XEDAR-related receptor). We show that the evolution of EDA pathway genes combines both strongly conserved features and evolutionary shifts. These shifts are found at different signaling levels (from the ligand to intracellular signaling) and at different taxonomic levels (class, suborder, and genera). Although conserved features likely participate to the similarities found in the early development of vertebrate skin appendages, these shifts might account for innovations and specializations. Moreover, our study demonstrates that we can now benefit from the large number of sequenced vertebrate genomes to explore the evolution of specific signaling pathways and thereby to open new perspectives for developmental biology and evolutionary developmental biology. PMID:18304980

  9. Boolean Modeling of Cellular and Molecular Pathways Involved in Influenza Infection.

    PubMed

    Anderson, Christopher S; DeDiego, Marta L; Topham, David J; Thakar, Juilee

    2016-01-01

    Systems virology integrates host-directed approaches with molecular profiling to understand viral pathogenesis. Self-contained statistical approaches that combine expression profiles of genes with the available databases defining the genes involved in the pathways (gene-sets) have allowed characterization of predictive gene-signatures associated with outcome of the influenza virus (IV) infection. However, such enrichment techniques do not take into account interactions among pathways that are responsible for the IV infection pathogenesis. We investigate dendritic cell response to seasonal H1N1 influenza A/New Caledonia/20/1999 (NC) infection and infer the Boolean logic rules underlying the interaction network of ligand induced signaling pathways and transcription factors. The model reveals several novel regulatory modes and provides insights into mechanism of cross talk between NFκB and IRF mediated signaling. Additionally, the logic rule underlying the regulation of IL2 pathway that was predicted by the Boolean model was experimentally validated. Thus, the model developed in this paper integrates pathway analysis tools with the dynamic modeling approaches to reveal the regulation between signaling pathways and transcription factors using genome-wide transcriptional profiles measured upon influenza infection. PMID:26981147

  10. Expression profile of critical genes involved in FGF signaling pathway in the developing human primary dentition.

    PubMed

    Huang, Feng; Hu, Xiaoxiao; Fang, Chunni; Liu, Hong; Lin, Chensheng; Zhang, Yanding; Hu, Xuefeng

    2015-11-01

    Mammalian tooth development is regulated by paracrine signal molecules of several conserved family interactions between epithelium and mesenchyme. The expression patterns and regulative roles of FGF signaling have been extensively studied in the mouse odontogenesis; however, that is not well known in human tooth development. In order to unveil the molecular mechanisms that regulate human tooth morphogenesis, we examined the expression patterns of the critical molecules involved in FGF signaling pathway in the developing human tooth germ by in situ hybridization, immunohistochemistry, and real-time RT-PCR, including FGF ligands, receptors, and intracellular transducer. We found overlapping but distinct expression pattern of FGF ligands and receptors in the different stages and components. Expression of FGF4, FGF7, FGF8, and FGF9 persists widespread in human tooth mesenchyme, which is quite different to that of in mouse. FGFR1 may be the major receptor in regulate mechanisms of FGF signals in human tooth development. Real-time RT-PCR indeed confirmed the results of in situ hybridization. Results of K-Ras, p-ERK1/2, p-p38, p-JNK, and p-PDK1 expression reveal spatial and temporal patterns of FGF signaling during morphogenesis and organogenesis of human tooth germ. Activity of the FGF signaling transducer protein in human tooth germ was much higher than that of in mouse. Our results provided important FGF singling information in the developing process, pinpoint to the domains where the downstream target genes of FGF signaling can be sought, and enlightened our knowledge about the nature of FGF signaling in human tooth germ. PMID:26266341

  11. An ATM-independent S-phase checkpoint response involves CHK1 pathway

    NASA Technical Reports Server (NTRS)

    Zhou, Xiang-Yang; Wang, Xiang; Hu, Baocheng; Guan, Jun; Iliakis, George; Wang, Ya

    2002-01-01

    After exposure to genotoxic stress, proliferating cells actively slow down the DNA replication through a S-phase checkpoint to provide time for repair. We report that in addition to the ataxia-telangiectasia mutated (ATM)-dependent pathway that controls the fast response, there is an ATM-independent pathway that controls the slow response to regulate the S-phase checkpoint after ionizing radiation in mammalian cells. The slow response of S-phase checkpoint, which is resistant to wortmannin, sensitive to caffeine and UCN-01, and related to cyclin-dependent kinase phosphorylation, is much stronger in CHK1 overexpressed cells, and it could be abolished by Chk1 antisense oligonucleotides. These results provide evidence that the ATM-independent slow response of S-phase checkpoint involves CHK1 pathway.

  12. The mitochondrial and death receptor pathways involved in the thymocytes apoptosis induced by aflatoxin B1.

    PubMed

    Peng, Xi; Yu, Zhengqiang; Liang, Na; Chi, Xiaofeng; Li, Xiaochong; Jiang, Min; Fang, Jing; Cui, Hengmin; Lai, Weimin; Zhou, Yi; Zhou, Shan

    2016-03-15

    Aflatoxin B1 (AFB1) is a potent immunosuppressive agent in endotherms, which can be related to the up-regulated apoptosis of immune organs. In this study, we investigated the roles of the mitochondrial, death receptor, and endoplasmic reticulum pathways in Aflatoxin B1 induced thymocytes apoptosis. Chickens were fed an aflatoxin B1 containing diet (0.6 mg/kg AFB1) for 3 weeks. Our results showed that (1) AFB1 diet induced the decrease of T-cell subsets, morphological changes, and excessive apoptosis of thymus. (2) The excessive apoptosis involved the mitochondrial pathway (up-regulation of Bax, Bak, cytC and down-regulation of Bcl-2 and Bcl-xL) and death receptor pathway (up-regulation of FasL, Fas and FADD). (3) Oxidative stress, an apoptosis inducer, was confirmed in the thymus. In conclusion, this is the first study to demonstrate that mitochondrial and death receptor pathways involved in AFB1 induced thymocytes apoptosis in broilers. PMID:26933817

  13. RICTOR involvement in the PI3K/AKT pathway regulation in melanocytes and melanoma

    PubMed Central

    Laugier, Florence; Finet-Benyair, Adeline; André, Jocelyne; Rachakonda, P. Sivaramakrishna; Kumar, Rajiv; Bensussan, Armand; Dumaz, Nicolas

    2015-01-01

    Several studies have highlighted the importance of the PI3K pathway in melanocytes and its frequent over-activation in melanoma. However, little is known about regulation of the PI3K pathway in melanocytic cells. We showed that normal human melanocytes are less sensitive to selective PI3K or mTOR inhibitors than to dual PI3K/mTOR inhibitors. The resistance to PI3K inhibitor was due to a rapid AKT reactivation limiting the inhibitor effect on proliferation. Reactivation of AKT was linked to a feedback mechanism involving the mTORC2 complex and in particular its scaffold protein RICTOR. RICTOR overexpression in melanocytes disrupted the negative feedback, activated the AKT pathway and stimulated clonogenicity highlighting the importance of this feedback to restrict melanocyte proliferation. We found that the RICTOR locus is frequently amplified and overexpressed in melanoma and that RICTOR over-expression in NRAS-transformed melanocytes stimulates their clonogenicity, demonstrating that RICTOR amplification can cooperate with NRAS mutation to stimulate melanoma proliferation. These results show that RICTOR plays a central role in PI3K pathway negative feedback in melanocytes and that its deregulation could be involved in melanoma development. PMID:26356562

  14. The mitochondrial and death receptor pathways involved in the thymocytes apoptosis induced by aflatoxin B1

    PubMed Central

    Chi, Xiaofeng; Li, Xiaochong; Jiang, Min; Fang, Jing; Cui, Hengmin; Lai, Weimin; Zhou, Yi; Zhou, Shan

    2016-01-01

    Aflatoxin B1 (AFB1) is a potent immunosuppressive agent in endotherms, which can be related to the up-regulated apoptosis of immune organs. In this study, we investigated the roles of the mitochondrial, death receptor, and endoplasmic reticulum pathways in Aflatoxin B1 induced thymocytes apoptosis. Chickens were fed an aflatoxin B1 containing diet (0.6 mg/kg AFB1) for 3 weeks. Our results showed that (1) AFB1 diet induced the decrease of T-cell subsets, morphological changes, and excessive apoptosis of thymus. (2) The excessive apoptosis involved the mitochondrial pathway (up-regulation of Bax, Bak, cytC and down-regulation of Bcl-2 and Bcl-xL) and death receptor pathway (up-regulation of FasL, Fas and FADD). (3) Oxidative stress, an apoptosis inducer, was confirmed in the thymus. In conclusion, this is the first study to demonstrate that mitochondrial and death receptor pathways involved in AFB1 induced thymocytes apoptosis in broilers. PMID:26933817

  15. Human umbilical vein: involvement of cyclooxygenase-2 pathway in bradykinin B1 receptor-sensitized responses.

    PubMed

    Errasti, A E; Rey-Ares, V; Daray, F M; Rogines-Velo, M P; Sardi, S P; Paz, C; Podestá, E J; Rothlin, R P

    2001-08-01

    In isolated human umbilical vein (HUV), the contractile response to des-Arg9-bradykinin (des-Arg9-BK), selective BK B1 receptor agonist, increases as a function of the incubation time. Here, we evaluated whether cyclooxygenase (COX) pathway is involved in BK B1-sensitized response obtained in 5-h incubated HUV rings. The effect of different concentrations of indomethacin, sodium salicylate, ibuprofen, meloxicam, lysine clonixinate or NS-398 administrated 30 min before concentration-response curves (CRC) was studied. All treatments produced a significant rightward shift of the CRC to des-Arg9-BK in a concentration-dependent manner, which provides pharmacological evidence that COX pathway is involved in the BK B1 responses. Moreover, in this tissue, the NS-398 pKb (5.2) observed suggests that COX-2 pathway is the most relevant. The strong correlation between published pIC50 for COX-2 and the NSAIDs' pKbs estimated further supports the hypothesis that COX-2 metabolites are involved in BK B1 receptor-mediated responses. In other rings, indomethacin (30, 100 micromol/l) or NS-398 (10, 30 micromol/l) produced a significant rightward shift of the CRC to BK, selective BK B2 agonist, and its pKbs were similar to the values to inhibit BK B1 receptor responses, suggesting that COX-2 pathway also is involved in BK B2 receptor responses. Western blot analysis shows that COX-1 and COX-2 isoenzymes are present before and after 5-h in vitro incubation and apparently COX-2 does not suffer additional induction. PMID:11534854

  16. Proteins involved on TGF-β pathway are up-regulated during the acute phase of experimental Chagas disease.

    PubMed

    Ferreira, Roberto Rodrigues; de Souza, Elen Mello; de Oliveira, Fabiane Loiola; Ferrão, Patrícia Mello; Gomes, Leonardo Henrique Ferreira; Mendonça-Lima, Leila; Meuser-Batista, Marcelo; Bailly, Sabine; Feige, Jean Jacques; de Araujo-Jorge, Tania Cremonini; Waghabi, Mariana Caldas

    2016-05-01

    Studies developed by our group in the last years have shown the involvement of TGF-β in acute and chronic Chagas heart disease, with elevated plasma levels and activated TGF-β cell signaling pathway as remarkable features of patients in the advanced stages of this disease, when high levels of cardiac fibrosis is present. Imbalance in synthesis and degradation of extracellular matrix components is the basis of pathological fibrosis and TGF-β is considered as one of the key regulators of this process. In the present study, we investigated the activity of the TGF-β signaling pathway, including receptors and signaling proteins activation in the heart of animals experimentally infected with Trypanosoma cruzi during the period that mimics the acute phase of Chagas disease. We observed that T. cruzi-infected animals presented increased expression of TGF-β receptors. Overexpression of receptors was followed by an increased phosphorylation of Smad2/3, p38 and ERK. Furthermore, we correlated these activities with cellular factors involved in the fibrotic process induced by TGF-β. We observed that the expression of collagen I, fibronectin and CTGF were increased in the heart of infected animals on day 15 post-infection. Correlated with the increased TGF-β activity in the heart, we found that serum levels of total TGF-β were significantly higher during acute infection. Taken together, our data suggest that the commitment of the heart associates with increased activity of TGF-β pathway and expression of its main components. Our results, confirm the importance of this cytokine in the development and maintenance of cardiac damage caused by T. cruzi infection. PMID:26852285

  17. Structural Basis for the Function of Complement Component C4 within the Classical and Lectin Pathways of Complement.

    PubMed

    Mortensen, Sofia; Kidmose, Rune T; Petersen, Steen V; Szilágyi, Ágnes; Prohászka, Zoltan; Andersen, Gregers R

    2015-06-01

    Complement component C4 is a central protein in the classical and lectin pathways within the complement system. During activation of complement, its major fragment C4b becomes covalently attached to the surface of pathogens and altered self-tissue, where it acts as an opsonin marking the surface for removal. Moreover, C4b provides a platform for assembly of the proteolytically active convertases that mediate downstream complement activation by cleavage of C3 and C5. In this article, we present the crystal and solution structures of the 195-kDa C4b. Our results provide the molecular details of the rearrangement accompanying C4 cleavage and suggest intramolecular flexibility of C4b. The conformations of C4b and its paralogue C3b are shown to be remarkably conserved, suggesting that the convertases from the classical and alternative pathways are likely to share their overall architecture and mode of substrate recognition. We propose an overall molecular model for the classical pathway C5 convertase in complex with C5, suggesting that C3b increases the affinity for the substrate by inducing conformational changes in C4b rather than a direct interaction with C5. C4b-specific features revealed by our structural studies are probably involved in the assembly of the classical pathway C3/C5 convertases and C4b binding to regulators. PMID:25911760

  18. Involvement of the ethylene response pathway in dormancy induction in chrysanthemum

    PubMed Central

    Sumitomo, Katsuhiko; Satoh, Shigeru; Hisamatsu, Tamotsu

    2008-01-01

    Temperature plays a significant role in the annual cycling between growth and dormancy of the herbaceous perennial chrysanthemum (Chrysanthemum morifolium Ramat.). After exposure to high summer temperatures, cool temperature triggers dormancy. The cessation of flowering and rosette formation by the cessation of elongation are characteristic of dormant plants, and can be stimulated by exogenous ethylene. Thus, the ethylene response pathway may be involved in temperature-induced dormancy of chrysanthemum. Transgenic chrysanthemums expressing a mutated ethylene receptor gene were used to assess this involvement. The transgenic lines showed reduced ethylene sensitivity: ethylene causes leaf yellowing in wild-type chrysanthemums, but leaves remained green in the transgenic lines. Extension growth and flowering of wild-type and transgenic lines varied between temperatures: at 20 °C, the transgenic lines showed the same stem elongation and flowering as the wild type; at cooler temperatures, the wild type formed rosettes with an inability to flower and entered dormancy, but some transgenic lines continued to elongate and flower. This supports the involvement of the ethylene response pathway in the temperature-induced dormancy of chrysanthemum. At the highest dosage of ethephon, an ethylene-releasing agent, wild-type plants formed rosettes with an inability to flower and became dormant, but one transgenic line did not. This confirms that dormancy is induced via the ethylene response pathway. PMID:18952907

  19. Helpful Components Involved in the Cognitive-Experiential Model of Dream Work

    ERIC Educational Resources Information Center

    Tien, Hsiu-Lan Shelley; Chen, Shuh-Chi; Lin, Chia-Huei

    2009-01-01

    The purpose of the study was to examine the helpful components involved in the Hill's cognitive-experiential dream work model. Participants were 27 volunteer clients from colleges and universities in northern and central parts of Taiwan. Each of the clients received 1-2 sessions of dream interpretations. The cognitive-experiential dream work model…

  20. Transcriptome Analysis and Discovery of Genes Involved in Immune Pathways from Hepatopancreas of Microbial Challenged Mitten Crab Eriocheir sinensis

    PubMed Central

    Li, Xihong; Cui, Zhaoxia; Liu, Yuan; Song, Chengwen; Shi, Guohui

    2013-01-01

    Background The Chinese mitten crab Eriocheir sinensis is an important economic crustacean and has been seriously attacked by various diseases, which requires more and more information for immune relevant genes on genome background. Recently, high-throughput RNA sequencing (RNA-seq) technology provides a powerful and efficient method for transcript analysis and immune gene discovery. Methods/Principal Findings A cDNA library from hepatopancreas of E. sinensis challenged by a mixture of three pathogen strains (Gram-positive bacteria Micrococcus luteus, Gram-negative bacteria Vibrio alginolyticus and fungi Pichia pastoris; 108 cfu·mL−1) was constructed and randomly sequenced using Illumina technique. Totally 39.76 million clean reads were assembled to 70,300 unigenes. After ruling out short-length and low-quality sequences, 52,074 non-redundant unigenes were compared to public databases for homology searching and 17,617 of them showed high similarity to sequences in NCBI non-redundant protein (Nr) database. For function classification and pathway assignment, 18,734 (36.00%) unigenes were categorized to three Gene Ontology (GO) categories, 12,243 (23.51%) were classified to 25 Clusters of Orthologous Groups (COG), and 8,983 (17.25%) were assigned to six Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways. Potentially, 24, 14, 47 and 132 unigenes were characterized to be involved in Toll, IMD, JAK-STAT and MAPK pathways, respectively. Conclusions/Significance This is the first systematical transcriptome analysis of components relating to innate immune pathways in E. sinensis. Functional genes and putative pathways identified here will contribute to better understand immune system and prevent various diseases in crab. PMID:23874555

  1. Involvement of the PI3K/Akt/GSK3β pathway in photodynamic injury of neurons and glial cells

    NASA Astrophysics Data System (ADS)

    Komandirov, M. A.; Knyazeva, E. A.; Fedorenko, Y. P.; Rudkovskii, M. V.; Stetsurin, D. A.; Uzdensky, A. B.

    2011-03-01

    Photodynamic treatment causes intense oxidative stress and kills cells. It is currently used in neurooncology. However, along with tumor it damages surrounding healthy neuronal and glial cells. In order to study the possible role of the phosphatidylinositol 3-kinase/protein kinase Akt/glycogen synthase kinase-3β signaling pathway in photodynamic damage to normal neurons and glia, we used isolated crayfish stretch receptor that consists only of a single neuron surrounded by glial cells. It was photosensitized with alumophthalocyanine Photosens (100 nM). The laser diode (670nm, 0.4W/cm2) was used as a light source. Application of specific inhibitors of the enzymes involved in this pathway showed that phosphatidylinositol 3-kinase did not participate in photoinduced death of neurons and glia. Protein kinase Akt was involved in photoinduced necrosis but not in apoptosis of neurons and glia. Glycogen synthase kinase-3β participated in photoinduced apoptosis of glial cells and in necrosis of neurons. Therefore, the phosphatidylinositol 3-kinase/protein kinase Akt/glycogen synthase kinase-3β pathway was not involved as a whole in photodynamic injury of crayfish neurons and glial cells but its components, protein kinase Akt and glycogen synthase kinase-3β, independently and cell-specifically regulated photoinduced death of neurons and glial cells. These data showed that in this system necrosis was not non-regulated and catastrophic mode of cell death. It was controlled by some signaling proteins. The obtained results may be used for search of pharmacological agents that selectively modulate injury of normal neurons and glial cells during photodynamic therapy of brain tumors.

  2. Involvement of the PI3K/Akt/GSK3β pathway in photodynamic injury of neurons and glial cells

    NASA Astrophysics Data System (ADS)

    Komandirov, M. A.; Knyazeva, E. A.; Fedorenko, Y. P.; Rudkovskii, M. V.; Stetsurin, D. A.; Uzdensky, A. B.

    2010-10-01

    Photodynamic treatment causes intense oxidative stress and kills cells. It is currently used in neurooncology. However, along with tumor it damages surrounding healthy neuronal and glial cells. In order to study the possible role of the phosphatidylinositol 3-kinase/protein kinase Akt/glycogen synthase kinase-3β signaling pathway in photodynamic damage to normal neurons and glia, we used isolated crayfish stretch receptor that consists only of a single neuron surrounded by glial cells. It was photosensitized with alumophthalocyanine Photosens (100 nM). The laser diode (670nm, 0.4W/cm2) was used as a light source. Application of specific inhibitors of the enzymes involved in this pathway showed that phosphatidylinositol 3-kinase did not participate in photoinduced death of neurons and glia. Protein kinase Akt was involved in photoinduced necrosis but not in apoptosis of neurons and glia. Glycogen synthase kinase-3β participated in photoinduced apoptosis of glial cells and in necrosis of neurons. Therefore, the phosphatidylinositol 3-kinase/protein kinase Akt/glycogen synthase kinase-3β pathway was not involved as a whole in photodynamic injury of crayfish neurons and glial cells but its components, protein kinase Akt and glycogen synthase kinase-3β, independently and cell-specifically regulated photoinduced death of neurons and glial cells. These data showed that in this system necrosis was not non-regulated and catastrophic mode of cell death. It was controlled by some signaling proteins. The obtained results may be used for search of pharmacological agents that selectively modulate injury of normal neurons and glial cells during photodynamic therapy of brain tumors.

  3. E-cadherin mediates contact inhibition of proliferation through Hippo signaling-pathway components

    PubMed Central

    Kim, Nam-Gyun; Koh, Eunjin; Chen, Xiao; Gumbiner, Barry M.

    2011-01-01

    Contact inhibition of cell growth is essential for embryonic development and maintenance of tissue architecture in adult organisms, and the growth of tumors is characterized by a loss of contact inhibition of proliferation. The recently identified Hippo signaling pathway has been implicated in contact inhibition of proliferation as well as organ size control. The modulation of the phosphorylation and nuclear localization of Yes-associated protein (YAP) by the highly conserved kinase cascade of the Hippo signaling pathway has been intensively studied. However, cell-surface receptors regulating the Hippo signaling pathway in mammals are not well understood. In this study, we show that Hippo signaling pathway components are required for E-cadherin–dependent contact inhibition of proliferation. Knockdown of the Hippo signaling components or overexpression of YAP inhibits the decrease in cell proliferation caused by E-cadherin homophilic binding at the cell surface, independent of other cell–cell interactions. We also demonstrate that the E-cadherin/catenin complex functions as an upstream regulator of the Hippo signaling pathway in mammalian cells. Expression of E-cadherin in MDA-MB-231 cells restores the density-dependent regulation of YAP nuclear exclusion. Knockdown of β-catenin in densely cultured MCF10A cells, which mainly depletes E-cadherin–bound β-catenin, induces a decrease in the phosphorylation of S127 residue of YAP and its nuclear accumulation. Moreover, E-cadherin homophilic binding independent of other cell interactions is sufficient to control the subcellular localization of YAP. Therefore, Our results indicate that, in addition to its role in cell–cell adhesion, E-cadherin-mediated cell–cell contact directly regulates the Hippo signaling pathway to control cell proliferation. PMID:21730131

  4. Human wild-type tau interacts with wingless pathway components and produces neurofibrillary pathology in Drosophila.

    PubMed

    Jackson, George R; Wiedau-Pazos, Martina; Sang, Tzu-Kang; Wagle, Naveed; Brown, Carlos A; Massachi, Sasan; Geschwind, Daniel H

    2002-05-16

    Pathologic alterations in the microtubule-associated protein tau have been implicated in a number of neurodegenerative disorders, including Alzheimer's disease (AD), progressive supranuclear palsy (PSP), and frontotemporal dementia (FTD). Here, we show that tau overexpression, in combination with phosphorylation by the Drosophila glycogen synthase kinase-3 (GSK-3) homolog and wingless pathway component (Shaggy), exacerbated neurodegeneration induced by tau overexpression alone, leading to neurofibrillary pathology in the fly. Furthermore, manipulation of other wingless signaling molecules downstream from shaggy demonstrated that components of the Wnt signaling pathway modulate neurodegeneration induced by tau pathology in vivo but suggested that tau phosphorylation by GSK-3beta differs from canonical Wnt effects on beta-catenin stability and TCF activity. The genetic system we have established provides a powerful reagent for identification of novel modifiers of tau-induced neurodegeneration that may serve as future therapeutic targets. PMID:12062036

  5. Association genetics and transcriptome analysis reveal a gibberellin-responsive pathway involved in regulating photosynthesis.

    PubMed

    Xie, Jianbo; Tian, Jiaxing; Du, Qingzhang; Chen, Jinhui; Li, Ying; Yang, Xiaohui; Li, Bailian; Zhang, Deqiang

    2016-05-01

    Gibberellins (GAs) regulate a wide range of important processes in plant growth and development, including photosynthesis. However, the mechanism by which GAs regulate photosynthesis remains to be understood. Here, we used multi-gene association to investigate the effect of genes in the GA-responsive pathway, as constructed by RNA sequencing, on photosynthesis, growth, and wood property traits, in a population of 435 Populus tomentosa By analyzing changes in the transcriptome following GA treatment, we identified many key photosynthetic genes, in agreement with the observed increase in measurements of photosynthesis. Regulatory motif enrichment analysis revealed that 37 differentially expressed genes related to photosynthesis shared two essential GA-related cis-regulatory elements, the GA response element and the pyrimidine box. Thus, we constructed a GA-responsive pathway consisting of 47 genes involved in regulating photosynthesis, including GID1, RGA, GID2, MYBGa, and 37 photosynthetic differentially expressed genes. Single nucleotide polymorphism (SNP)-based association analysis showed that 142 SNPs, representing 40 candidate genes in this pathway, were significantly associated with photosynthesis, growth, and wood property traits. Epistasis analysis uncovered interactions between 310 SNP-SNP pairs from 37 genes in this pathway, revealing possible genetic interactions. Moreover, a structural gene-gene matrix based on a time-course of transcript abundances provided a better understanding of the multi-gene pathway affecting photosynthesis. The results imply a functional role for these genes in mediating photosynthesis, growth, and wood properties, demonstrating the potential of combining transcriptome-based regulatory pathway construction and genetic association approaches to detect the complex genetic networks underlying quantitative traits. PMID:27091876

  6. Shade Avoidance Components and Pathways in Adult Plants Revealed by Phenotypic Profiling

    PubMed Central

    Nozue, Kazunari; Tat, An V.; Kumar Devisetty, Upendra; Robinson, Matthew; Mumbach, Maxwell R.; Ichihashi, Yasunori; Lekkala, Saradadevi; Maloof, Julin N.

    2015-01-01

    Shade from neighboring plants limits light for photosynthesis; as a consequence, plants have a variety of strategies to avoid canopy shade and compete with their neighbors for light. Collectively the response to foliar shade is called the shade avoidance syndrome (SAS). The SAS includes elongation of a variety of organs, acceleration of flowering time, and additional physiological responses, which are seen throughout the plant life cycle. However, current mechanistic knowledge is mainly limited to shade-induced elongation of seedlings. Here we use phenotypic profiling of seedling, leaf, and flowering time traits to untangle complex SAS networks. We used over-representation analysis (ORA) of shade-responsive genes, combined with previous annotation, to logically select 59 known and candidate novel mutants for phenotyping. Our analysis reveals shared and separate pathways for each shade avoidance response. In particular, auxin pathway components were required for shade avoidance responses in hypocotyl, petiole, and flowering time, whereas jasmonic acid pathway components were only required for petiole and flowering time responses. Our phenotypic profiling allowed discovery of seventeen novel shade avoidance mutants. Our results demonstrate that logical selection of mutants increased success of phenotypic profiling to dissect complex traits and discover novel components. PMID:25874869

  7. Aberrant levels of Wnt/β-catenin pathway components in a rat model of endometriosis.

    PubMed

    de Mattos, Rômulo Medina; Pereira, Paula Rodrigues; Barros, Eliane Gouvêa de Oliveira; da Silva, Julianna Henriques; Palmero, Celia Yelimar; da Costa, Nathália Meireles; Pinto, Luis Felipe Ribeiro; Gimba, Etel Rodrigues Pereira; Hecht, Fabio; Ferreira, Luciana Bueno; Machado, Daniel Escorsim; de Oliveira, Felipe Leite; Nasciutti, Luiz Eurico

    2016-08-01

    Endometriosis is a benign gynecological disease affecting approximately 10-15% of women of reproductive age and 25-50% of all infertile women. It is characterized by the presence of glands and/or endometrial stroma outside the uterine cavity. Angiogenesis is a crucial process for the development and maintenance of endometriotic lesions. The Wnt/β-catenin pathway is a major promoter of angiogenesis in both physiological and pathological conditions. In the present study, we evaluated the expression of molecules related to the Wnt/β-catenin pathway in a rat model of peritoneal endometriosis. mRNA analyses showed significantly increased expression of Wnt4 and Wnt7b and decreased expression of Gsk3beta and E-cadherin in endometriotic lesions. However, there were no differences in β-catenin and Fzd2 mRNA expression. In addition, we observed a significant increase of nuclear β-catenin in endometriotic lesions, a hallmark of Wnt/ β-catenin pathway activation. Stromal β-catenin staining was found in 45.4% of endometrial tissues and 77.8% of endometriotic lesions. β-catenin nuclear localization was found in 18.2% of the endometrial tissues and 33.3% of endometriotic lesions. Finally, the expression of galectin-3, a regulator of this pathway, was increased in endometriosis. In summary, this pattern of Wnt/β-catenin components expression suggests an increased activity of this pathway in endometriosis. PMID:26853489

  8. Expression analysis of eight amphioxus genes involved in the Wnt/β-catenin signaling pathway

    PubMed Central

    WANG, Jing; LI, Guang; QIAN, Guang-Hui; HUA, Jun-Hao; WANG, Yi-Quan

    2016-01-01

    The Wnt/β-catenin signaling pathway plays a crucial role in the embryonic development of metazoans. Although the pathway has been studied extensively in many model animals, its function in amphioxus, the most primitive chordate, remains largely uncharacterized. To obtain basic data for functional analysis, we identified and isolated seven genes (Lrp5/6, Dvl, APC, CkIα, CkIδ, Gsk3β, and Gro) of the Wnt/β-catenin signaling pathway from the amphioxus (Branchiostoma floridae) genome. Phylogenetic analysis revealed that amphioxus had fewer members of each gene family than that found in vertebrates. Whole-mount in situ hybridization showed that the genes were maternally expressed and broadly distributed throughout the whole embryo at the cleavage and blastula stages. Among them, Dvl was expressed asymmetrically towards the animal pole, while the others were evenly distributed in all blastomeres. At the mid-gastrula stage, the genes were specifically expressed in the primitive endomesoderm, but displayed different patterns. When the embryo developed into the neurula stage, the gene expressions were mainly detected in either paraxial somites or the tail bud. With the development of the embryo, the expression levels further decreased gradually and remained only in some pharyngeal regions or the tail bud at the larva stage. Our results suggest that the Wnt/β-catenin pathway might be involved in amphioxus somite formation and posterior growth, but not in endomesoderm specification. PMID:27265651

  9. Prediction of Metabolic Pathway Involvement in Prokaryotic UniProtKB Data by Association Rule Mining

    PubMed Central

    Hoehndorf, Robert; Martin, Maria J.; Solovyev, Victor

    2016-01-01

    The widening gap between known proteins and their functions has encouraged the development of methods to automatically infer annotations. Automatic functional annotation of proteins is expected to meet the conflicting requirements of maximizing annotation coverage, while minimizing erroneous functional assignments. This trade-off imposes a great challenge in designing intelligent systems to tackle the problem of automatic protein annotation. In this work, we present a system that utilizes rule mining techniques to predict metabolic pathways in prokaryotes. The resulting knowledge represents predictive models that assign pathway involvement to UniProtKB entries. We carried out an evaluation study of our system performance using cross-validation technique. We found that it achieved very promising results in pathway identification with an F1-measure of 0.982 and an AUC of 0.987. Our prediction models were then successfully applied to 6.2 million UniProtKB/TrEMBL reference proteome entries of prokaryotes. As a result, 663,724 entries were covered, where 436,510 of them lacked any previous pathway annotations. PMID:27390860

  10. Expression analysis of eight amphioxus genes involved in the Wnt/β-catenin signaling pathway.

    PubMed

    Wang, Jing; Li, Guang; Qian, Guang-Hui; Hua, Jun-Hao; Wang, Yi-Quan

    2016-05-18

    The Wnt/β-catenin signaling pathway plays a crucial role in the embryonic development of metazoans. Although the pathway has been studied extensively in many model animals, its function in amphioxus, the most primitive chordate, remains largely uncharacterized. To obtain basic data for functional analysis, we identified and isolated seven genes (Lrp5/6, Dvl, APC, CkIα, CkIδ, Gsk3β, and Gro) of the Wnt/β-catenin signaling pathway from the amphioxus (Branchiostoma floridae) genome. Phylogenetic analysis revealed that amphioxus had fewer members of each gene family than that found in vertebrates. Whole-mount in situ hybridization showed that the genes were maternally expressed and broadly distributed throughout the whole embryo at the cleavage and blastula stages. Among them, Dvl was expressed asymmetrically towards the animal pole, while the others were evenly distributed in all blastomeres. At the mid-gastrula stage, the genes were specifically expressed in the primitive endomesoderm, but displayed different patterns. When the embryo developed into the neurula stage, the gene expressions were mainly detected in either paraxial somites or the tail bud. With the development of the embryo, the expression levels further decreased gradually and remained only in some pharyngeal regions or the tail bud at the larva stage. Our results suggest that the Wnt/β-catenin pathway might be involved in amphioxus somite formation and posterior growth, but not in endomesoderm specification. PMID:27265651

  11. Modelling and pathway identification involving the transport mechanism of a complex metabolic system in batch culture

    NASA Astrophysics Data System (ADS)

    Yuan, Jinlong; Zhang, Xu; Zhu, Xi; Feng, Enmin; Yin, Hongchao; Xiu, Zhilong

    2014-06-01

    The bio-dissimilation of glycerol to 1,3-propanediol (1,3-PD) by Klebsiella pneumoniae (K. pneumoniae) can be characterized by a complex metabolic system of interactions among biochemical fluxes, metabolic compounds, key enzymes and genetic regulation. In this paper, in consideration of the fact that the transport ways of 1,3-PD and glycerol with different weights across cell membrane are still unclear in batch culture, we consider 121 possible metabolic pathways and establish a novel mathematical model which is represented by a complex metabolic system. Taking into account the difficulty in accurately measuring the concentration of intracellular substances and the absence of equilibrium point for the metabolic system of batch culture, the novel approach used here is to define quantitatively biological robustness of the intracellular substance concentrations for the overall process of batch culture. To determine the most possible metabolic pathway, we take the defined biological robustness as cost function and establish an identification model, in which 1452 system parameters and 484 pathway parameters are involved. Simultaneously, the identification model is subject to the metabolic system, continuous state constraints and parameter constraints. As such, solving the identification model by a serial program is a very complicated task. We propose a parallel migration particle swarm optimization algorithm (MPSO) capable of solving the identification model in conjunction with the constraint transcription and smoothing approximation techniques. Numerical results show that the most possible metabolic pathway and the corresponding metabolic system can reasonably describe the process of batch culture.

  12. Involvement of PDK1, PKC and TOR signaling pathways in basal fluconazole tolerance in Cryptococcus neoformans

    PubMed Central

    Lee, Hyeseung; Lamichhane, Ami Khanal; Garraffo, H. Martin; Kwon-Chung, Kyung J.; Chang, Yun C.

    2012-01-01

    Summary This study shows the importance of PDK1, TOR and PKC signaling pathways to the basal tolerance of Cryptococcus neoformans toward fluconazole, the widely used drug for treatment of cryptococcosis. Mutations in genes integral to these pathway resulted in hypersensitivity to the drug. Upon fluconazole treatment, Mpk1, the downstream target of PKC was phosphorylated and its phosphorylation required Pdk1. We show genetically that the PDK1 and TOR phosphorylation sites in Ypk1 as well as the kinase activity of Ypk1 are required for the fluconazole basal tolerance. The involvement of these pathways in fluconazole basal tolerance was associated with sphingolipid homeostasis. Deletion of PDK1, SIN1, or YPK1 but not MPK1 affected cell viability in the presence of sphingolipid biosynthesis inhibitors. Concurrently, pdk1Δ, sinΔ1, ypk1Δ, and mpk1Δ exhibited altered sphingolipid content and elevated fluconazole accumulation compared with the wild-type. The fluconazole hypersensitivity phenotype of these mutants, therefore, appears to be the result of malfunction of the influx/efflux systems due to modifications of membrane sphingolipid content. Interestingly, the reduced virulence of these strains in mice suggests that the cryptococcal PDK1, PKC, and likely the TOR pathways play an important role in managing stress exerted either by fluconazole or by the host environment. PMID:22339665

  13. Pathways and genes involved in steroid hormone metabolism in male pigs: a review and update.

    PubMed

    Robic, Annie; Faraut, Thomas; Prunier, Armelle

    2014-03-01

    This paper reviews state-of-the-art knowledge on steroid biosynthesis pathways in the pig and provides an updated characterization of the porcine genes involved in these pathways with particular focus on androgens, estrogens, and 16-androstenes. At least 21 different enzymes appear to be involved in these pathways in porcine tissues together with at least five cofactors. Until now, data on several porcine genes were scarce or confusing. We characterized the complete genomic and transcript sequences of the single porcine CYP11B gene. We analyzed the porcine AKR1 gene cluster and identified four AKR1C, one AKR1C like genes and one AKR1E2 gene. We provide evidence that porcine AKR1C genes are not orthologous to human AKR1C. A new nomenclature is thus needed for this gene family in the pig. Thirty-two genes are now described: transcript (30+2 characterized in this study) and genomic (complete: 18+1 and partial: 12+1) sequences are identified. However, despite increasing knowledge on steroid metabolism in the pig, there is still no explanation of why porcine testes can produce androstenone and epiandrosterone, but not dihydrotestosterone (DHT), which is also a reduced steroid. PMID:24239507

  14. On the possible involvement of bovine serum albumin precursor in lipofection pathway.

    PubMed

    Mukherjee, Anubhab; Bhattacharyya, Jayanta; Chaudhuri, Arabinda

    2014-03-01

    Protein factors involved in lipofection pathways remain elusive. Using avidin-biotin affinity chromatography and mass finger printing analysis technique, herein we report the identification of a 70 kDa size protein (bovine serum albumin precursor, BSAP) which binds strongly with lipoplexes and may play role in lipofection pathway. Using multiple cultured animal cells and three structurally different cationic transfection lipids, we show that the efficiencies of liposomal transfection vectors get significantly enhanced (by ~2.5- to 5.0-fold) in cells pre-transfected with lipoplexes of reporter plasmid construct encoding BSAP. Findings in the cellular uptake experiments in A549 cells cultured in DMEM supplemented with 10 percent (w/w) BODIPY-labelled BSAP are consistent with the supposition that BSAP enters cell cytoplasm from the cell culture medium (DMEM supplemented with 10 percent FBS) used in lipofection. Cellular uptake studies by confocal microscopy using BODIPY-labelled BSAP and FITC-labelled plasmid DNA revealed co-localization of plasmid DNA and BSAP within the cell cytoplasm and nucleus. In summary, the present findings hint at the possible involvement of BSAP in lipofection pathway. PMID:24499789

  15. Tributyltin induces apoptotic signaling in hepatocytes through pathways involving the endoplasmic reticulum and mitochondria

    SciTech Connect

    Grondin, Melanie; Marion, Michel; Denizeau, Francine; Averill-Bates, Diana A. . E-mail: averill.diana@uqam.ca

    2007-07-01

    Tri-n-butyltin is a widespread environmental toxicant, which accumulates in the liver. This study investigates whether tri-n-butyltin induces pro-apoptotic signaling in rat liver hepatocytes through pathways involving the endoplasmic reticulum and mitochondria. Tri-n-butyltin activated the endoplasmic reticulum pathway of apoptosis, which was demonstrated by the activation of the protease calpain, its translocation to the plasma membrane, followed by cleavage of the calpain substrates, cytoskeletal protein vinculin, and caspase-12. Caspase-12 is localized to the cytoplasmic side of the endoplasmic reticulum and is involved in apoptosis mediated by the endoplasmic reticulum. Tri-n-butyltin also caused translocation of the pro-apoptotic proteins Bax and Bad from the cytosol to mitochondria, as well as changes in mitochondrial membrane permeability, events which can activate the mitochondrial death pathway. Tri-n-butyltin induced downstream apoptotic events in rat hepatocytes at the nuclear level, detected by chromatin condensation and by confocal microscopy using acridine orange. We investigated whether the tri-n-butyltin-induced pro-apoptotic events in hepatocytes could be linked to perturbation of intracellular calcium homeostasis, using confocal microscopy. Tri-n-butyltin caused changes in intracellular calcium distribution, which were similar to those induced by thapsigargin. Calcium was released from a subcellular compartment, which is likely to be the endoplasmic reticulum, into the cytosol. Cytosolic acidification, which is known to trigger apoptosis, also occurred and involved the Cl{sup -}/HCO{sub 3} {sup -} exchanger. Pro-apoptotic events in hepatocytes were inhibited by the calcium chelator, Bapta-AM, and by a calpain inhibitor, which suggests that changes in intracellular calcium homeostasis are involved in tri-n-butyltin-induced apoptotic signaling in rat hepatocytes.

  16. The Rtr1p CTD phosphatase autoregulates its mRNA through a degradation pathway involving the REX exonucleases

    PubMed Central

    Hodko, Domagoj; Ward, Taylor; Chanfreau, Guillaume

    2016-01-01

    Rtr1p is a phosphatase that impacts gene expression by modulating the phosphorylation status of the C-terminal domain of the large subunit of RNA polymerase II. Here, we show that Rtr1p is a component of a novel mRNA degradation pathway that promotes its autoregulation through turnover of its own mRNA. We show that the 3′UTR of the RTR1 mRNA contains a cis element that destabilizes this mRNA. RTR1 mRNA turnover is achieved through binding of Rtr1p to the RTR1 mRNP in a manner that is dependent on this cis element. Genetic evidence shows that Rtr1p-mediated decay of the RTR1 mRNA involves the 5′-3′ DExD/H-box RNA helicase Dhh1p and the 3′-5′ exonucleases Rex2p and Rex3p. Rtr1p and Rex3p are found associated with Dhh1p, suggesting a model for recruiting the REX exonucleases to the RTR1 mRNA for degradation. Rtr1p-mediated decay potentially impacts additional transcripts, including the unspliced BMH2 pre-mRNA. We propose that Rtr1p may imprint its RNA targets cotranscriptionally and determine their downstream degradation mechanism by directing these transcripts to a novel turnover pathway that involves Rtr1p, Dhh1p, and the REX family of exonucleases. PMID:26843527

  17. Prolonged sleep restriction induces changes in pathways involved in cholesterol metabolism and inflammatory responses.

    PubMed

    Aho, Vilma; Ollila, Hanna M; Kronholm, Erkki; Bondia-Pons, Isabel; Soininen, Pasi; Kangas, Antti J; Hilvo, Mika; Seppälä, Ilkka; Kettunen, Johannes; Oikonen, Mervi; Raitoharju, Emma; Hyötyläinen, Tuulia; Kähönen, Mika; Viikari, Jorma S A; Härmä, Mikko; Sallinen, Mikael; Olkkonen, Vesa M; Alenius, Harri; Jauhiainen, Matti; Paunio, Tiina; Lehtimäki, Terho; Salomaa, Veikko; Orešič, Matej; Raitakari, Olli T; Ala-Korpela, Mika; Porkka-Heiskanen, Tarja

    2016-01-01

    Sleep loss and insufficient sleep are risk factors for cardiometabolic diseases, but data on how insufficient sleep contributes to these diseases are scarce. These questions were addressed using two approaches: an experimental, partial sleep restriction study (14 cases and 7 control subjects) with objective verification of sleep amount, and two independent epidemiological cohorts (altogether 2739 individuals) with questions of sleep insufficiency. In both approaches, blood transcriptome and serum metabolome were analysed. Sleep loss decreased the expression of genes encoding cholesterol transporters and increased expression in pathways involved in inflammatory responses in both paradigms. Metabolomic analyses revealed lower circulating large HDL in the population cohorts among subjects reporting insufficient sleep, while circulating LDL decreased in the experimental sleep restriction study. These findings suggest that prolonged sleep deprivation modifies inflammatory and cholesterol pathways at the level of gene expression and serum lipoproteins, inducing changes toward potentially higher risk for cardiometabolic diseases. PMID:27102866

  18. Prolonged sleep restriction induces changes in pathways involved in cholesterol metabolism and inflammatory responses

    PubMed Central

    Aho, Vilma; Ollila, Hanna M.; Kronholm, Erkki; Bondia-Pons, Isabel; Soininen, Pasi; Kangas, Antti J.; Hilvo, Mika; Seppälä, Ilkka; Kettunen, Johannes; Oikonen, Mervi; Raitoharju, Emma; Hyötyläinen, Tuulia; Kähönen, Mika; Viikari, Jorma S.A.; Härmä, Mikko; Sallinen, Mikael; Olkkonen, Vesa M.; Alenius, Harri; Jauhiainen, Matti; Paunio, Tiina; Lehtimäki, Terho; Salomaa, Veikko; Orešič, Matej; Raitakari, Olli T.; Ala-Korpela, Mika; Porkka-Heiskanen, Tarja

    2016-01-01

    Sleep loss and insufficient sleep are risk factors for cardiometabolic diseases, but data on how insufficient sleep contributes to these diseases are scarce. These questions were addressed using two approaches: an experimental, partial sleep restriction study (14 cases and 7 control subjects) with objective verification of sleep amount, and two independent epidemiological cohorts (altogether 2739 individuals) with questions of sleep insufficiency. In both approaches, blood transcriptome and serum metabolome were analysed. Sleep loss decreased the expression of genes encoding cholesterol transporters and increased expression in pathways involved in inflammatory responses in both paradigms. Metabolomic analyses revealed lower circulating large HDL in the population cohorts among subjects reporting insufficient sleep, while circulating LDL decreased in the experimental sleep restriction study. These findings suggest that prolonged sleep deprivation modifies inflammatory and cholesterol pathways at the level of gene expression and serum lipoproteins, inducing changes toward potentially higher risk for cardiometabolic diseases. PMID:27102866

  19. Nitrogen modulation of legume root architecture signaling pathways involves phytohormones and small regulatory molecules

    PubMed Central

    Mohd-Radzman, Nadiatul A.; Djordjevic, Michael A.; Imin, Nijat

    2013-01-01

    Nitrogen, particularly nitrate is an important yield determinant for crops. However, current agricultural practice with excessive fertilizer usage has detrimental effects on the environment. Therefore, legumes have been suggested as a sustainable alternative for replenishing soil nitrogen. Legumes can uniquely form nitrogen-fixing nodules through symbiotic interaction with specialized soil bacteria. Legumes possess a highly plastic root system which modulates its architecture according to the nitrogen availability in the soil. Understanding how legumes regulate root development in response to nitrogen availability is an important step to improving root architecture. The nitrogen-mediated root development pathway starts with sensing soil nitrogen level followed by subsequent signal transduction pathways involving phytohormones, microRNAs and regulatory peptides that collectively modulate the growth and shape of the root system. This review focuses on the current understanding of nitrogen-mediated legume root architecture including local and systemic regulations by different N-sources and the modulations by phytohormones and small regulatory molecules. PMID:24098303

  20. Identification of Sleep-Modulated Pathways Involved in Neuroprotection from Stroke

    PubMed Central

    Pace, Marta; Baracchi, Francesca; Gao, Bo; Bassetti, Claudio

    2015-01-01

    Study Objectives: Sleep deprivation (SDp) performed before stroke induces an ischemic tolerance state as observed in other forms of preconditioning. As the mechanisms underlying this effect are not well understood, we used DNA oligonucleotide microarray analysis to identify the genes and the gene-pathways underlying SDp preconditioning effects. Design: Gene expression was analyzed 3 days after stroke in 4 experimental groups: (i) SDp performed before focal cerebral ischemia (IS) induction; (ii) SDp performed before sham surgery; (iii) IS without SDp; and (iv) sham surgery without SDp. SDp was performed by gentle handling during the last 6 h of the light period, and ischemia was induced immediately after. Settings: Basic sleep research laboratory. Measurements and Results: Stroke induced a massive alteration in gene expression both in sleep deprived and non-sleep deprived animals. However, compared to animals that underwent ischemia alone, SDp induced a general reduction in transcriptional changes with a reduction in the upregulation of genes involved in cell cycle regulation and immune response. Moreover, an upregulation of a new neuroendocrine pathway which included melanin concentrating hormone, glycoprotein hormones-α-polypeptide and hypocretin was observed exclusively in rats sleep deprived before stroke. Conclusion: Our data indicate that sleep deprivation before stroke reprogrammed the signaling response to injury. The inhibition of cell cycle regulation and inflammation are neuroprotective mechanisms reported also for other forms of preconditioning treatment, whereas the implication of the neuroendocrine function is novel and has never been described before. These results therefore provide new insights into neuroprotective mechanisms involved in ischemic tolerance mechanisms. Citation: Pace M, Baracchi F, Gao B, Bassetti C. Identification of sleep-modulated pathways involved in neuroprotection from stroke. SLEEP 2015;38(11):1707–1718. PMID:26085290

  1. Core regulatory components of the PHO pathway are conserved in the methylotrophic yeast Hansenula polymorpha.

    PubMed

    Zhou, Ying; Yuikawa, Naoya; Nakatsuka, Hiroki; Maekawa, Hiromi; Harashima, Satoshi; Nakanishi, Yoichi; Kaneko, Yoshinobu

    2016-08-01

    To gain better understanding of the diversity and evolution of the gene regulation system in eukaryotes, the phosphate signal transduction (PHO) pathway in non-conventional yeasts has been studied in recent years. Here we characterized the PHO pathway of Hansenula polymorpha, which is genetically tractable and distantly related to Saccharomyces cerevisiae and Schizosaccharomyces pombe, in order to get more information for the diversity and evolution of the PHO pathway in yeasts. We generated several pho gene-deficient mutants based on the annotated draft genome of H. polymorpha BY4329. Except for the Hppho2-deficient mutant, these mutants exhibited the same phenotype of repressible acid phosphatase (APase) production as their S. cerevisiae counterparts. Subsequently, Hppho80 and Hppho85 mutants were isolated as suppressors of the Hppho81 mutation and Hppho4 was isolated from Hppho80 and Hppho85 mutants as the sole suppressor of the Hppho80 and Hppho85 mutations. To gain more complete delineation of the PHO pathway in H. polymorpha, we screened for UV-irradiated mutants that expressed APase constitutively. As a result, three classes of recessive constitutive mutations and one dominant constitutive mutation were isolated. Genetic analysis showed that one group of recessive constitutive mutations was allelic to HpPHO80 and that the dominant mutation occurred in the HpPHO81 gene. Epistasis analysis between Hppho81 and the other two classes of recessive constitutive mutations suggested that the corresponding new genes, named PHO51 and PHO53, function upstream of HpPHO81 in the PHO pathway. Taking these findings together, we conclude that the main components of the PHO pathway identified in S. cerevisiae are conserved in the methylotrophic yeast H. polymorpha, even though these organisms separated from each other before duplication of the whole genome. This finding is useful information for the study of evolution of the PHO regulatory system in yeasts. PMID:26794724

  2. Dissection of the bifunctional ARGRII protein involved in the regulation of arginine anabolic and catabolic pathways.

    PubMed Central

    Qui, H F; Dubois, E; Messenguy, F

    1991-01-01

    ARGRII is a regulatory protein which regulates the arginine anabolic and catabolic pathways in combination with ARGRI and ARGRIII. We have investigated, by deletion analysis and fusion to LexA protein, the different domains of ARGRII protein. In contrast to other yeast regulatory proteins, 92% of ARGRII is necessary for its anabolic repression function and 80% is necessary for its catabolic activator function. We can define three domains in this protein: a putative DNA-binding domain containing a zinc finger motif, a region more involved in the repression activity located around the RNase-like sequence, and a large activation domain. Images PMID:2005903

  3. A Novel Component of the Disulfide-Reducing Pathway Required for Cytochrome c Assembly in Plastids

    PubMed Central

    Gabilly, Stéphane T.; Kropat, Janette; Karamoko, Mohamed; Page, M. Dudley; Nakamoto, Stacie S.; Merchant, Sabeeha S.; Hamel, Patrice P.

    2011-01-01

    In plastids, the conversion of energy in the form of light to ATP requires key electron shuttles, the c-type cytochromes, which are defined by the covalent attachment of heme to a CXXCH motif. Plastid c-type cytochrome biogenesis occurs in the thylakoid lumen and requires a system for transmembrane transfer of reductants. Previously, CCDA and CCS5/HCF164, found in all plastid-containing organisms, have been proposed as two components of the disulfide-reducing pathway. In this work, we identify a small novel protein, CCS4, as a third component in this pathway. CCS4 was genetically identified in the green alga Chlamydomonas reinhardtii on the basis of the rescue of the ccs4 mutant, which is blocked in the synthesis of holoforms of plastid c-type cytochromes, namely cytochromes f and c6. Although CCS4 does not display sequence motifs suggestive of redox or heme-binding function, biochemical and genetic complementation experiments suggest a role in the disulfide-reducing pathway required for heme attachment to apoforms of cytochromes c. Exogenous thiols partially rescue the growth phenotype of the ccs4 mutant concomitant with recovery of holocytochrome f accumulation, as does expression of an ectopic copy of the CCDA gene, encoding a trans-thylakoid transporter of reducing equivalents. We suggest that CCS4 might function to stabilize CCDA or regulate its activity. PMID:21220358

  4. Phosphate-responsive signaling pathway is a novel component of NAD+ metabolism in Saccharomyces cerevisiae.

    PubMed

    Lu, Shu-Ping; Lin, Su-Ju

    2011-04-22

    Nicotinamide adenine dinucleotide (NAD(+)) is an essential cofactor involved in various cellular biochemical reactions. To date the signaling pathways that regulate NAD(+) metabolism remain unclear due to the dynamic nature and complexity of the NAD(+) metabolic pathways and the difficulty of determining the levels of the interconvertible pyridine nucleotides. Nicotinamide riboside (NmR) is a key pyridine metabolite that is excreted and re-assimilated by yeast and plays important roles in the maintenance of NAD(+) pool. In this study we establish a NmR-specific reporter system and use it to identify yeast mutants with altered NmR/NAD(+) metabolism. We show that the phosphate-responsive signaling (PHO) pathway contributes to control NAD(+) metabolism. Yeast strains with activated PHO pathway show increases in both the release rate and internal concentration of NmR. We further identify Pho8, a PHO-regulated vacuolar phosphatase, as a potential NmR production factor. We also demonstrate that Fun26, a homolog of human ENT (equilibrative nucleoside transporter), localizes to the vacuolar membrane and establishes the size of the vacuolar and cytosolic NmR pools. In addition, the PHO pathway responds to depletion of cellular nicotinic acid mononucleotide (NaMN) and mediates nicotinamide mononucleotide (NMN) catabolism, thereby contributing to both NmR salvage and phosphate acquisition. Therefore, NaMN is a putative molecular link connecting the PHO signaling and NAD(+) metabolic pathways. Our findings may contribute to the understanding of the molecular basis and regulation of NAD(+) metabolism in higher eukaryotes. PMID:21349851

  5. Rabies virus-induced apoptosis involves caspase-dependent and caspase-independent pathways.

    PubMed

    Sarmento, Luciana; Tseggai, Tesfai; Dhingra, Vikas; Fu, Zhen F

    2006-11-01

    Previously, it has been shown that the laboratory attenuated rabies virus CVS-B2C, but not the wild-type virus SHBRV, induces apoptosis in mice and the induction of apoptosis is mediated by viral glycoprotein. Induction of apoptosis by CVS-B2C limits the spread of the virus in the CNS. In the present study, we characterized the pathways by which CVS-B2C induces apoptosis. BSR cells were infected with CVS-B2C or SHBRV and harvested at different time points for detection of apoptosis by immunofluorescence and flow cytometry. Apoptosis was detected only in cells infected with CVS-B2C, but not SHBRV. Caspase activity and expression of several apoptotic proteins were analyzed by fluorometric assay and Western blotting. Activation of caspase-8 and -3, but not of caspase-9, was observed in CVS-B2C-infected cells. In addition, the level of expression of Apaf-1 did not change. Furthermore, PARP was cleaved confirming activation of downstream caspases. All these data suggest that CVS-B2C infection activates the extrinsic, but not the intrinsic, apoptotic pathway. In addition, AIF, a caspase-independent apoptotic protein was up-regulated and translocated from the cytoplasm to the nucleus post-infection, suggesting that apoptosis induced by CVS-B2C also involves the activation of a caspase-independent pathway. PMID:16814422

  6. Intracellular Signaling Pathways Involved in Childhood Acute Lymphoblastic Leukemia; Molecular Targets.

    PubMed

    Layton Tovar, Cristian Fabián; Mendieta Zerón, Hugo

    2016-06-01

    Acute lymphoblastic leukemia (ALL) is a malignant disease characterized by an uncontrolled proliferation of immature lymphoid cells. ALL is the most common hematologic malignancy in early childhood, and it reaches peak incidence between the ages of 2 and 3 years. The prognosis of ALL is associated with aberrant gene expression, in addition to the presence of numerical or structural chromosomal alterations, age, race, and immunophenotype. The Relapse rate with regard to pharmacological treatment rises in childhood; thus, the expression of biomarkers associated with the activation of cell signaling pathways is crucial to establish the disease prognosis. Intracellular pathways involved in ALL are diverse, including Janus kinase/Signal transducers and transcription activators (JAK-STAT), Phosphoinositide-3-kinase-protein kinase B (PI3K-AKT), Ras mitogen-activated protein kinase (Ras-MAPK), Glycogen synthase kinase-3β (GSK-3β), Nuclear factor-kappa beta (NF-κB), and Hypoxia-inducible transcription factor 1α (HIF-1α), among others. In this review, we present several therapeutic targets, intracellular pathways, and molecular markers that are being studied extensively at present. PMID:27065575

  7. Characterization of Differentially Expressed Genes Involved in Pathways Associated with Gastric Cancer

    PubMed Central

    Li, Hao; Yu, Beiqin; Li, Jianfang; Su, Liping; Yan, Min; Zhang, Jun; Li, Chen; Zhu, Zhenggang; Liu, Bingya

    2015-01-01

    To explore the patterns of gene expression in gastric cancer, a total of 26 paired gastric cancer and noncancerous tissues from patients were enrolled for gene expression microarray analyses. Limma methods were applied to analyze the data, and genes were considered to be significantly differentially expressed if the False Discovery Rate (FDR) value was < 0.01, P-value was <0.01 and the fold change (FC) was >2. Subsequently, Gene Ontology (GO) categories were used to analyze the main functions of the differentially expressed genes. According to the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, we found pathways significantly associated with the differential genes. Gene-Act network and co-expression network were built respectively based on the relationships among the genes, proteins and compounds in the database. 2371 mRNAs and 350 lncRNAs considered as significantly differentially expressed genes were selected for the further analysis. The GO categories, pathway analyses and the Gene-Act network showed a consistent result that up-regulated genes were responsible for tumorigenesis, migration, angiogenesis and microenvironment formation, while down-regulated genes were involved in metabolism. These results of this study provide some novel findings on coding RNAs, lncRNAs, pathways and the co-expression network in gastric cancer which will be useful to guide further investigation and target therapy for this disease. PMID:25928635

  8. Directing folding pathways for multi-component DNA origami nanostructures with complex topology

    NASA Astrophysics Data System (ADS)

    Marras, A. E.; Zhou, L.; Kolliopoulos, V.; Su, H.-J.; Castro, C. E.

    2016-05-01

    Molecular self-assembly has become a well-established technique to design complex nanostructures and hierarchical mesoscale assemblies. The typical approach is to design binding complementarity into nucleotide or amino acid sequences to achieve the desired final geometry. However, with an increasing interest in dynamic nanodevices, the need to design structures with motion has necessitated the development of multi-component structures. While this has been achieved through hierarchical assembly of similar structural units, here we focus on the assembly of topologically complex structures, specifically with concentric components, where post-folding assembly is not feasible. We exploit the ability to direct folding pathways to program the sequence of assembly and present a novel approach of designing the strand topology of intermediate folding states to program the topology of the final structure, in this case a DNA origami slider structure that functions much like a piston-cylinder assembly in an engine. The ability to program the sequence and control orientation and topology of multi-component DNA origami nanostructures provides a foundation for a new class of structures with internal and external moving parts and complex scaffold topology. Furthermore, this work provides critical insight to guide the design of intermediate states along a DNA origami folding pathway and to further understand the details of DNA origami self-assembly to more broadly control folding states and landscapes.

  9. Intracellular signaling pathways involved in the relaxin-induced proliferation of rat Sertoli cells.

    PubMed

    Nascimento, Aline Rosa; Pimenta, Maristela Taliari; Lucas, Thais F G; Royer, Carine; Porto, Catarina Segreti; Lazari, Maria Fatima Magalhaes

    2012-09-15

    Regulation of Sertoli cell number is a key event to determine normal spermatogenesis. We have previously shown that relaxin and its G-protein coupled receptor RXFP1 are expressed in rat Sertoli cells, and that relaxin stimulates Sertoli cell proliferation. This study examined the mechanisms underlying the mitogenic effect of relaxin in a primary culture of Sertoli cells removed from testes of immature rats. Stimulation with exogenous relaxin increased Sertoli cell number and the expression of the proliferating cell nuclear antigen (PCNA), but did not affect the mRNA level of the differentiation markers cadherins 1 and 2. Relaxin-induced Sertoli cell proliferation was blocked by inhibition of MEK/ERK1/2 or PI3K/AKT pathways, but not by inhibition of PKC or EGFR activity. Relaxin induced a rapid and transient activation of ERK1/2 phosphorylation, which was MEK and SRC-dependent, and involved upstream activation of G(i). AKT activation could be detected 5 min after relaxin stimulation, and was still detected after 24h of stimulation with relaxin. Relaxin-induced AKT phosphorylation was G(i)- but not PKA-dependent, and it was blocked by both PI3K and MEK inhibitors. In conclusion, the mitogenic effect of relaxin in Sertoli cell involves coupling to G(i) and activation of both MEK/ERK1/2 and PI3K/AKT pathways. PMID:22819701

  10. Genes and Pathways Involved in Adult Onset Disorders Featuring Muscle Mitochondrial DNA Instability

    PubMed Central

    Ahmed, Naghia; Ronchi, Dario; Comi, Giacomo Pietro

    2015-01-01

    Replication and maintenance of mtDNA entirely relies on a set of proteins encoded by the nuclear genome, which include members of the core replicative machinery, proteins involved in the homeostasis of mitochondrial dNTPs pools or deputed to the control of mitochondrial dynamics and morphology. Mutations in their coding genes have been observed in familial and sporadic forms of pediatric and adult-onset clinical phenotypes featuring mtDNA instability. The list of defects involved in these disorders has recently expanded, including mutations in the exo-/endo-nuclease flap-processing proteins MGME1 and DNA2, supporting the notion that an enzymatic DNA repair system actively takes place in mitochondria. The results obtained in the last few years acknowledge the contribution of next-generation sequencing methods in the identification of new disease loci in small groups of patients and even single probands. Although heterogeneous, these genes can be conveniently classified according to the pathway to which they belong. The definition of the molecular and biochemical features of these pathways might be helpful for fundamental knowledge of these disorders, to accelerate genetic diagnosis of patients and the development of rational therapies. In this review, we discuss the molecular findings disclosed in adult patients with muscle pathology hallmarked by mtDNA instability. PMID:26251896

  11. Implications of the involvement of the endoplasmic reticulum stress pathway in drug-induced apoptosis.

    PubMed

    Cory, Ann H; Chen, Jianming; Cory, Joseph G

    2008-01-01

    Apoptosis occurs by distinct pathways that involve the cell surface, mitochondria or the endoplasmic reticulum. Previous studies had shown that deoxyadenosine-resistant L1210 cells (Y8) proceeded to apoptosis under conditions in which the parental L1210 cell line (WT) did not undergo an apoptotic response. Combinations of drugs, acting at different molecular targets, markedly potentiated the apoptotic response in the Y8 cells without inducing apoptosis in the WT cells. In the present study, induction of apoptosis by parthenolide and BAY 11-7085, drugs that targeted nuclear factor kappa B activation, was blocked by the presence of N-acetylcysteine (NAC). On the other hand, the levels of apoptosis induced by parthenolide or BAY 11-7085 were increased by pre-treatment of the cells with glutathione lowering L-buthionine-(S,R)-sulfoximine (BSO). Western blot analyses showed that the levels of the stress proteins, Grp 78 and Gadd 153 were reduced in the parthenolide-treated Y8 cells, but not in those co-treated with NAC. Protection of the cells from apoptosis induced by parthenolide or BAY 11-7085 by NAC was relatively specific as the induction of apoptosis in the Y8 cells by MG-132, flavopiridol, Gemcitabine or PRIMA-1 was not decreased by NAC. These data suggest that multiple pathways, one of which is ER-stress induced, may ultimately be involved and interactive in the induction of apoptosis in specific cell lines. PMID:18507007

  12. Musashi2 modulates K562 leukemic cell proliferation and apoptosis involving the MAPK pathway

    SciTech Connect

    Zhang, Huijuan; Tan, Shi; Wang, Juan; Chen, Shana; Quan, Jing; Xian, Jingrong; Zhang, Shuai shuai; He, Jingang; Zhang, Ling

    2014-01-01

    The RNA-binding protein Musashi2 (Msi2) has been identified as a master regulator within a variety of stem cell populations via the regulation of translational gene expression. A recent study has suggested that Msi2 is strongly expressed in leukemic cells of acute myeloid leukemia patients, and elevated Msi2 is associated with poor prognosis. However, the potential role of Msi2 in leukemogenesis is still not well understood. Here, we investigated the effect of Msi2 knockdown on the biological properties of leukemic cells. High expression of Msi2 was found in K562 and KG-1a leukemic cell lines, and low expression was observed in the U937 cell line. We transduced K562 cells with two independent adenoviral shRNA vectors targeting Msi2 and confirmed knockdown of Msi2 at the mRNA and protein levels. Msi2 silencing inhibited cell growth and caused cell cycle arrest by increasing the expression of p21 and decreasing the expression of cyclin D1 and cdk2. In addition, knockdown of Msi2 promoted cellular apoptosis via the upregulation of Bax and downregulation of Bcl-2 expression. Furthermore, Msi2 knockdown resulted in the inactivation of the ERK/MAPK and p38/MAPK pathways, but no remarkable change in p-AKT was observed. These data provide evidence that Msi2 plays an important role in leukemogenesis involving the MAPK signaling pathway, which indicates that Msi2 may be a novel target for leukemia treatment. - Highlights: • Knockdown of Msi2 inhibited K562 cell growth and arrested cell cycle progression. • Knockdown of Msi2 induced K562 cell apoptosis via the regulation of Bax and Bcl-2. • The MAPK pathway was involved in the process of Msi2-mediated leukemogenesis. • Our data indicate that Msi2 is a potential new target for leukemia treatment.

  13. GSK-3/CREB pathway involved in the gx-50's effect on Alzheimer's disease.

    PubMed

    Tang, Maoping; Shi, Shi; Guo, Yubing; Xu, Wangjie; Wang, Lianyun; Chen, Yi; Wang, Zhaoxia; Qiao, Zhongdong

    2014-06-01

    Aggregation of amyloid-beta (Aβ) fragments is one of the major pathological hallmarks of Alzheimer's disease (AD). Our previous study has demonstrated that a novel compound named N-[2-(3, 4-dimethoxyphenyl) ethyl]-3-phenyl-acrylamide (gx-50) can decrease the accumulation of Aβ oligomers in the cerebral cortex and improve the cognitive abilities in transgenic demented mice. To further study the mechanism of the neuroprotective effect of gx-50 against AD, we employed microarray to investigate the gene expression profile of the primary cultured neurons treated with gx-50 or/and Aβ. Microarray disclosed 351 genes associated with AD in the gx-50 plus Aβ treated group, out of the 22,523 probes. 217 of the 351 genes were significantly up-regulated, 134 of them were down-regulated. The 351 genes were mainly involved in neurotransmission, signal transduction, nervous system development, protein phosphorylation, transcription and apoptosis. By the Onto-pathway analysis, a network involved two molecules - GSK-3, CREB and another two closely linked proteins - AKT, BDNF was discovered. The GSK/CREB pathway was further studied at the gene and protein level both in vivo and in vitro. Western blot and immunohistochemistry analysis showed that the gx-50 elevated the AKT phosphorylation and inhibited its downstream protein - GSK-3's activity, then restored the CREB's transcriptional activity, and finally enhanced the expression of the CREB target gene - BDNF. In addition, the real-time PCR results displayed the same tendency. In conclusion, studies in this research indicated that the gx-50 may improve the cognitive ability of AD via the GSK-3/CREB pathway. PMID:24565641

  14. Effect of CCL2 on BV2 microglial cell migration: Involvement of probable signaling pathways.

    PubMed

    Bose, Shambhunath; Kim, Sunyoung; Oh, Yeonsoo; Moniruzzaman, Md; Lee, Gyeongjun; Cho, Jungsook

    2016-05-01

    Microglia, the resident macrophages of the central nervous system, play a vital role in the regulation of innate immune function and neuronal homeostasis of the brain. Currently, much interest is being generated regarding the investigation of the microglial migration that results in their accumulation at focal sites of injury. Chemokines including CCL2 are known to cause the potential induction of migration of microglial cells, although the underlying mechanisms are not well understood. In the present study, using murine neonatal BV2 microglial cells as a model, we investigate the impact of CCL2 on the migration of microglial cells and address the probable molecular events within the cellular signaling cascades mediating CCL2-induced cell migration. Our results demonstrate concentration- and time-dependent induction of BV2 cell migration by CCL2 and reveal complex mechanisms involving the activation of MEK, ERK1/2, and Akt, and their cross-talk. In addition, we demonstrate that the MEK/ERK pathway activated by CCL2 treatment mediate p90RSK activation in BV2 cells. Moreover, our findings indicate that Akt, ERK1/2, and p90RSK are the downstream effectors of PI3K in the CCL2-induced signaling. Finally, phosphorylation of the transcription factors c-jun and ATF-1 is found to be a further downstream signaling cascade in the CCL2-mediated action. Our results suggest that CCL2-induced activation of c-jun and ATF-1 is likely to be linked to the MEK/ERK and PI3K signaling pathways, respectively. Taken together, these findings contribute to a better understanding of CCL2-induced microglial migration and the probable signaling pathways involved. PMID:26878647

  15. Involvement of net and Hif1alpha in distinct yet intricately linked hypoxia-induced signaling pathways.

    PubMed

    Serchov, Tsvetan; Dubois-Pot-Schneider, Helene; Charlot, Celine; Rösl, Frank; Wasylyk, Bohdan

    2010-07-01

    The present study compares negative Ets transcription factor (Net) and hypoxia-inducible factor 1alpha (HIF1alpha) regulation by hypoxia. Their protein stabilities are differently regulated by hypoxia, defining three periods in the kinetics: normoxia (high Net levels and low HIF1alpha levels), early hypoxia (high levels of Net and HIF1alpha), and late hypoxia (degradation of Net and HIF1alpha). Modulators of prolyl hydroxylase domain protein (PHD) activity induce a mobility shift of Net, similar to HIF1alpha, suggesting that post-translational modifications of both factors depend on PHD activity. The three PHDs have different roles in the regulation of Net protein levels; PHD1 and PHD3 are involved in the stabilization of Net, whereas PHD2 controls its degradation in late hypoxia. Net physically interacts with PHD2 in hypoxia, whereas PHD1 and PHD3 bind to Net in normoxia and hypoxia. Under the same conditions, PHD2 and PHD3 regulate both HIF1alpha stabilization in early hypoxia and its degradation at late hypoxia, whereas PHD1 is involved in HIF1alpha degradation in late hypoxia. We describe interconnections between the regulation of both Net and HIF1alpha at the protein level. Evidence is provided for a direct physical interaction between Net and HIF1alpha and indirect transcriptional regulation loops that involve the PHDs. Taken together our results indicate that Net and HIF1alpha are components of distinct signaling pathways that are intricately linked. PMID:20427288

  16. Altered Expression of Wnt Signaling Pathway Components in Osteogenesis of Mesenchymal Stem Cells in Osteoarthritis Patients

    PubMed Central

    Herranz, Eva; Rodríguez-Rodríguez, Luis; Mucientes, Arkaitz; Abásolo, Lydia; Marco, Fernando; Fernández-Gutiérrez, Benjamín; Lamas, José Ramón

    2015-01-01

    Introduction Osteoarthritis (OA) is characterized by altered homeostasis of joint cartilage and bone, whose functional properties rely on chondrocytes and osteoblasts, belonging to mesenchymal stem cells (MSCs). WNT signaling acts as a hub integrating and crosstalking with other signaling pathways leading to the regulation of MSC functions. The aim of this study was to evaluate the existence of a differential signaling between Healthy and OA-MSCs during osteogenesis. Methods MSCs of seven OA patients and six healthy controls were isolated, characterised and expanded. During in vitro osteogenesis, cells were recovered at days 1, 10 and 21. RNA and protein content was obtained. Expression of WNT pathway genes was evaluated using RT-qPCR. Functional studies were also performed to study the MSC osteogenic commitment and functional and post-traslational status of β-catenin and several receptor tyrosine kinases. Results Several genes were downregulated in OA-MSCs during osteogenesis in vitro. These included soluble Wnts, inhibitors, receptors, co-receptors, several kinases and transcription factors. Basal levels of β-catenin were higher in OA-MSCs, but calcium deposition and expression of osteogenic genes was similar between Healthy and OA-MSCs. Interestingly an increased phosphorylation of p44/42 MAPK (ERK1/2) signaling node was present in OA-MSCs. Conclusion Our results point to the existence in OA-MSCs of alterations in expression of Wnt pathway components during in vitro osteogenesis that are partially compensated by post-translational mechanisms modulating the function of other pathways. We also point the relevance of other signaling pathways in OA pathophysiology suggesting their role in the maintenance of joint homeostasis through modulation of MSC osteogenic potential. PMID:26352263

  17. Transcription profiling provides insights into gene pathways involved in horn and scurs development in cattle

    PubMed Central

    2010-01-01

    Background Two types of horns are evident in cattle - fixed horns attached to the skull and a variation called scurs, which refers to small loosely attached horns. Cattle lacking horns are referred to as polled. Although both the Poll and Scurs loci have been mapped to BTA1 and 19 respectively, the underlying genetic basis of these phenotypes is unknown, and so far, no candidate genes regulating these developmental processes have been described. This study is the first reported attempt at transcript profiling to identify genes and pathways contributing to horn and scurs development in Brahman cattle, relative to polled counterparts. Results Expression patterns in polled, horned and scurs tissues were obtained using the Agilent 44 k bovine array. The most notable feature when comparing transcriptional profiles of developing horn tissues against polled was the down regulation of genes coding for elements of the cadherin junction as well as those involved in epidermal development. We hypothesize this as a key event involved in keratinocyte migration and subsequent horn development. In the polled-scurs comparison, the most prevalent differentially expressed transcripts code for genes involved in extracellular matrix remodelling, which were up regulated in scurs tissues relative to polled. Conclusion For this first time we describe networks of genes involved in horn and scurs development. Interestingly, we did not observe differential expression in any of the genes present on the fine mapped region of BTA1 known to contain the Poll locus. PMID:20537189

  18. Role of Host Type IA Phosphoinositide 3-Kinase Pathway Components in Invasin-Mediated Internalization of Yersinia enterocolitica.

    PubMed

    Dowd, Georgina C; Bhalla, Manmeet; Kean, Bernard; Thomas, Rowan; Ireton, Keith

    2016-06-01

    Many bacterial pathogens subvert mammalian type IA phosphoinositide 3-kinase (PI3K) in order to induce their internalization into host cells. How PI3K promotes internalization is not well understood. Also unclear is whether type IA PI3K affects different pathogens through similar or distinct mechanisms. Here, we performed an RNA interference (RNAi)-based screen to identify components of the type IA PI3K pathway involved in invasin-mediated entry of Yersinia enterocolitica, an enteropathogen that causes enteritis and lymphadenitis. The 69 genes targeted encode known upstream regulators or downstream effectors of PI3K. A similar RNAi screen was previously performed with the food-borne bacterium Listeria monocytogenes The results of the screen with Y. enterocolitica indicate that at least nine members of the PI3K pathway are needed for invasin-mediated entry. Several of these proteins, including centaurin-α1, Dock180, focal adhesion kinase (FAK), Grp1, LL5α, LL5β, and PLD2 (phospholipase D2), were recruited to sites of entry. In addition, centaurin-α1, FAK, PLD2, and mTOR were required for remodeling of the actin cytoskeleton during entry. Six of the human proteins affecting invasin-dependent internalization also promote InlB-mediated entry of L. monocytogenes Our results identify several host proteins that mediate invasin-induced effects on the actin cytoskeleton and indicate that a subset of PI3K pathway components promote internalization of both Y. enterocolitica and L. monocytogenes. PMID:27068087

  19. A 2-component system is involved in the early stages of the Pisolithus tinctorius-Pinus greggii symbiosis.

    PubMed

    Herrera-Martínez, Aseneth; Ruiz-Medrano, Roberto; Galván-Gordillo, Santiago Valentín; Toscano Morales, Roberto; Gómez-Silva, Lidia; Valdés, María; Hinojosa-Moya, Jesús; Xoconostle-Cázares, Beatriz

    2014-01-01

    Ectomycorrhizal symbiosis results in profound morphological and physiological modifications in both plant and fungus. This in turn is the product of differential gene expression in both co-symbionts, giving rise to specialized cell types capable of performing novel functions. During the precolonization stage, chemical signals from root exudates are sensed by the ectomycorrizal fungus, and vice versa, which are in principle responsible for the observed change in the developmental symbionts program. Little is known about the molecular mechanisms involved in the signaling and recognition between ectomycorrhizal fungi and their host plants. In the present work, we characterized a novel lactone, termed pinelactone, and identified a gene encoding for a histidine kinase in Pisolithus tictorius, which function is proposed to be the perception of the aforementioned metabolites. In this study, the use of closantel, a specific inhibitor of histidine kinase phosphorylation, affected the capacity for fungal colonization in the symbiosis between Pisolithus tinctorius and Pinus greggii, indicating that a 2-component system (TCS) may operate in the early events of plant-fungus interaction. Indeed, the metabolites induced the accumulation of Pisolithus tinctorius mRNA for a putative histidine kinase (termed Pthik1). Of note, Pthik1 was able to partially complement a S. cerevisiae histidine kinase mutant, demonstrating its role in the response to the presence of the aforementioned metabolites. Our results indicate a role of a 2-component pathway in the early stages of ectomycorrhizal symbiosis before colonization. Furthermore, a novel lactone from Pinus greggii root exudates may activate a signal transduction pathway that contributes to the establishment of the ectomycorrhizal symbiosis. PMID:24704731

  20. Involvement of the GABAergic septo-hippocampal pathway in brain stimulation reward.

    PubMed

    Vega-Flores, Germán; Gruart, Agnès; Delgado-García, José M

    2014-01-01

    The hippocampus is a structure related to several cognitive processes, but not very much is known about its putative involvement in positive reinforcement. In its turn, the septum has been related to instrumental brain stimulation reward (BSR) by its electrical stimulation with trains of pulses. Although the anatomical relationships of the septo-hippocampal pathway are well established, the functional relationship between these structures during rewarding behaviors remains poorly understood. To explore hippocampal mechanisms involved in BSR, CA3-evoked field excitatory and inhibitory postsynaptic potentials (fEPSPs, fIPSPs) were recorded in the CA1 area during BSR in alert behaving mice. The synaptic efficiency was determined from changes in fEPSP and fIPSP amplitudes across the learning of a BSR task. The successive BSR sessions evoked a progressive increase of the performance in inverse relationship with a decrease in the amplitude of fEPSPs, but not of fIPSPs. Additionally, we evaluated CA1 local field potentials (LFPs) during a preference task, comparing 8-, 20-, and 100-Hz trains of septal BSR. We corroborate a clear preference for BSR at 100 Hz (in comparison with BSR at 20 Hz or 8 Hz), in parallel with an increase in the spectral power of the low theta band, and a decrease in the gamma. These results were replicated by intrahippocampal injections of a GABAB antagonist. Thus, the GABAergic septo-hippocampal pathway seems to carry information involved in the encoding of reward properties, where GABAB receptors seem to play a key role. With regard to the dorsal hippocampus, fEPSPs evoked at the CA3-CA1 synapse seem to reflect the BSR learning process, while hippocampal rhythmic activities are more related to reward properties. PMID:25415445

  1. Involvement of the GABAergic Septo-Hippocampal Pathway in Brain Stimulation Reward

    PubMed Central

    Vega-Flores, Germán; Gruart, Agnès; Delgado-García, José M.

    2014-01-01

    The hippocampus is a structure related to several cognitive processes, but not very much is known about its putative involvement in positive reinforcement. In its turn, the septum has been related to instrumental brain stimulation reward (BSR) by its electrical stimulation with trains of pulses. Although the anatomical relationships of the septo-hippocampal pathway are well established, the functional relationship between these structures during rewarding behaviors remains poorly understood. To explore hippocampal mechanisms involved in BSR, CA3-evoked field excitatory and inhibitory postsynaptic potentials (fEPSPs, fIPSPs) were recorded in the CA1 area during BSR in alert behaving mice. The synaptic efficiency was determined from changes in fEPSP and fIPSP amplitudes across the learning of a BSR task. The successive BSR sessions evoked a progressive increase of the performance in inverse relationship with a decrease in the amplitude of fEPSPs, but not of fIPSPs. Additionally, we evaluated CA1 local field potentials (LFPs) during a preference task, comparing 8-, 20-, and 100-Hz trains of septal BSR. We corroborate a clear preference for BSR at 100 Hz (in comparison with BSR at 20 Hz or 8 Hz), in parallel with an increase in the spectral power of the low theta band, and a decrease in the gamma. These results were replicated by intrahippocampal injections of a GABAB antagonist. Thus, the GABAergic septo-hippocampal pathway seems to carry information involved in the encoding of reward properties, where GABAB receptors seem to play a key role. With regard to the dorsal hippocampus, fEPSPs evoked at the CA3-CA1 synapse seem to reflect the BSR learning process, while hippocampal rhythmic activities are more related to reward properties. PMID:25415445

  2. Expression of hedgehog pathway components in prostate carcinoma microenvironment: shifting the balance towards autocrine signalling

    PubMed Central

    Tzelepi, Vassiliki; Karlou, Maria; Wen, Sijin; Hoang, Anh; Logothetis, Christopher; Troncoso, Patricia; Efstathiou, Eleni

    2016-01-01

    Aims The hedgehog (Hh) signalling pathway has been implicated in the pathogenesis and aggressiveness of prostate cancer through epithelial–mesenchymal interactions. The aim of this study was to elucidate the cell-type partitioned expression of the Hh pathway biomarkers in the non-neoplastic and tumour microenvironments and to correlate it with the grade and stage of prostate cancer. Methods and results Expression of the Hh pathway components (Shh, Smo, Ptch, Gli1) in the microenvironment of non-neoplastic peripheral zone (n = 119), hormone-naive primary prostate carcinoma (n = 141) and castrate-resistant bone marrow metastases (n = 53) was analysed using immunohistochemistry in tissue microarrays and bone marrow sections. Results showed that epithelial Shh, Smo and Ptch expression was up-regulated, whereas stromal Smo, Ptch, and Gli1 expression was down-regulated in prostate carcinomas compared to non-neoplastic peripheral zone tissue. Ptch expression was modulated further in high-grade and high-stage primary tumours and in bone marrow metastases. Hh signalling correlated with ki67 and vascular endothelial growth factor (VEGF) but not with CD31 expression. Conclusion Our results highlight the importance of Hh-mediated epithelial–mesenchymal interactions in the non-neoplastic prostate and imply that shifting the balance from paracrine towards autocrine signalling is important in the pathogenesis and progression of prostate carcinoma. PMID:21707705

  3. Virtual reality and claustrophobia: multiple components therapy involving game editor virtual environments exposure.

    PubMed

    Malbos, E; Mestre, D R; Note, I D; Gellato, C

    2008-12-01

    The effectiveness of a multiple components therapy regarding claustrophobia and involving virtual reality (VR) will be demonstrated through a trial which immersed six claustrophobic patients in multiple context-graded enclosed virtual environments (VE) using affordable VR apparatus and software. The results of the questionnaires and behavior tests exhibited a significant reduction in fear towards the enclosed space and quality of life improvement. Such gains were maintained at 6-month follow-up. Presence score indicated the patients felt immersed and present inside the game editor VE. PMID:18954278

  4. A Computational Model of a Descending Mechanosensory Pathway Involved in Active Tactile Sensing

    PubMed Central

    Ache, Jan M.; Dürr, Volker

    2015-01-01

    Many animals, including humans, rely on active tactile sensing to explore the environment and negotiate obstacles, especially in the dark. Here, we model a descending neural pathway that mediates short-latency proprioceptive information from a tactile sensor on the head to thoracic neural networks. We studied the nocturnal stick insect Carausius morosus, a model organism for the study of adaptive locomotion, including tactually mediated reaching movements. Like mammals, insects need to move their tactile sensors for probing the environment. Cues about sensor position and motion are therefore crucial for the spatial localization of tactile contacts and the coordination of fast, adaptive motor responses. Our model explains how proprioceptive information about motion and position of the antennae, the main tactile sensors in insects, can be encoded by a single type of mechanosensory afferents. Moreover, it explains how this information is integrated and mediated to thoracic neural networks by a diverse population of descending interneurons (DINs). First, we quantified responses of a DIN population to changes in antennal position, motion and direction of movement. Using principal component (PC) analysis, we find that only two PCs account for a large fraction of the variance in the DIN response properties. We call the two-dimensional space spanned by these PCs ‘coding-space’ because it captures essential features of the entire DIN population. Second, we model the mechanoreceptive input elements of this descending pathway, a population of proprioceptive mechanosensory hairs monitoring deflection of the antennal joints. Finally, we propose a computational framework that can model the response properties of all important DIN types, using the hair field model as its only input. This DIN model is validated by comparison of tuning characteristics, and by mapping the modelled neurons into the two-dimensional coding-space of the real DIN population. This reveals the

  5. A Computational Model of a Descending Mechanosensory Pathway Involved in Active Tactile Sensing.

    PubMed

    Ache, Jan M; Dürr, Volker

    2015-07-01

    Many animals, including humans, rely on active tactile sensing to explore the environment and negotiate obstacles, especially in the dark. Here, we model a descending neural pathway that mediates short-latency proprioceptive information from a tactile sensor on the head to thoracic neural networks. We studied the nocturnal stick insect Carausius morosus, a model organism for the study of adaptive locomotion, including tactually mediated reaching movements. Like mammals, insects need to move their tactile sensors for probing the environment. Cues about sensor position and motion are therefore crucial for the spatial localization of tactile contacts and the coordination of fast, adaptive motor responses. Our model explains how proprioceptive information about motion and position of the antennae, the main tactile sensors in insects, can be encoded by a single type of mechanosensory afferents. Moreover, it explains how this information is integrated and mediated to thoracic neural networks by a diverse population of descending interneurons (DINs). First, we quantified responses of a DIN population to changes in antennal position, motion and direction of movement. Using principal component (PC) analysis, we find that only two PCs account for a large fraction of the variance in the DIN response properties. We call the two-dimensional space spanned by these PCs 'coding-space' because it captures essential features of the entire DIN population. Second, we model the mechanoreceptive input elements of this descending pathway, a population of proprioceptive mechanosensory hairs monitoring deflection of the antennal joints. Finally, we propose a computational framework that can model the response properties of all important DIN types, using the hair field model as its only input. This DIN model is validated by comparison of tuning characteristics, and by mapping the modelled neurons into the two-dimensional coding-space of the real DIN population. This reveals the versatility

  6. Differential regulation of calcium signalling pathways by components of Piper methysticum ('Awa).

    PubMed

    Shimoda, L M N; Showman, A; Baker, J D; Lange, I; Koomoa, D L; Stokes, A J; Borris, R P; Turner, H

    2015-04-01

    Kava is a soporific, anxiolytic and relaxant in widespread ritual and recreational use throughout the Pacific. Traditional uses of kava by indigenous Pacific Island peoples reflect a complex pharmacopeia, centered on GABA-ergic effects of the well-characterized kavalactones. However, peripheral effects of kava suggest active components other than the CNS-targeted kavalactones. We have previously shown that immunocytes exhibit calcium mobilization in response to traditionally prepared kava extracts, and that the kavalactones do not induce these calcium responses. Here, we characterize the complex calcium-mobilizing activity of traditionally prepared and partially HPLC-purified kava extracts, noting induction of both calcium entry and store release pathways. Kava components activate intracellular store depletion of thapsigargin-sensitive and -insensitive stores that are coupled to the calcium release activated (CRAC) current, and cause calcium entry through non-store-operated pathways. Together with the pepper-like potency reported by kava users, these studies lead us to hypothesize that kava extracts contain one or more ligands for the transient receptor potential (TRP) family of ion channels. Indeed, TRP-like conductances are observed in kava-treated cells under patch clamp. Thus TRP-mediated cellular effects may be responsible for some of the reported pharmacology of kava. PMID:25640812

  7. Differential regulation of calcium signalling pathways by components of Piper methysticum (‘Awa)

    PubMed Central

    Shimoda, L.M.N; Showman, A.; Baker, J.D.; Lange, I.; Koomoa, D.L.; Stokes, A.J.; Borris, R.P.; Turner, H.

    2015-01-01

    Kava is a soporific, anxiolytic and relaxant in widespread ritual and recreational use throughout the Pacific. Traditional uses of kava by indigenous Pacific Island peoples reflect a complex pharmacopeia, centered on GABA-ergic effects of the well-characterized kavalactones. However, peripheral effects of kava suggest active components other than the CNS-targeted kavalactones. We have previously shown that immunocytes exhibit calcium mobilization in response to traditionally-prepared kava extracts, and that the kavalactones do not induce these calcium responses. Here, we characterize the complex calcium-mobilizing activity of traditionally-prepared and partially HPLC-purified kava extracts, noting induction of both calcium entry and store release pathways. Kava components activate intracellular store depletion of thapsigargin-sensitive and –insensitive stores that are coupled to the calcium release activated (CRAC) current, and cause calcium entry through non-store-operated pathways. Together with the pepper-like potency reported by kava users, these studies lead us to hypothesize that kava extracts contain one or more ligands for the transient receptor potential (TRP) family of ion channels. Indeed, TRP-like conductances are observed in kava-treated cells under patch clamp. Thus TRP-mediated cellular effects may be responsible for some of the reported pharmacology of kava. PMID:25640812

  8. The mammalian molecular clockwork controls rhythmic expression of its own input pathway components.

    PubMed

    Pfeffer, Martina; Müller, Christian M; Mordel, Jérôme; Meissl, Hilmar; Ansari, Nariman; Deller, Thomas; Korf, Horst-Werner; von Gall, Charlotte

    2009-05-13

    The core molecular clockwork in the suprachiasmatic nucleus (SCN) is based on autoregulatory feedback loops of transcriptional activators (CLOCK/NPAS2 and BMAL1) and inhibitors (mPER1-2 and mCRY1-2). To synchronize the phase of the molecular clockwork to the environmental day and night condition, light at dusk and dawn increases mPer expression. However, the signal transduction pathways differ remarkably between the day/night and the night/day transition. Light during early night leads to intracellular Ca(2+) release by neuronal ryanodine receptors (RyRs), resulting in phase delays. Light during late night triggers an increase in guanylyl cyclase activity, resulting in phase advances. To date, it is still unknown how the core molecular clockwork regulates the availability of the respective input pathway components. Therefore, we examined light resetting mechanisms in mice with an impaired molecular clockwork (BMAL1(-/-)) and the corresponding wild type (BMAL1(+/+)) using in situ hybridization, real-time PCR, immunohistochemistry, and a luciferase reporter system. In addition, intracellular calcium concentrations (Ca(2+)(i)) were measured in SCN slices using two-photon microscopy. In the SCN of BMAL1(-/-) mice Ryr mRNA and RyR protein levels were reduced, and light-induced mPer expression was selectively impaired during early night. Transcription assays with NIH3T3 fibroblasts showed that Ryr expression was activated by CLOCK::BMAL1 and inhibited by mCRY1. The Ca(2+)(i) response of SCN cells to the RyR agonist caffeine was reduced in BMAL1(-/-) compared with BMAL1(+/+) mice. Our findings provide the first evidence that the mammalian molecular clockwork influences Ryr expression and thus controls its own photic input pathway components. PMID:19439589

  9. Tanshinol Rescues the Impaired Bone Formation Elicited by Glucocorticoid Involved in KLF15 Pathway

    PubMed Central

    Yang, Yajun; Su, Yanjie; Wang, Dongtao; Chen, Yahui; Liu, Yuyu; Luo, Shiying; Wu, Tie

    2016-01-01

    Decreased bone formation is responsible for the pathogenesis of glucocorticoid- (GC-) induced osteoporosis (GIO), while the mechanism remains to be elucidated. The aim was to investigate how natural antioxidant tanshinol attenuates oxidative stress and rescues impaired bone formation elicited by GC in Sprague-Dawley rats and in C2C12 cells and/or MC3T3-E1 cells. The results showed that tanshinol prevented bone loss and decreased biomechanical characteristics and suppressed reduction of biomarkers related to osteogenesis in GIO rats. Further study revealed that tanshinol reversed decrease of transcription activity of Osterix-luc and rescued impairment of osteoblastic differentiation and bone formation involved in induction of KLF15 mRNA. Meanwhile, tanshinol diminished inhibition of protein expression of β-catenin and Tcf4 and transcription activity of Tcf4-luc induced by GC, especially under conditions of KLF siRNA in vitro. Additionally, tanshinol attenuated increase of reactive oxygen species (ROS) generation, phosphorylation of p66Shc expression, TUNEL-positive cells, and caspase-3 activity elicited by KLF15 under conditions of GC. Taken together, the present findings suggest that tanshinol attenuated the decrease of bone formation and bone mass and bone quality elicited by GC involved in KLF15/Wnt signaling transduction and counteracted GC-evoked oxidative stress and subsequent cell apoptosis involved in KLF15/p66Shc pathway cascade. PMID:27051474

  10. The mouse and human genes encoding the recognition component of the N-end rule pathway

    PubMed Central

    Kwon, Yong Tae; Reiss, Yuval; Fried, Victor A.; Hershko, Avram; Yoon, Jeong Kyo; Gonda, David K.; Sangan, Pitchai; Copeland, Neal G.; Jenkins, Nancy A.; Varshavsky, Alexander

    1998-01-01

    The N-end rule relates the in vivo half-life of a protein to the identity of its N-terminal residue. The N-end rule pathway is one proteolytic pathway of the ubiquitin system. The recognition component of this pathway, called N-recognin or E3, binds to a destabilizing N-terminal residue of a substrate protein and participates in the formation of a substrate-linked multiubiquitin chain. We report the cloning of the mouse and human Ubr1 cDNAs and genes that encode a mammalian N-recognin called E3α. Mouse UBR1p (E3α) is a 1,757-residue (200-kDa) protein that contains regions of sequence similarity to the 225-kDa Ubr1p of the yeast Saccharomyces cerevisiae. Mouse and human UBR1p have apparent homologs in other eukaryotes as well, thus defining a distinct family of proteins, the UBR family. The residues essential for substrate recognition by the yeast Ubr1p are conserved in the mouse UBR1p. The regions of similarity among the UBR family members include a putative zinc finger and RING-H2 finger, another zinc-binding domain. Ubr1 is located in the middle of mouse chromosome 2 and in the syntenic 15q15-q21.1 region of human chromosome 15. Mouse Ubr1 spans ≈120 kilobases of genomic DNA and contains ≈50 exons. Ubr1 is ubiquitously expressed in adults, with skeletal muscle and heart being the sites of highest expression. In mouse embryos, the Ubr1 expression is highest in the branchial arches and in the tail and limb buds. The cloning of Ubr1 makes possible the construction of Ubr1-lacking mouse strains, a prerequisite for the functional understanding of the mammalian N-end rule pathway. PMID:9653112

  11. Expression Analysis of Genes Involved in the RB/E2F Pathway in Astrocytic Tumors

    PubMed Central

    Ferreira, Wallax Augusto Silva; Araújo, Mariana Diniz; de Oliveira, Edivaldo Herculano Correa; Brito, José Reginaldo Nascimento; Burbano, Rommel Rodriguez; Harada, Maria Lúcia; Borges, Bárbara do Nascimento

    2015-01-01

    Astrocytic gliomas, which are derived from glial cells, are considered the most common primary neoplasias of the central nervous system (CNS) and are histologically classified as low grade (I and II) or high grade (III and IV). Recent studies have shown that astrocytoma formation is the result of the deregulation of several pathways, including the RB/E2F pathway, which is commonly deregulated in various human cancers via genetic or epigenetic mechanisms. On the basis of the assumption that the study of the mechanisms controlling the INK4/ARF locus can help elucidate the molecular pathogenesis of astrocytic tumors, identify diagnostic and prognostic markers, and help select appropriate clinical treatments, the present study aimed to evaluate and compare methylation patterns using bisulfite sequencing PCR and evaluate the gene expression profile using real-time PCR in the genes CDKN2A, CDKN2B, CDC6, Bmi-1, CCND1, and RB1 in astrocytic tumors. Our results indicate that all the evaluated genes are not methylated independent of the tumor grade. However, the real-time PCR results indicate that these genes undergo progressive deregulation as a function of the tumor grade. In addition, the genes CDKN2A, CDKN2B, and RB1 were underexpressed, whereas CDC6, Bmi-1, and CCND1 were overexpressed; the increase in gene expression was significantly associated with decreased patient survival. Therefore, we propose that the evaluation of the expression levels of the genes involved in the RB/E2F pathway can be used in the monitoring of patients with astrocytomas in clinical practice and for the prognostic indication of disease progression. PMID:26317630

  12. Expression Analysis of Genes Involved in the RB/E2F Pathway in Astrocytic Tumors.

    PubMed

    Ferreira, Wallax Augusto Silva; Araújo, Mariana Diniz; Anselmo, Nilson Praia; de Oliveira, Edivaldo Herculano Correa; Brito, José Reginaldo Nascimento; Burbano, Rommel Rodriguez; Harada, Maria Lúcia; Borges, Bárbara do Nascimento

    2015-01-01

    Astrocytic gliomas, which are derived from glial cells, are considered the most common primary neoplasias of the central nervous system (CNS) and are histologically classified as low grade (I and II) or high grade (III and IV). Recent studies have shown that astrocytoma formation is the result of the deregulation of several pathways, including the RB/E2F pathway, which is commonly deregulated in various human cancers via genetic or epigenetic mechanisms. On the basis of the assumption that the study of the mechanisms controlling the INK4/ARF locus can help elucidate the molecular pathogenesis of astrocytic tumors, identify diagnostic and prognostic markers, and help select appropriate clinical treatments, the present study aimed to evaluate and compare methylation patterns using bisulfite sequencing PCR and evaluate the gene expression profile using real-time PCR in the genes CDKN2A, CDKN2B, CDC6, Bmi-1, CCND1, and RB1 in astrocytic tumors. Our results indicate that all the evaluated genes are not methylated independent of the tumor grade. However, the real-time PCR results indicate that these genes undergo progressive deregulation as a function of the tumor grade. In addition, the genes CDKN2A, CDKN2B, and RB1 were underexpressed, whereas CDC6, Bmi-1, and CCND1 were overexpressed; the increase in gene expression was significantly associated with decreased patient survival. Therefore, we propose that the evaluation of the expression levels of the genes involved in the RB/E2F pathway can be used in the monitoring of patients with astrocytomas in clinical practice and for the prognostic indication of disease progression. PMID:26317630

  13. Pathways Involved in the Synergistic Activation of Macrophages by Lipoteichoic Acid and Hemoglobin

    PubMed Central

    Cox, Kathleen H.; Cox, Michelle E.; Woo-Rasberry, Virginia; Hasty, David L.

    2012-01-01

    Lipoteichoic acid (LTA) is a Gram-positive cell surface molecule that is found in both a cell-bound form and cell-free form in the host during an infection. Hemoglobin (Hb) can synergize with LTA, a TLR2 ligand, to potently activate macrophage innate immune responses in a TLR2- and TLR4-dependent way. At low levels of LTA, the presence of Hb can result in a 200-fold increase in the secretion of IL-6 following macrophage activation. Six hours after activation, the macrophage genes that are most highly up-regulated by LTA plus Hb activation compared to LTA alone are cytokines, chemokines, receptors and interferon-regulated genes. Several of these genes exhibit a unique TLR4-dependent increase in mRNA levels that continued to rise more than eight hours after stimulation. This prolonged increase in mRNA levels could be the result of an extended period of NF-κB nuclear localization and the concurrent absence of the NF-κB inhibitor, IκBα, after stimulation with LTA plus Hb. Dynasore inhibition experiments indicate that an endocytosis-dependent pathway is required for the TLR4-dependent up-regulation of IL-6 secretion following activation with LTA plus Hb. In addition, interferon-β mRNA is present after activation with LTA plus Hb, suggesting that the TRIF/TRAM-dependent pathway may be involved. Hb alone can elicit the TLR4-dependent secretion of TNF-α from macrophages, so it may be the TLR4 ligand. Hb also led to secretion of high mobility group box 1 protein (HMGB1), which synergized with LTA to increase secretion of IL-6. The activation of both the TLR2 and TLR4 pathways by LTA plus Hb leads to an enhanced innate immune response. PMID:23071790

  14. Identification of Interphase Functions for the NIMA Kinase Involving Microtubules and the ESCRT Pathway

    PubMed Central

    Govindaraghavan, Meera; McGuire Anglin, Sarah Lea; Shen, Kuo-Fang; Shukla, Nandini; De Souza, Colin P.; Osmani, Stephen A.

    2014-01-01

    The Never in Mitosis A (NIMA) kinase (the founding member of the Nek family of kinases) has been considered a mitotic specific kinase with nuclear restricted roles in the model fungus Aspergillus nidulans. By extending to A. nidulans the results of a synthetic lethal screen performed in Saccharomyces cerevisiae using the NIMA ortholog KIN3, we identified a conserved genetic interaction between nimA and genes encoding proteins of the Endosomal Sorting Complex Required for Transport (ESCRT) pathway. Absence of ESCRT pathway functions in combination with partial NIMA function causes enhanced cell growth defects, including an inability to maintain a single polarized dominant cell tip. These genetic insights suggest NIMA potentially has interphase functions in addition to its established mitotic functions at nuclei. We therefore generated endogenously GFP-tagged NIMA (NIMA-GFP) which was fully functional to follow its interphase locations using live cell spinning disc 4D confocal microscopy. During interphase some NIMA-GFP locates to the tips of rapidly growing cells and, when expressed ectopically, also locates to the tips of cytoplasmic microtubules, suggestive of non-nuclear interphase functions. In support of this, perturbation of NIMA function either by ectopic overexpression or through partial inactivation results in marked cell tip growth defects with excess NIMA-GFP promoting multiple growing cell tips. Ectopic NIMA-GFP was found to locate to the plus ends of microtubules in an EB1 dependent manner, while impairing NIMA function altered the dynamic localization of EB1 and the cytoplasmic microtubule network. Together, our genetic and cell biological analyses reveal novel non-nuclear interphase functions for NIMA involving microtubules and the ESCRT pathway for normal polarized fungal cell tip growth. These insights extend the roles of NIMA both spatially and temporally and indicate that this conserved protein kinase could help integrate cell cycle progression

  15. Involvement of multiple cellular pathways in regulating resistance to tamoxifen in BIK-suppressed MCF-7 cells.

    PubMed

    Viedma-Rodríguez, Rubí; Ruiz Esparza-Garrido, Ruth; Baiza-Gutman, Luis Arturo; Velázquez-Flores, Miguel Ángel; García-Carrancá, Alejandro; Salamanca-Gómez, Fabio; Arenas-Aranda, Diego

    2015-09-01

    Majority of women with estrogen receptor (ER)-positive breast cancers initially respond to hormone therapies such as tamoxifen (TAM; antagonist of estrogen). However, many tumors eventually become resistant to TAM. Therefore, understanding the various cellular components involved in causing resistance to TAM is of paramount importance in designing novel entities for efficacious hormone therapy. Previously, we found that suppression of BIK gene expression induced TAM resistance in MCF-7 breast cancer cells. In order to understand the response of these cells to TAM and its association with resistance, a microarray analysis of gene expression was performed in the BIK-suppressed MCF-7 cells and compared it to the TAM-only-treated cells (controls). Several genes participating in various cellular pathways were identified. Molecules identified in the drug resistance pathway were 14-3-3z or YWHAZ, WEE1, PRKACA, NADK, and HSP90AA 1. Further, genes involved in cell cycle control, apoptosis, and cell proliferation were also found differentially expressed in these cells. Transcriptional and translational analysis of key molecules such as STAT2, AKT 3, and 14-3-3z revealed similar changes at the messenger RNA (mRNA) as well as at the protein level. Importantly, there was no cytotoxic effect of TAM on BIK-suppressed MCF-7 cells. Further, these cells were not arrested at the G0-G1 phase of the cell cycle although 30 % of BIK-suppressed cells were arrested at the G2 phase of the cycle on TAM treatment. Furthermore, we found a relevant interaction between 14-3-3z and WEE1, suggesting that the cytotoxic effect of TAM was prevented in BIK-suppressed cells because this interaction leads to transitory arrest in the G2 phase leading to the repair of damaged DNA and allowing the cells to proliferate. PMID:25861752

  16. Metagenomic identification of bacterioplankton taxa and pathways involved in microcystin degradation in lake erie.

    PubMed

    Mou, Xiaozhen; Lu, Xinxin; Jacob, Jisha; Sun, Shulei; Heath, Robert

    2013-01-01

    Cyanobacterial harmful blooms (CyanoHABs) that produce microcystins are appearing in an increasing number of freshwater ecosystems worldwide, damaging quality of water for use by human and aquatic life. Heterotrophic bacteria assemblages are thought to be important in transforming and detoxifying microcystins in natural environments. However, little is known about their taxonomic composition or pathways involved in the process. To address this knowledge gap, we compared the metagenomes of Lake Erie free-living bacterioplankton assemblages in laboratory microcosms amended with microcystins relative to unamended controls. A diverse array of bacterial phyla were responsive to elevated supply of microcystins, including Acidobacteria, Actinobacteria, Bacteroidetes, Planctomycetes, Proteobacteria of the alpha, beta, gamma, delta and epsilon subdivisions and Verrucomicrobia. At more detailed taxonomic levels, Methylophilales (mainly in genus Methylotenera) and Burkholderiales (mainly in genera Bordetella, Burkholderia, Cupriavidus, Polaromonas, Ralstonia, Polynucleobacter and Variovorax) of Betaproteobacteria were suggested to be more important in microcystin degradation than Sphingomonadales of Alphaproteobacteria. The latter taxa were previously thought to be major microcystin degraders. Homologs to known microcystin-degrading genes (mlr) were not overrepresented in microcystin-amended metagenomes, indicating that Lake Erie bacterioplankton might employ alternative genes and/or pathways in microcystin degradation. Genes for xenobiotic metabolism were overrepresented in microcystin-amended microcosms, suggesting they are important in bacterial degradation of microcystin, a phenomenon that has been identified previously only in eukaryotic systems. PMID:23637924

  17. Quantitative evaluation of pathways involved in trichloroethylene reduction by zero-valent metals: Iron and zinc

    SciTech Connect

    Arnold, W.; Roberts, A.L.; Burris, D.R.; Campbell, T.J.

    1995-12-31

    In order to design in situ remediation systems using zero-valent metals, the mechanism and kinetics of chlorinated solvent degradation by zero-valent metals need clarification. These issues are addressed by conducting detailed investigations of the pathways involved in trichloroethylene (TCE) reduction by two zero-valent metals. Analyses are based on batch reaction data for chloroethylene reduction by iron and zinc. Experiments were conducted using TCE and each readily available reaction product of TCE degradation as a starting material and monitoring the disappearance of the parent chemical and the appearance of reaction products over time. Models were developed by working backwards through the hypothesized reaction sequence. Determining rate constants for the latter steps in the pathway, inserting them into the more complex models for more highly oxidized compounds, and obtaining rate constants for the remaining steps in the transformation of the oxidized species was repeated until a model for trichloroethylene was developed. Results indicate that reactions may not occur via a process of sequential hydrogenolysis or hydrogenation. Ethylene and/or ethane production are too rapid to be accounted for in this manner. The product distribution, especially the presence of acetylene, can only be explained by invoking reductive elimination reactions.

  18. Signaling pathways involved in megakaryocyte-mediated proliferation of osteoblast lineage cells.

    PubMed

    Cheng, Ying-Hua; Streicher, Drew A; Waning, David L; Chitteti, Brahmananda R; Gerard-O'Riley, Rita; Horowitz, Mark C; Bidwell, Joseph P; Pavalko, Fredrick M; Srour, Edward F; Mayo, Lindsey D; Kacena, Melissa A

    2015-03-01

    Recent studies suggest that megakaryocytes (MKs) may play a significant role in skeletal homeostasis, as evident by the occurrence of osteosclerosis in multiple MK related diseases (Lennert et al., 1975; Thiele et al., 1999; Chagraoui et al., 2006). We previously reported a novel interaction whereby MKs enhanced proliferation of osteoblast lineage/osteoprogenitor cells (OBs) by a mechanism requiring direct cell-cell contact. However, the signal transduction pathways and the downstream effector molecules involved in this process have not been characterized. Here we show that MKs contact with OBs, via beta1 integrin, activate the p38/MAPKAPK2/p90RSK kinase cascade in the bone cells, which causes Mdm2 to neutralizes p53/Rb-mediated check point and allows progression through the G1/S. Interestingly, activation of MAPK (ERK1/2) and AKT, collateral pathways that regulate the cell cycle, remained unchanged with MK stimulation of OBs. The MK-to-OB signaling ultimately results in significant increases in the expression of c-fos and cyclin A, necessary for sustaining the OB proliferation. Overall, our findings show that OBs respond to the presence of MKs, in part, via an integrin-mediated signaling mechanism, activating a novel response axis that de-represses cell cycle activity. Understanding the mechanisms by which MKs enhance OB proliferation will facilitate the development of novel anabolic therapies to treat bone loss associated with osteoporosis and other bone-related diseases. PMID:25160801

  19. PI3K/Akt signaling pathway is involved in the neurotrophic effect of senegenin.

    PubMed

    Pi, Ting; Zhou, Xiao-Wen; Cai, Liang; Zhang, Wei; Su, Chao-Fen; Wu, Wu-Tian; Ren, Xiao-Ming; Luo, Huan-Min

    2016-02-01

    Neurodegenerative diseases are frequently associated with the loss of synapses and neurons. Senegenin, extracted from the Chinese herb Polygala tenuifolia Willd, was previously found to promote neurite outgrowth and neuronal survival in primary cultured rat cortical neurons. The aim of the present study was to investigate the underlying mechanisms of senegenin-induced neurotrophic effects on rat cortical neurons. Primary cortical rat neurons were treated with various pharmacological antagonists and with or without senegenin, and subjected to MTT and western blot analysis to explore the effects of senegenin on cell survival as well as the activation of signaling pathways. Neurite outgrowth and neuronal survival induced by senegenin were significantly inhibited by A2A receptor antagonist ZM241385 and specific phosphoinositide-3 kinase (PI3K) inhibitor LY294002, but not by tropomyosin receptor kinase A receptor inhibitor K252a, mitogen-activated protein kinase kinase inhibitor PD98059 or protein kinase C inhibitor GÖ6976. Furthermore, senegenin enhanced the phosphorylation of Akt, which was blocked by LY294002. The present study revealed that the PI3K/Akt signaling pathway may be involved in the neurotrophic effects of senegenin. PMID:26647727

  20. Metagenomic Identification of Bacterioplankton Taxa and Pathways Involved in Microcystin Degradation in Lake Erie

    PubMed Central

    Mou, Xiaozhen; Lu, Xinxin; Jacob, Jisha; Sun, Shulei; Heath, Robert

    2013-01-01

    Cyanobacterial harmful blooms (CyanoHABs) that produce microcystins are appearing in an increasing number of freshwater ecosystems worldwide, damaging quality of water for use by human and aquatic life. Heterotrophic bacteria assemblages are thought to be important in transforming and detoxifying microcystins in natural environments. However, little is known about their taxonomic composition or pathways involved in the process. To address this knowledge gap, we compared the metagenomes of Lake Erie free-living bacterioplankton assemblages in laboratory microcosms amended with microcystins relative to unamended controls. A diverse array of bacterial phyla were responsive to elevated supply of microcystins, including Acidobacteria, Actinobacteria, Bacteroidetes, Planctomycetes, Proteobacteria of the alpha, beta, gamma, delta and epsilon subdivisions and Verrucomicrobia. At more detailed taxonomic levels, Methylophilales (mainly in genus Methylotenera) and Burkholderiales (mainly in genera Bordetella, Burkholderia, Cupriavidus, Polaromonas, Ralstonia, Polynucleobacter and Variovorax) of Betaproteobacteria were suggested to be more important in microcystin degradation than Sphingomonadales of Alphaproteobacteria. The latter taxa were previously thought to be major microcystin degraders. Homologs to known microcystin-degrading genes (mlr) were not overrepresented in microcystin-amended metagenomes, indicating that Lake Erie bacterioplankton might employ alternative genes and/or pathways in microcystin degradation. Genes for xenobiotic metabolism were overrepresented in microcystin-amended microcosms, suggesting they are important in bacterial degradation of microcystin, a phenomenon that has been identified previously only in eukaryotic systems. PMID:23637924

  1. Involvement of Autophagic Pathway in the Progression of Retinal Degeneration in a Mouse Model of Diabetes

    PubMed Central

    Piano, Ilaria; Novelli, Elena; Della Santina, Luca; Strettoi, Enrica; Cervetto, Luigi; Gargini, Claudia

    2016-01-01

    The notion that diabetic retinopathy (DR) is essentially a micro-vascular disease has been recently challenged by studies reporting that vascular changes are preceded by signs of damage and loss of retinal neurons. As to the mode by which neuronal death occurs, the evidence that apoptosis is the main cause of neuronal loss is far from compelling. The objective of this study was to investigate these controversies in a mouse model of streptozotocin (STZ) induced diabetes. Starting from 8 weeks after diabetes induction there was loss of rod but not of cone photoreceptors, together with reduced thickness of the outer and inner synaptic layers. Correspondingly, rhodopsin expression was downregulated and the scotopic electroretinogram (ERG) is suppressed. In contrast, cone opsin expression and photopic ERG response were not affected. Suppression of the scotopic ERG preceded morphological changes as well as any detectable sign of vascular alteration. Only sparse apoptotic figures were detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and glia was not activated. The physiological autophagy flow was altered instead, as seen by increased LC3 immunostaining at the level of outer plexiform layer (OPL) and upregulation of the autophagic proteins Beclin-1 and Atg5. Collectively, our results show that the streptozotocin induced DR in mouse initiates with a functional loss of the rod visual pathway. The pathogenic pathways leading to cell death develop with the initial dysregulation of autophagy well before the appearance of signs of vascular damage and without strong involvement of apoptosis. PMID:26924963

  2. Neuroprotective action of cycloheximide involves induction of bcl-2 and antioxidant pathways.

    PubMed

    Furukawa, K; Estus, S; Fu, W; Mark, R J; Mattson, M P

    1997-03-10

    The ability of the protein synthesis inhibitor cycloheximide (CHX) to prevent neuronal death in different paradigms has been interpreted to indicate that the cell death process requires synthesis of "killer" proteins. On the other hand, data indicate that neurotrophic factors protect neurons in the same death paradigms by inducing expression of neuroprotective gene products. We now provide evidence that in embryonic rat hippocampal cell cultures, CHX protects neurons against oxidative insults by a mechanism involving induction of neuroprotective gene products including the antiapoptotic gene bcl-2 and antioxidant enzymes. Neuronal survival after exposure to glutamate, FeSO4, and amyloid beta-peptide was increased in cultures pretreated with CHX at concentrations of 50-500 nM; higher and lower concentrations were ineffective. Neuroprotective concentrations of CHX caused only a moderate (20-40%) reduction in overall protein synthesis, and induced an increase in c-fos, c-jun, and bcl-2 mRNAs and protein levels as determined by reverse transcription-PCR analysis and immunocytochemistry, respectively. At neuroprotective CHX concentrations, levels of c-fos heteronuclear RNA increased in parallel with c-fos mRNA, indicating that CHX acts by inducing transcription. Neuroprotective concentrations of CHX suppressed accumulation of H2O2 induced by FeSO4, suggesting activation of antioxidant pathways. Treatment of cultures with an antisense oligodeoxynucleotide directed against bcl-2 mRNA decreased Bcl-2 protein levels and significantly reduced the neuroprotective action of CHX, suggesting that induction of Bcl-2 expression was mechanistically involved in the neuroprotective actions of CHX. In addition, activity levels of the antioxidant enzymes Cu/Zn-superoxide dismutase, Mn-superoxide dismutase, and catalase were significantly increased in cultures exposed to neuroprotective levels of CHX. Our data suggest that low concentrations of CHX can promote neuron survival by

  3. Genetic risk for schizophrenia: convergence on synaptic pathways involved in plasticity.

    PubMed

    Hall, Jeremy; Trent, Simon; Thomas, Kerrie L; O'Donovan, Michael C; Owen, Michael J

    2015-01-01

    Recent large-scale genomic studies have revealed two broad classes of risk alleles for schizophrenia: a polygenic component of risk mediated through multiple common risk variants and rarer more highly penetrant submicroscopic chromosomal deletions and duplications, known as copy number variants. The focus of this review is on the emerging findings from the latter and subsequent exome sequencing data of smaller, deleterious single nucleotide variants and indels. In these studies, schizophrenia patients were found to have enriched de novo mutations in genes belonging to the postsynaptic density at glutamatergic synapses, particularly components of the N-methyl-D-aspartate receptor signaling complex, including the PSD-95 complex, activity-regulated cytoskeleton-associated protein interactors, the fragile X mental retardation protein complex, voltage-gated calcium channels, and genes implicated in actin cytoskeletal dynamics. The convergence of these implicated genes onto a coherent biological pathway at the synapse, with a specific role in plasticity, provides a significant advance in understanding pathogenesis and points to new targets for biological investigation. We consider the implications of these studies in the context of existing genetic data and the potential need to reassess diagnostic boundaries of neuropsychiatric disorders before discussing ways forward for more directed mechanistic studies to develop stratified, novel therapeutic approaches in the future. PMID:25152434

  4. Delineation of Molecular Pathways Involved in Cardiomyopathies Caused by Troponin T Mutations.

    PubMed

    Gilda, Jennifer E; Lai, Xianyin; Witzmann, Frank A; Gomes, Aldrin V

    2016-06-01

    Familial hypertrophic cardiomyopathy (FHC) is associated with mild to severe cardiac problems and is the leading cause of sudden death in young people and athletes. Although the genetic basis for FHC is well-established, the molecular mechanisms that ultimately lead to cardiac dysfunction are not well understood. To obtain important insights into the molecular mechanism(s) involved in FHC, hearts from two FHC troponin T models (Ile79Asn [I79N] and Arg278Cys [R278C]) were investigated using label-free proteomics and metabolomics. Mutations in troponin T are the third most common cause of FHC, and the I79N mutation is associated with a high risk of sudden cardiac death. Most FHC-causing mutations, including I79N, increase the Ca(2+) sensitivity of the myofilament; however, the R278C mutation does not alter Ca(2+) sensitivity and is associated with a better prognosis than most FHC mutations. Out of more than 1200 identified proteins, 53 and 76 proteins were differentially expressed in I79N and R278C hearts, respectively, when compared with wild-type hearts. Interestingly, more than 400 proteins were differentially expressed when the I79N and R278C hearts were directly compared. The three major pathways affected in I79N hearts relative to R278C and wild-type hearts were the ubiquitin-proteasome system, antioxidant systems, and energy production pathways. Further investigation of the proteasome system using Western blotting and activity assays showed that proteasome dysfunction occurs in I79N hearts. Metabolomic results corroborate the proteomic data and suggest the glycolytic, citric acid, and electron transport chain pathways are important pathways that are altered in I79N hearts relative to R278C or wild-type hearts. Our findings suggest that impaired energy production and protein degradation dysfunction are important mechanisms in FHCs associated with poor prognosis and that cardiac hypertrophy is not likely needed for a switch from fatty acid to glucose metabolism

  5. Early Brain Response to Low-Dose Radiation Exposure Involves Molecular Networks and Pathways Associated with Cognitive Functions, Advanced Aging and Alzheimer's Disease

    SciTech Connect

    Lowe, Xiu R; Bhattacharya, Sanchita; Marchetti, Francesco; Wyrobek, Andrew J.

    2008-06-06

    Understanding the cognitive and behavioral consequences of brain exposures to low-dose ionizing radiation has broad relevance for health risks from medical radiation diagnostic procedures, radiotherapy, environmental nuclear contamination, as well as earth orbit and space missions. Analyses of transcriptome profiles of murine brain tissue after whole-body radiation showed that low-dose exposures (10 cGy) induced genes not affected by high dose (2 Gy), and low-dose genes were associated with unique pathways and functions. The low-dose response had two major components: pathways that are consistently seen across tissues, and pathways that were brain tissue specific. Low-dose genes clustered into a saturated network (p < 10{sup -53}) containing mostly down-regulated genes involving ion channels, long-term potentiation and depression, vascular damage, etc. We identified 9 neural signaling pathways that showed a high degree of concordance in their transcriptional response in mouse brain tissue after low-dose radiation, in the aging human brain (unirradiated), and in brain tissue from patients with Alzheimer's disease. Mice exposed to high-dose radiation did not show these effects and associations. Our findings indicate that the molecular response of the mouse brain within a few hours after low-dose irradiation involves the down-regulation of neural pathways associated with cognitive dysfunctions that are also down regulated in normal human aging and Alzheimer's disease.

  6. The centrosomal component CEP161 of Dictyostelium discoideum interacts with the Hippo signaling pathway

    PubMed Central

    Sukumaran, Salil K.; Blau-Wasser, Rosemarie; Rohlfs, Meino; Gallinger, Christoph; Schleicher, Michael; Noegel, Angelika A

    2015-01-01

    CEP161 is a novel component of the Dictyostelium discoideum centrosome which was identified as binding partner of the pericentriolar component CP250. Here we show that the amino acids 1-763 of the 1381 amino acids CEP161 are sufficient for CP250 binding, centrosomal targeting and centrosome association. Analysis of AX2 cells over-expressing truncated and full length CEP161 proteins revealed defects in growth and development. By immunoprecipitation experiments we identified the Hippo related kinase SvkA (Hrk-svk) as binding partner for CEP161. Both proteins colocalize at the centrosome. In in vitro kinase assays the N-terminal domain of CEP161 (residues 1-763) inhibited the kinase activity of Hrk-svk. A comparison of D. discoideum Hippo kinase mutants with mutants overexpressing CEP161 polypeptides revealed similar defects. We propose that the centrosomal component CEP161 is a novel player in the Hippo signaling pathway and affects various cellular properties through this interaction. PMID:25607232

  7. On the right path? Exploring the experiences and opinions of clinicians involved in developing and implementing HealthPathways Barwon.

    PubMed

    Mansfield, Sarah J; Quirk, Frances; von Treuer, Kathryn; Gill, Gerard

    2016-04-01

    The aims of this paper are to present the findings of a process evaluation exploring the experiences and opinions of clinicians who have been involved in the HealthPathways Barwon clinical workgroups and discuss implications for further development of the program, as well as regional health service initiatives more broadly. HealthPathways Barwon is a web-based program comprising locally agreed-upon evidence-based clinical pathways that assist with assessment, management and region-specific referral for various clinical conditions. Clinical workgroup members participated in focus groups. Coding and thematic analysis were performed and findings were compared with similar evaluations of HealthPathways in other jurisdictions. Five broad themes emerged from the focus group, each with several subthemes: (1) purpose of HealthPathways; (2) workgroup process; (3) barriers and facilitators to HealthPathways use; (4) impact of HealthPathways on clinical practice; and (5) measuring performance. Findings of particular interest were that the perceived drivers for implementation of HealthPathways Barwon are broad, HealthPathways Barwon is viewed positively by clinicians, the workgroup process itself has a positive impact on relationships between primary and secondary care clinicians, existing habits of clinicians are a major barrier to adoption of HealthPathways Barwon, the sustainability of HealthPathways Barwon is a concern and it is difficult to measure the outcomes of HealthPathways. Although HealthPathways Barwon is viewed positively by clinicians and is seen to have the potential to address many issues at the primary-secondary care interface, successful implementation and uptake will depend on buy-in from clinicians, as well as continuous evaluation to inform improved development and implementation. More broadly, health service initiatives like HealthPathways Barwon require longer-term certainty of funding and administration to become established and produce meaningful

  8. Assembly and release of infectious hepatitis C virus involving unusual organization of the secretory pathway

    PubMed Central

    Triyatni, Miriam; Berger, Edward A; Saunier, Bertrand

    2016-01-01

    AIM: To determine if calnexin (CANX), RAB1 and alpha-tubulin were involved in the production of hepatitis C virus (HCV) particles by baby hamster kidney-West Nile virus (BHK-WNV) cells. METHODS: Using a siRNA-based approach complemented with immuno-fluorescence confocal microscope and Western blot studies, we examined the roles of CANX, RAB1 and alpha-tubulin in the production of HCV particles by permissive BHK-WNV cells expressing HCV structural proteins or the full-length genome of HCV genotype 1a. Immuno-fluorescence studies in producer cells were performed with monoclonal antibodies against HCV structural proteins, as well as immunoglobulin from the serum of a patient recently cured from an HCV infection of same genotype. The cellular compartment stained by the serum immunoglobulin was also observed in thin section transmission electron microscopy. These findings were compared with the JFH-1 strain/Huh-7.5 cell model. RESULTS: We found that CANX was necessary for the production of HCV particles by BHK-WNV cells. This process involved the recruitment of a subset of HCV proteins, detected by immunoglobulin of an HCV-cured patient, in a compartment of rearranged membranes bypassing the endoplasmic reticulum-Golgi intermediary compartment and surrounded by mitochondria. It also involved the maturation of N-linked glycans on HCV envelope proteins, which was required for assembly and/or secretion of HCV particles. The formation of this specialized compartment required RAB1; upon expression of HCV structural genes, this compartment developed large vesicles with viral particles. RAB1 and alpha-tubulin were required for the release of HCV particles. These cellular factors were also involved in the production of HCVcc in the JFH-1 strain/Huh-7.5 cell system, which involves HCV RNA replication. The secretion of HCV particles by BHK-WNV cells presents similarities with a pathway involving caspase-1; a caspase-1 inhibitor was found to suppress the production of HCV

  9. Protective effect of relaxin in cardiac anaphylaxis: involvement of the nitric oxide pathway

    PubMed Central

    Masini, E; Zagli, G; Ndisang, J F; Solazzo, M; Mannaioni, P F; Bani, D

    2002-01-01

    Relaxin (RLX) is a multifunctional hormone best known for its role in pregnancy and parturition, that has been also shown to influence coronary perfusion and mast cell activation through the generation of endogenous nitric oxide (NO). In this study we report on the effects of RLX on the biochemical and mechanical changes of ex vivo perfused hearts isolated from ovalbumin-sensitized guinea-pigs induced by challenge with the specific antigen. The possible involvement of NO in the RLX action has been also investigated. A 30-min perfusion with RLX (30 ng ml−1) before ovalbumin challenge fully abated the positive chronotropic and inotropic effects evoked by anaphylactic reaction to the antigen. RLX also blunted the short-term coronary constriction following to antigen challenge. Conversely, perfusion with chemically inactivated RLX had no effect. The release of histamine in the perfusate and the accumulation of calcium in heart tissue induced by antigen challenge were significantly decreased by RLX, while the amounts of nitrites in the perfusate were significantly increased, as were NO synthase activity and expression and cGMP levels in heart tissue. These findings indicate that RLX has a protective effect in cardiac anaphylaxis which involves an up-regulation of the NO biosynthetic pathway. PMID:12237253

  10. Establishing an in vivo assay system to identify components involved in environmental RNA interference in the western corn rootworm.

    PubMed

    Miyata, Keita; Ramaseshadri, Parthasarathy; Zhang, Yuanji; Segers, Gerrit; Bolognesi, Renata; Tomoyasu, Yoshinori

    2014-01-01

    The discovery of environmental RNA interference (RNAi), in which gene expression is suppressed via feeding with double-stranded RNA (dsRNA) molecules, opened the door to the practical application of RNAi-based techniques in crop pest management. The western corn rootworm (WCR, Diabrotica virgifera virgifera) is one of the most devastating corn pests in North America. Interestingly, WCR displays a robust environmental RNAi response, raising the possibility of applying an RNAi-based pest management strategy to this pest. Understanding the molecular mechanisms involved in the WCR environmental RNAi process will allow for determining the rate limiting steps involved with dsRNA toxicity and potential dsRNA resistance mechanisms in WCR. In this study, we have established a two-step in vivo assay system, which allows us to evaluate the involvement of genes in environmental RNAi in WCR. We show that laccase 2 and ebony, critical cuticle pigmentation/tanning genes, can be used as marker genes in our assay system, with ebony being a more stable marker to monitor RNAi activity. In addition, we optimized the dsRNA dose and length for the assay, and confirmed that this assay system is sensitive to detect well-known RNAi components such as Dicer-2 and Argonaute-2. We also evaluated two WCR sid1- like (sil) genes with this assay system. This system will be useful to quickly survey candidate systemic RNAi genes in WCR, and also will be adaptable for a genome-wide RNAi screening to give us an unbiased view of the environmental/systemic RNAi pathway in WCR. PMID:25003334

  11. Properdin and C3 Proactivator: Alternate Pathway Components in Human Glomerulonephritis

    PubMed Central

    McLean, Robert H.; Michael, Alfred F.

    1973-01-01

    Serological and immunopathological studies of human glomerulonephritis have suggested that alternate pathways of activation of the third component of complement may be important in some forms of glomerulonephritis. We have investigated the role of two alternate pathway proteins, properdin and C3 proactivator, in 22 patients with chronic membranoproliferative glomerulonephritis, 21 patients with systemic lupus erythematosus, 20 patients with acute poststreptococcal glomerulonephritis, and 19 patients with other forms of renal disease. C3 (measured at β1A), properdin, and C3 proactivator were assayed by single radial immunodiffusion. In sera with low β1A (< 2 SD), mean properdin was most significantly decreased in patients with acute poststreptococcal glomerulonephritis but was also significantly decreased in chronic membranoproliferative glomerulonephritis and in untreated systemic lupus erythematosus. Properdin levels in other renal disease, acute glomerulonephritis, and chronic membranoproliferative glomerulonephritis with normal β1A levels were not significantly different from normal. A positive correlation between β1A and properdin levels in individual sera was present in all diseases except systemic lupus erythematosus. Serum C3 proactivator was markedly decreased in active systemic lupus erythematosus and there was a positive correlation between β1A and C3 proactivator levels in systemic lupus erythematosus and other renal diseases but not acute poststreptococcal glomerulonephritis. Properdin in fresh sera from four patients with systemic lupus erythematosus and five with chronic membranoproliferative glomerulonephritis showed increased migration toward the cathode on immunoelectrophoresis, suggesting in vivo change of the properdin molecule. The observation of reduced serum levels of properdin and C3 proactivator and altered electrophoretic migration of properdin in some patients with glomerulonephritis provide new evidence for participation of these

  12. Phenotypes associated with inherited and developmental somatic mutations in genes encoding mTOR pathway components.

    PubMed

    Saxena, Anurag; Sampson, Julian R

    2014-12-01

    Mutations affecting the genes that encode upstream components in the mammalian (or mechanistic) target of rapamycin signalling pathway are associated with a group of rare inherited and developmental disorders that show overlapping clinical features. These include predisposition to a variety of benign or malignant tumours, localized overgrowth, developmental abnormalities of the brain, neurodevelopmental disorders and epilepsy. Many of these features have been linked to hyperactivation of signalling via mammalian target of rapamycin complex 1, suggesting that inhibitors of this complex such as rapamycin and its derivatives may offer new opportunities for therapy. In this review we describe this group of inherited and developmental disorders and discuss recent progress in their treatment via mTORC1 inhibition. PMID:25263008

  13. Identification of signaling pathways in macrophage exposed to Porphyromonas gingivalis or to its purified cell wall components.

    PubMed

    Zhou, Qingde; Amar, Salomon

    2007-12-01

    Porphyromonas gingivalis (P. gingivalis) can trigger an inflammatory condition leading to the destruction of periodontal tissues. However P. gingivalis LPS and its fimbriae (FimA) play different roles compared with the live bacteria in the context of intracellular molecule induction and cytokine secretion. To elucidate whether this difference results from different signaling pathways in host immune response to P. gingivalis, its LPS, or its FimA, we examined gene expression profile of human macrophages exposed to P. gingivalis, its LPS, or its FimA. A comparison of gene expression resulted in the identification of three distinct groups of expressed genes. Furthermore, computer-assisted promoter analysis of a subset of each group of differentially regulated genes revealed four putative transcriptional regulation models that associate with transcription factors NFkappaB, IRF7, and KLF4. Using gene knockout mice and siRNA to silence mouse genes, we showed that both TLR2 and TLR7 are essential for the induction of NFkappaB-containing genes and NFkappaB-IFN-sensitive response element (ISRE) cocontaining genes by either P. gingivalis or its purified components. The gene induction via either TLR2 or TLR7 is dependent on both MyD88 and p38 MAPK. However, the unique induction of IFN-beta by P. gingivalis LPS requires TLR7 and IFNalphabetaR cosignaling, and the induction of ISRE-containing gene is dependent on the activation of IFN-beta autocrine loop. Taken together, these data demonstrate that P. gingivalis and its components induce NFkappaB-containing genes through either TLR2- or TLR7-MyD88-p38 MAPK pathway, while P. gingivalis LPS uniquely induces ISRE-containing genes, which requires IFNalphabetaR signaling involving IRF7, KLF4, and pY701 STAT1. PMID:18025224

  14. The heparan and heparin metabolism pathway is involved in regulation of fatty acid composition.

    PubMed

    Jiang, Zhihua; Michal, Jennifer J; Wu, Xiao-Lin; Pan, Zengxiang; MacNeil, Michael D

    2011-01-01

    Six genes involved in the heparan sulfate and heparin metabolism pathway, DSEL (dermatan sulfate epimerase-like), EXTL1 (exostoses (multiple)-like 1), HS6ST1 (heparan sulfate 6-O-sulfotransferase 1), HS6ST3 (heparan sulfate 6-O-sulfotransferase 3), NDST3 (N-deacetylase/N-sulfotransferase (heparan glucosaminyl) 3), and SULT1A1 (sulfotransferase family, cytosolic, 1A, phenol-preferring, member 1), were investigated for their associations with muscle lipid composition using cattle as a model organism. Nineteen single nucleotide polymorphisms (SNPs)/multiple nucleotide length polymorphisms (MNLPs) were identified in five of these six genes. Six of these mutations were then genotyped on 246 Wagyu x Limousin F(2) animals, which were measured for 5 carcass, 6 eating quality and 8 fatty acid composition traits. Association analysis revealed that DSEL, EXTL1 and HS6ST1 significantly affected two stearoyl-CoA desaturase activity indices, the amount of conjugated linoleic acid (CLA), and the relative amount of saturated fatty acids (SFA) and monounsaturated fatty acids (MUFA) in skeletal muscle (P<0.05). In particular, HS6ST1 joined our previously reported SCD1 and UQCRC1 genes to form a three gene network for one of the stearoyl-CoA desaturase activity indices. These results provide evidence that genes involved in heparan sulfate and heparin metabolism are also involved in regulation of lipid metabolism in bovine muscle. Whether the SNPs affected heparan sulfate proteoglycan structure is unknown and warrants further investigation. PMID:21647334

  15. Degradation of the antiviral component ARGONAUTE1 by the autophagy pathway

    PubMed Central

    Derrien, Benoît; Baumberger, Nicolas; Schepetilnikov, Mikhail; Viotti, Corrado; De Cillia, Julia; Ziegler-Graff, Véronique; Isono, Erika; Schumacher, Karin; Genschik, Pascal

    2012-01-01

    Posttranscriptional gene silencing (PTGS) mediated by siRNAs is an evolutionarily conserved antiviral defense mechanism in higher plants and invertebrates. In this mechanism, viral-derived siRNAs are incorporated into the RNA-induced silencing complex (RISC) to guide degradation of the corresponding viral RNAs. In Arabidopsis, a key component of RISC is ARGONAUTE1 (AGO1), which not only binds to siRNAs but also carries the RNA slicer activity. At present little is known about posttranslational mechanisms regulating AGO1 turnover. Here we report that the viral suppressor of RNA silencing protein P0 triggers AGO1 degradation by the autophagy pathway. Using a P0-inducible transgenic line, we observed that AGO1 degradation is blocked by inhibition of autophagy. The engineering of a functional AGO1 fluorescent reporter protein further indicated that AGO1 colocalizes with autophagy-related (ATG) protein 8a (ATG8a) positive bodies when degradation is impaired. Moreover, this pathway also degrades AGO1 in a nonviral context, especially when the production of miRNAs is impaired. Our results demonstrate that a selective process such as ubiquitylation can lead to the degradation of a key regulatory protein such as AGO1 by a degradation process generally believed to be unspecific. We anticipate that this mechanism will not only lead to degradation of AGO1 but also of its associated proteins and eventually small RNAs. PMID:23019378

  16. Systemic Lupus Erythematosus and Deficiencies of Early Components of the Complement Classical Pathway

    PubMed Central

    Macedo, Ana Catarina Lunz; Isaac, Lourdes

    2016-01-01

    The complement system plays an important role in the innate and acquired immune response against pathogens. It consists of more than 30 proteins found in soluble form or attached to cell membranes. Most complement proteins circulate in inactive forms and can be sequentially activated by the classical, alternative, or lectin pathways. Biological functions, such as opsonization, removal of apoptotic cells, adjuvant function, activation of B lymphocytes, degranulation of mast cells and basophils, and solubilization and clearance of immune complex and cell lysis, are dependent on complement activation. Although the activation of the complement system is important to avoid infections, it also can contribute to the inflammatory response triggered by immune complex deposition in tissues in autoimmune diseases. Paradoxically, the deficiency of early complement proteins from the classical pathway (CP) is strongly associated with development of systemic lupus erythematous (SLE) – mainly C1q deficiency (93%) and C4 deficiency (75%). The aim of this review is to focus on the deficiencies of early components of the CP (C1q, C1r, C1s, C4, and C2) proteins in SLE patients. PMID:26941740

  17. A 2-component system is involved in the early stages of the Pisolithus tinctorius-Pinus greggii symbiosis

    PubMed Central

    Herrera-Martínez, Aseneth; Ruiz-Medrano, Roberto; Galván-Gordillo, Santiago Valentín; Toscano-Morales, Roberto; Gómez-Silva, Lidia; Valdés, María; Hinojosa-Moya, Jesús; Xoconostle-Cázares, Beatriz

    2014-01-01

    Ectomycorrhizal symbiosis results in profound morphological and physiological modifications in both plant and fungus. This in turn is the product of differential gene expression in both co-symbionts, giving rise to specialized cell types capable of performing novel functions. During the precolonization stage, chemical signals from root exudates are sensed by the ectomycorrhizal fungus, and vice versa, which are in principle responsible for the observed change in the symbionts developmental program. Little is known about the molecular mechanisms involved in the signaling and recognition between ectomycorrhizal fungi and their host plants. In the present work, we characterized a novel lactone, termed pinelactone, and identified a gene encoding for a histidine kinase in Pisolithus tictorius, the function of which is proposed to be the perception of the aforementioned metabolites. In this study, the use of closantel, a specific inhibitor of histidine kinase phosphorylation, affected the capacity for fungal colonization in the symbiosis between Pisolithus tinctorius and Pinus greggii, indicating that a 2-component system (TCS) may operate in the early events of plant-fungus interaction. Indeed, the metabolites induced the accumulation of Pisolithus tinctorius mRNA for a putative histidine kinase (termed Pthik1). Of note, Pthik1 was able to partially complement a S. cerevisiae histidine kinase mutant, demonstrating its role in the response to the presence of these metabolites. Our results indicate a role of a TCS pathway in the early stages of ectomycorrhizal symbiosis before colonization. Furthermore, a novel lactone from Pinus greggii root exudates may activate a signal transduction pathway that contributes to the establishment of the ectomycorrhizal symbiosis. PMID:24704731

  18. Involvement of ER stress and activation of apoptotic pathways in fisetin induced cytotoxicity in human melanoma.

    PubMed

    Syed, Deeba N; Lall, Rahul K; Chamcheu, Jean Christopher; Haidar, Omar; Mukhtar, Hasan

    2014-12-01

    The prognosis of malignant melanoma remains poor in spite of recent advances in therapeutic strategies for the deadly disease. Fisetin, a dietary flavonoid is currently being investigated for its growth inhibitory properties in various cancer models. We previously showed that fisetin inhibited melanoma growth in vitro and in vivo. Here, we evaluated the molecular basis of fisetin induced cytotoxicity in metastatic human melanoma cells. Fisetin treatment induced endoplasmic reticulum (ER) stress in highly aggressive A375 and 451Lu human melanoma cells, as revealed by up-regulation of ER stress markers including IRE1α, XBP1s, ATF4 and GRP78. Time course analysis indicated that the ER stress was associated with activation of the extrinsic and intrinsic apoptotic pathways. Fisetin treated 2-D melanoma cultures displayed autophagic response concomitant with induction of apoptosis. Prolonged treatment (16days) with fisetin in a 3-D reconstituted melanoma model resulted in inhibition of melanoma progression with significant apoptosis, as evidenced by increased staining of cleaved Caspase-3 in the treated constructs. However, no difference in the expression of autophagic marker LC-3 was noted between treated and control groups. Fisetin treatment to 2-D melanoma cultures resulted in phosphorylation and activation of the multifunctional AMP-activated protein kinase (AMPK) involved in the regulation of diverse cellular processes, including autophagy and apoptosis. Silencing of AMPK failed to prevent cell death indicating that fisetin induced cytotoxicity is mediated through both AMPK-dependent and -independent mechanisms. Taken together, our studies confirm apoptosis as the primary mechanism through which fisetin inhibits melanoma cell growth and that activation of both extrinsic and intrinsic pathways contributes to fisetin induced cytotoxicity. PMID:25016296

  19. Alpha-synuclein aggregation involves a bafilomycin A1-sensitive autophagy pathway

    PubMed Central

    Klucken, Jochen; Poehler, Anne-Maria; Ebrahimi-Fakhari, Darius; Schneider, Jacqueline; Nuber, Silke; Rockenstein, Edward; Schlötzer-Schrehardt, Ursula; Hyman, Bradley T.; McLean, Pamela J.; Masliah, Eliezer; Winkler, Juergen

    2012-01-01

    Synucleinopathies like Parkinson disease and dementia with Lewy bodies (DLB) are characterized by α-synuclein aggregates within neurons (Lewy bodies) and their processes (Lewy neurites). Whereas α-synuclein has been genetically linked to the disease process, the pathological relevance of α-synuclein aggregates is still debated. Impaired degradation is considered to result in aggregation of α-synuclein. In addition to the ubiquitin-proteasome degradation, the autophagy-lysosomal pathway (ALP) is involved in intracellular degradation processes for α-synuclein. Here, we asked if modulation of ALP affects α-synuclein aggregation and toxicity. We have identified an induction of the ALP markers LAMP-2A and LC3-II in human brain tissue from DLB patients, in a transgenic mouse model of synucleinopathy, and in a cell culture model for α-synuclein aggregation. ALP inhibition using bafilomycin A1 (BafA1) significantly potentiates toxicity of aggregated α-synuclein species in transgenic mice and in cell culture. Surprisingly, increased toxicity is paralleled by reduced aggregation in both in vivo and in vitro models. The dichotomy of effects on aggregating and nonaggregating species of α-synuclein was specifically sensitive to BafA1 and could not be reproduced by other ALP inhibitors. The present study expands on the accumulating evidence regarding the function of ALP for α-synuclein degradation by isolating an aggregation specific, BafA1-sensitive, ALP-related pathway. Our data also suggest that protein aggregation may represent a detoxifying event rather than being causal for cellular toxicity. PMID:22647715

  20. Alpha-synuclein aggregation involves a bafilomycin A 1-sensitive autophagy pathway.

    PubMed

    Klucken, Jochen; Poehler, Anne-Maria; Ebrahimi-Fakhari, Darius; Schneider, Jacqueline; Nuber, Silke; Rockenstein, Edward; Schlötzer-Schrehardt, Ursula; Hyman, Bradley T; McLean, Pamela J; Masliah, Eliezer; Winkler, Juergen

    2012-05-01

    Synucleinopathies like Parkinson disease and dementia with Lewy bodies (DLB) are characterized by α-synuclein aggregates within neurons (Lewy bodies) and their processes (Lewy neurites). Whereas α-synuclein has been genetically linked to the disease process, the pathological relevance of α-synuclein aggregates is still debated. Impaired degradation is considered to result in aggregation of α-synuclein. In addition to the ubiquitin-proteasome degradation, the autophagy-lysosomal pathway (ALP) is involved in intracellular degradation processes for α-synuclein. Here, we asked if modulation of ALP affects α-synuclein aggregation and toxicity. We have identified an induction of the ALP markers LAMP-2A and LC3-II in human brain tissue from DLB patients, in a transgenic mouse model of synucleinopathy, and in a cell culture model for α-synuclein aggregation. ALP inhibition using bafilomycin A 1 (BafA1) significantly potentiates toxicity of aggregated α-synuclein species in transgenic mice and in cell culture. Surprisingly, increased toxicity is paralleled by reduced aggregation in both in vivo and in vitro models. The dichotomy of effects on aggregating and nonaggregating species of α-synuclein was specifically sensitive to BafA1 and could not be reproduced by other ALP inhibitors. The present study expands on the accumulating evidence regarding the function of ALP for α-synuclein degradation by isolating an aggregation specific, BafA1-sensitive, ALP-related pathway. Our data also suggest that protein aggregation may represent a detoxifying event rather than being causal for cellular toxicity. PMID:22647715

  1. atRA-induced apoptosis of mouse embryonic palate mesenchymal cells involves activation of MAPK pathway

    SciTech Connect

    Yu Zengli . E-mail: yuzengli@263.net; Xing Ying . E-mail: xingy@zzu.edu.cn

    2006-08-15

    Our previous studies have shown that atRA treatment resulted in cell-cycle block and growth inhibition in mouse embryonic palatal mesenchymal (MEPM). In the current study, gestation day (GD) 13 MEPM cells were used to test the hypothesis that the growth inhibition by atRA is due to apoptosis. The effects of atRA on apoptosis were assessed by performing MTT assay, Cell Death Detection ELISA and flow cytometry, respectively. Data analysis confirmed that atRA treatment induced apoptosis-like cell death, as shown by decreased cell viability and increased fragmented DNA and sub-G1 fraction. atRA-induced apoptosis was associated with upregulation of bcl-2, translocation of bax protein to the mitochondria from the cytosol, activation of caspase-3 and cytochrome c release into cytosol. atRA-induced apoptosis was abrogated by z-DEVD-fmk, a caspase-3 specific inhibitor, and z-VAD-fmk, a general caspase inhibitor, suggesting that the atRA-induced cell death of MEPM cells occurs through the cytochrome c- and caspase-3-dependent pathways. In addition, atRA treatment caused a strong and sustained activation of c-Jun N-terminal kinase (JNK) and p38 kinase (p38), as well as an early but transient activation of extracellular signal-regulated kinase (ERK). Importantly, atRA-induced DNA fragmentation and capase-3 activation were prevented by pretreatment with the JNK inhibitor (SP600125) and the p38 MAPK inhibitor (SB202190), but not by pretreatment with MEK inhibitor (U0126). From these results, we suggest that mitogen-activated protein kinase-dependent pathways is involved in the atRA-induced apoptosis of MEPM cells.

  2. Involvement of the Notch pathway in terminal astrocytic differentiation: role of PKA

    PubMed Central

    Angulo-Rojo, Carla; Manning-Cela, Rebeca; Aguirre, Adán; Ortega, Arturo; López-Bayghen, Esther

    2013-01-01

    The Notch pathway is a highly conserved signaling system essential for modulating neurogenesis and promoting astrogenesis. Similarly, the cAMP signaling cascade can promote astrocytic commitment in several cell culture models, such as the C6 glioma cell line. These cells have the capacity to differentiate into oligodendrocytes or astrocytes, characteristics that allow their use as a glial progenitor model. In this context, we explore here the plausible involvement of cAMP in Notch-dependent signal transactions. The exposure of C6 cells to a non-hydrolysable cAMP analogue resulted in a sustained augmentation of Notch activity, as detected by nuclear translocation of its intracellular domain portion (NICD) and transcriptional activity. The cAMP effect is mediated through the activation of the γ-secretase complex, responsible for Notch cleavage and is sensitive to inhibitors of the cAMP-dependent protein kinase, PKA. As expected, Notch cleavage and nuclear translocation resulted in the up-regulation of the mRNA levels of one of its target genes, the transcription factor Hair and enhancer of split 5. Moreover, the glutamate uptake activity, as well as the expression of astrocytic markers such as glial fibrillary acidic protein, S100β protein and GLAST was also enhanced in cAMP-exposed cells. Our results clearly suggest that during the process of C6 astrocytic differentiation, cAMP activates the PKA/γ-secretase/NICD/RBPJκ pathway and Notch1 expression, leading to transcriptional activation of the genes responsible for glial progenitor cell fate decision. PMID:24286475

  3. Involvement of ER stress and activation of apoptotic pathways in fisetin induced cytotoxicity in human melanoma

    PubMed Central

    Chamcheu, Jean Christopher; Haidar, Omar; Mukhtar, Hasan

    2014-01-01

    The prognosis of malignant melanoma remains poor in spite of recent advances in therapeutic strategies for the deadly disease. Fisetin, a dietary flavonoid is currently being investigated for its growth inhibitory properties in various cancer models. We previously showed that fisetin inhibited melanoma growth in vitro and in vivo. Here, we evaluated the molecular basis of fisetin induced cytoxicity in metastatic human melanoma cells. Fisetin treatment induced endoplasmic reticulum (ER) stress in highly aggressive A375 and 451Lu human melanoma cells, as revealed by up- regulation of ER stress markers including IRE1α, XBP1s, ATF4 and GRP78. Time course analysis indicated that the ER stress was associated with activation of the extrinsic and intrinsic apoptotic pathways. Fisetin treated 2-D melanoma cultures displayed autophagic response concomitant with induction of apoptosis. Prolonged treatment (16 days) with fisetin in a 3-D reconstituted melanoma model resulted in inhibition of melanoma progression with significant apoptosis, as evidenced by increased staining of cleaved Caspase-3 in the treated constructs. However, no difference in the expression of autophagic marker LC-3 was noted between treated and control groups. Fisetin treatment to 2-D melanoma cultures resulted in phosphorylation and activation of the multifunctional AMPK-activated protein kinase (AMPK) involved in the regulation of diverse cellular processes, including autophagy and apoptosis. Silencing of AMPK failed to prevent cell death indicating that fisetin induced cytotoxicity is mediated through both AMPK-dependent and -independent mechanisms. Taken together, our studies confirm apoptosis as the primary mechanism through which fisetin inhibits melanoma cell growth and that activation of both extrinsic and intrinsic pathways contributes to fisetin induced cytotoxicity. PMID:25016296

  4. Plk1-dependent recruitment of gamma-tubulin complexes to mitotic centrosomes involves multiple PCM components.

    PubMed

    Haren, Laurence; Stearns, Tim; Lüders, Jens

    2009-01-01

    The nucleation of microtubules requires protein complexes containing gamma-tubulin, which are present in the cytoplasm and associate with the centrosome and with the mitotic spindle. We have previously shown that these interactions require the gamma-tubulin targeting factor GCP-WD/NEDD1, which has an essential role in spindle formation. The recruitment of additional gamma-tubulin to the centrosomes occurs during centrosome maturation at the G2/M transition and is regulated by the mitotic kinase Plk1. However, the molecular details of this important pathway are unknown and a Plk1 substrate that controls gamma-tubulin recruitment has not been identified. Here we show that Plk1 associates with GCP-WD in mitosis and Plk1 activity contributes to phosphorylation of GCP-WD. Plk1 depletion or inhibition prevents accumulation of GCP-WD at mitotic centrosomes, but GCP-WD mutants that are defective in Plk1-binding and -phosphorylation still accumulate at mitotic centrosomes and recruit gamma-tubulin. Moreover, Plk1 also controls the recruitment of other PCM proteins implicated in centrosomal gamma-tubulin attachment (Cep192/hSPD2, pericentrin, Cep215/Cdk5Rap2). Our results support a model in which Plk1-dependent recruitment of gamma-tubulin to mitotic centrosomes is regulated upstream of GCP-WD, involves multiple PCM proteins and therefore potentially multiple Plk1 substrates. PMID:19543530

  5. Predicting and exploring network components involved in pathogenesis in the malaria parasite via novel subnetwork alignments

    PubMed Central

    2015-01-01

    Background Malaria is a major health threat, affecting over 40% of the world's population. The latest report released by the World Health Organization estimated about 207 million cases of malaria infection, and about 627,000 deaths in 2012 alone. During the past decade, new therapeutic targets have been identified and are at various stages of characterization, thanks to the emerging omics-based technologies. However, the mechanism of malaria pathogenesis remains largely unknown. In this paper, we employ a novel neighborhood subnetwork alignment approach to identify network components that are potentially involved in pathogenesis. Results Our module-based subnetwork alignment approach identified 24 functional homologs of pathogenesis-related proteins in the malaria parasite P. falciparum, using the protein-protein interaction networks in Escherichia coli as references. Eighteen out of these 24 proteins are associated with 418 other proteins that are related to DNA replication, transcriptional regulation, translation, signaling, metabolism, cell cycle regulation, as well as cytoadherence and entry to the host. Conclusions The subnetwork alignments and subsequent protein-protein association network mining predicted a group of malarial proteins that may be involved in parasite development and parasite-host interaction, opening a new systems-level view of parasite pathogenesis and virulence. PMID:26100579

  6. Involvement of calcitonin gene-related peptide and receptor component protein in experimental autoimmune encephalomyelitis

    PubMed Central

    Sardi, Claudia; Zambusi, Laura; Finardi, Annamaria; Ruffini, Francesca; Tolun, Adviye A.; Dickerson, Ian M.; Righi, Marco; Zacchetti, Daniele; Grohovaz, Fabio; Provini, Luciano; Furlan, Roberto; Morara, Stefano

    2015-01-01

    Calcitonin Gene-Related Peptide (CGRP) inhibits microglia inflammatory activation in vitro. We here analyzed the involvement of CGRP and Receptor Component Protein (RCP) in experimental autoimmune encephalomyelitis (EAE). Alpha-CGRP deficiency increased EAE scores which followed the scale alpha-CGRP null > heterozygote > wild type. In wild type mice, CGRP delivery into the cerebrospinal fluid (CSF) 1) reduced chronic EAE (C-EAE) signs, 2) inhibited microglia activation (revealed by quantitative shape analysis), and 3) did not alter GFAP expression, cell density, lymphocyte infiltration, and peripheral lymphocyte production of IFN-gamma, TNF-alpha, IL-17, IL-2, and IL-4. RCP (probe for receptor involvement) was expressed in white matter microglia, astrocytes, oligodendrocytes, and vascular-endothelial cells: in EAE, also in infiltrating lymphocytes. In relapsing–remitting EAE (R-EAE) RCP increased during relapse, without correlation with lymphocyte density. RCP nuclear localization (stimulated by CGRP in vitro) was I) increased in microglia and decreased in astrocytes (R-EAE), and II) increased in microglia by CGRP CSF delivery (C-EAE). Calcitonin like receptor was rarely localized in nuclei of control and relapse mice. CGRP increased in motoneurons. In conclusion, CGRP can inhibit microglia activation in vivo in EAE. CGRP and its receptor may represent novel protective factors in EAE, apparently acting through the differential cell-specific intracellular translocationof RCP. PMID:24746422

  7. Two-component systems in Streptomyces: key regulators of antibiotic complex pathways

    PubMed Central

    2013-01-01

    Streptomyces, the main antibiotic-producing bacteria, responds to changing environmental conditions through a complex sensing mechanism and two-component systems (TCSs) play a crucial role in this extraordinary “sensing” device. Moreover, TCSs are involved in the biosynthetic control of a wide range of secondary metabolites, among them commercial antibiotics. Increased knowledge about TCSs can be a powerful asset in the manipulation of bacteria through genetic engineering with a view to obtaining higher efficiencies in secondary metabolite production. In this review we summarise the available information about Streptomyces TCSs, focusing specifically on their connections to antibiotic production. PMID:24354561

  8. Involvement of the MEK/ERK pathway in EGF-induced E-cadherin down-regulation.

    PubMed

    Tashiro, Etsu; Henmi, Shizuka; Odake, Hiroyuki; Ino, Seitaro; Imoto, Masaya

    2016-09-01

    E-cadherin is a major component of the epithelial adherens junction. However, the regulatory mechanism of E-cadherin expression is still poorly understood. In this study, we found that EGF decreased E-cadherin expression at both mRNA and protein levels in colorectal carcinoma LoVo cells. Since E-cadherin down-regulation is a well-known hallmark of the EMT (Epithelial-Mesenchymal Transition), we investigated whether EGF induced E-cadherin down-regulation during the EMT. EGF was unable to affect the expression of mesenchymal markers (such as N-cadherin, vimentin or fibronectin) or EMT-regulating transcription factors (such as SNAIL, SLUG, ZEB1, ZEB2 or TWIST), suggesting that EGF induced E-cadherin down-regulation via an EMT-independent mechanism. On the other hand, the MEK inhibitor U0126 was found to suppress EGF-induced E-cadherin down-regulation at the transcriptional level, suggesting that the MEK/ERK pathway is involved in EGF-induced E-cadherin down-regulation. Moreover, we also found that EGF disrupted cell-cell contact, stimulated cells to form an elongated shape with filamentous protrusions, and induced cell migration in LoVo cells. These effects were suppressed by U0126. Therefore, EGF is suggested to induce E-cadherin down-regulation at the transcriptional level through the MEK/ERK pathway, which might result in, at least in part, the induction of cellular morphological changes and cell migration in LoVo cells. PMID:27369075

  9. Involvement of SH2-containing phosphotyrosine phosphatase Syp in erythropoietin receptor signal transduction pathways.

    PubMed

    Tauchi, T; Feng, G S; Shen, R; Hoatlin, M; Bagby, G C; Kabat, D; Lu, L; Broxmeyer, H E

    1995-03-10

    Erythropoietin (Epo) regulates the proliferation and differentiation of erythroid precursors. The phosphorylation of proteins at tyrosine residues is critical in the growth signaling induced by Epo. This mechanism is regulated by the activities of both protein-tyrosine kinases and protein tyrosine phosphatases. The discovery of phosphotyrosine phosphatases that contain SH2 domains suggests roles for these molecules in growth factor signaling pathways. We found that Syp, a phosphotyrosine phosphatase, widely expressed in all tissues in mammals became phosphorylated on tyrosine after stimulation with Epo in M07ER cells engineered to express high levels of human EpoR. Syp was complexed with Grb2 in Epo-stimulated M07ER cells. Direct binding between Syp and Grb2 was also observed in vitro. Furthermore, Syp appeared to bind directly to tyrosine-phosphorylated EpoR in M07ER cells. Both NH2-terminal and COOH-terminal SH2 domains of Syp, made as glutathione S-transferase fusion proteins, were able to bind to the tyrosine-phosphorylated EpoR in vitro. These results suggest that Syp may be an important signaling component downstream of the EpoR and may regulate the proliferation and differentiation of hematopoietic cells. PMID:7534299

  10. Transcriptome Characterization by RNA-Seq Reveals the Involvement of the Complement Components in Noise-Traumatized Rat Cochleae

    PubMed Central

    Patel, Minal; Hu, Zihua; Bard, Jonathan; Jamison, Jennifer; Cai, Qunfeng; Hu, Bo Hua

    2013-01-01

    Acoustic trauma, a leading cause of sensorineural hearing loss in adults, induces a complex degenerative process in the cochlea. Although previous investigations have identified multiple stress pathways, a comprehensive analysis of cochlear responses to acoustic injury is still lacking. In the current study, we used the next-generation RNA-sequencing (RNA-seq) technique to sequence the whole transcriptome of the normal and noise-traumatized cochlear sensory epithelia (CSE). CSE tissues were collected from rat inner ears 1 d after the rats were exposed to a 120-dB (sound pressure level) noise for 2 h. The RNA-seq generated over 176 million sequence reads for the normal CSE and over 164 million reads for the noise-traumatized CSE. Alignment of these sequences with the rat Rn4 genome revealed the expression of over 17000 gene transcripts in the CSE, over 2000 of which were exclusively expressed in either the normal or noise-traumatized CSE. Seventy-eight gene transcripts were differentially expressed (70 upregulated and 8 downregulated) after acoustic trauma. Many of the differentially expressed genes are related to the innate immune system. Further expression analyses using qRT-PCR confirmed the constitutive expression of multiple complement genes in the normal organ of Corti and the changes in the expression levels of the complement factor I (Cfi) and complement component 1, s subcomponent (C1s) after acoustic trauma. Moreover, protein expression analysis revealed strong expression of Cfi and C1s proteins in the organ of Corti. Importantly, these proteins exhibited expression changes following acoustic trauma. Collectively, the results of the current investigation suggest the involvement of the complement components in cochlear responses to acoustic trauma. PMID:23727008

  11. Early Components of the Complement Classical Activation Pathway in Human Systemic Autoimmune Diseases

    PubMed Central

    Lintner, Katherine E.; Wu, Yee Ling; Yang, Yan; Spencer, Charles H.; Hauptmann, Georges; Hebert, Lee A.; Atkinson, John P.; Yu, C. Yung

    2016-01-01

    The complement system consists of effector proteins, regulators, and receptors that participate in host defense against pathogens. Activation of the complement system, via the classical pathway (CP), has long been recognized in immune complex-mediated tissue injury, most notably systemic lupus erythematosus (SLE). Paradoxically, a complete deficiency of an early component of the CP, as evidenced by homozygous genetic deficiencies reported in human, are strongly associated with the risk of developing SLE or a lupus-like disease. Similarly, isotype deficiency attributable to a gene copy-number (GCN) variation and/or the presence of autoantibodies directed against a CP component or a regulatory protein that result in an acquired deficiency are relatively common in SLE patients. Applying accurate assay methodologies with rigorous data validations, low GCNs of total C4, and heterozygous and homozygous deficiencies of C4A have been shown as medium to large effect size risk factors, while high copy numbers of total C4 or C4A as prevalent protective factors, of European and East-Asian SLE. Here, we summarize the current knowledge related to genetic deficiency and insufficiency, and acquired protein deficiencies for C1q, C1r, C1s, C4A/C4B, and C2 in disease pathogenesis and prognosis of SLE, and, briefly, for other systemic autoimmune diseases. As the complement system is increasingly found to be associated with autoimmune diseases and immune-mediated diseases, it has become an attractive therapeutic target. We highlight the recent developments and offer a balanced perspective concerning future investigations and therapeutic applications with a focus on early components of the CP in human systemic autoimmune diseases. PMID:26913032

  12. INVOLVEMENT OF PEPTIDOGLYCAN RECOGNITION PROTEIN L6 IN ACTIVATION OF IMMUNE DEFICIENCY PATHWAY IN THE IMMUNE RESPONSIVE SILKWORM CELLS.

    PubMed

    Tanaka, Hiromitsu; Sagisaka, Aki

    2016-06-01

    The immune deficiency (Imd) signaling pathway is activated by Gram-negative bacteria for producing antimicrobial peptides (AMPs). In Drosophila melanogaster, the activation of this pathway is initiated by the recognition of Gram-negative bacteria by peptidoglycan (PGN) recognition proteins (PGRPs), PGRP-LC and PGRP-LE. In this study, we found that the Imd pathway is involved in enhancing the promoter activity of AMP gene in response to Gram-negative bacteria or diaminopimelic (DAP) type PGNs derived from Gram-negative bacteria in an immune responsive silkworm cell line, Bm-NIAS-aff3. Using gene knockdown experiments, we further demonstrated that silkworm PGRP L6 (BmPGRP-L6) is involved in the activation of E. coli or E. coli-PGN mediated AMP promoter activation. Domain analysis revealed that BmPGRP-L6 contained a conserved PGRP domain, transmembrane domain, and RIP homotypic interaction motif like motif but lacked signal peptide sequences. BmPGRP-L6 overexpression enhances AMP promoter activity through the Imd pathway. BmPGRP-L6 binds to DAP-type PGNs, although it also binds to lysine-type PGNs that activate another immune signal pathway, the Toll pathway in Drosophila. These results indicate that BmPGRP-L6 is a key PGRP for activating the Imd pathway in immune responsive silkworm cells. PMID:26991439

  13. Changes in actin dynamics are involved in salicylic acid signaling pathway.

    PubMed

    Matoušková, Jindřiška; Janda, Martin; Fišer, Radovan; Sašek, Vladimír; Kocourková, Daniela; Burketová, Lenka; Dušková, Jiřina; Martinec, Jan; Valentová, Olga

    2014-06-01

    Changes in actin cytoskeleton dynamics are one of the crucial players in many physiological as well as non-physiological processes in plant cells. Positioning of actin filament arrays is necessary for successful establishment of primary lines of defense toward pathogen attack, depolymerization leads very often to the enhanced susceptibility to the invading pathogen. On the other hand it was also shown that the disruption of actin cytoskeleton leads to the induction of defense response leading to the expression of PATHOGENESIS RELATED proteins (PR). In this study we show that pharmacological actin depolymerization leads to the specific induction of genes in salicylic acid pathway but not that involved in jasmonic acid signaling. Life imaging of leafs of Arabidopsis thaliana with GFP-tagged fimbrin (GFP-fABD2) treated with 1 mM salicylic acid revealed rapid disruption of actin filaments resembling the pattern viewed after treatment with 200 nM latrunculin B. The effect of salicylic acid on actin filament fragmentation was prevented by exogenous addition of phosphatidic acid, which binds to the capping protein and thus promotes actin polymerization. The quantitative evaluation of actin filament dynamics is also presented. PMID:24767113

  14. Molecular pathway of near-infrared laser phototoxicity involves ATF-4 orchestrated ER stress.

    PubMed

    Khan, Imran; Tang, Elieza; Arany, Praveen

    2015-01-01

    High power lasers are used extensively in medicine while lower power applications are popular for optical imaging, optogenetics, skin rejuvenation and a therapeutic modality termed photobiomodulation (PBM). This study addresses the therapeutic dose limits, biological safety and molecular pathway of near-infrared (NIR) laser phototoxicity. Increased erythema and tissue damage were noted in mice skin and cytotoxicity in cell cultures at phototoxic laser doses involving generation of reactive oxygen species (ROS) coupled with a rise in surface temperature (>45 °C). NIR laser phototoxicity results from Activating Transcription Factor-4 (ATF-4) mediated endoplasmic reticulum stress and autophagy. Neutralizations of heat or ROS and overexpressing ATF-4 were noted to rescue NIR laser phototoxicity. Further, NIR laser mediated phototoxicity was noted to be non-genotoxic and non-mutagenic. This study outlines the mechanism of NIR laser phototoxicity and the utility of monitoring surface temperature and ATF4 expression as potential biomarkers to develop safe and effective clinical applications. PMID:26030745

  15. Molecular pathway of near-infrared laser phototoxicity involves ATF-4 orchestrated ER stress

    PubMed Central

    Khan, Imran; Tang, Elieza; Arany, Praveen

    2015-01-01

    High power lasers are used extensively in medicine while lower power applications are popular for optical imaging, optogenetics, skin rejuvenation and a therapeutic modality termed photobiomodulation (PBM). This study addresses the therapeutic dose limits, biological safety and molecular pathway of near-infrared (NIR) laser phototoxicity. Increased erythema and tissue damage were noted in mice skin and cytotoxicity in cell cultures at phototoxic laser doses involving generation of reactive oxygen species (ROS) coupled with a rise in surface temperature (>45 °C). NIR laser phototoxicity results from Activating Transcription Factor-4 (ATF-4) mediated endoplasmic reticulum stress and autophagy. Neutralizations of heat or ROS and overexpressing ATF-4 were noted to rescue NIR laser phototoxicity. Further, NIR laser mediated phototoxicity was noted to be non-genotoxic and non-mutagenic. This study outlines the mechanism of NIR laser phototoxicity and the utility of monitoring surface temperature and ATF4 expression as potential biomarkers to develop safe and effective clinical applications. PMID:26030745

  16. The GABAergic septohippocampal pathway is directly involved in internal processes related to operant reward learning.

    PubMed

    Vega-Flores, Germán; Rubio, Sara E; Jurado-Parras, M Teresa; Gómez-Climent, María Ángeles; Hampe, Christiane S; Manto, Mario; Soriano, Eduardo; Pascual, Marta; Gruart, Agnès; Delgado-García, José M

    2014-08-01

    We studied the role of γ-aminobutyric acid (GABA)ergic septohippocampal projections in medial septum (MS) self-stimulation of behaving mice. Self-stimulation was evoked in wild-type (WT) mice using instrumental conditioning procedures and in J20 mutant mice, a type of mouse with a significant deficit in GABAergic septohippocampal projections. J20 mice showed a significant modification in hippocampal activities, including a different response for input/output curves and the paired-pulse test, a larger long-term potentiation (LTP), and a delayed acquisition and lower performance in the MS self-stimulation task. LTP evoked at the CA3-CA1 synapse further decreased self-stimulation performance in J20, but not in WT, mice. MS self-stimulation evoked a decrease in the amplitude of field excitatory postsynaptic potentials (fEPSPs) at the CA3-CA1 synapse in WT, but not in J20, mice. This self-stimulation-dependent decrease in the amplitude of fEPSPs was also observed in the presence of another positive reinforcer (food collected during an operant task) and was canceled by the local administration of an antibody-inhibiting glutamate decarboxylase 65 (GAD65). LTP evoked in the GAD65Ab-treated group was also larger than in controls. The hippocampus has a different susceptibility to septal GABAergic inputs depending on ongoing cognitive processes, and the GABAergic septohippocampal pathway is involved in consummatory processes related to operant rewards. PMID:23479403

  17. Nanometer Scale Titanium Surface Texturing Are Detected by Signaling Pathways Involving Transient FAK and Src Activations

    PubMed Central

    Zambuzzi, Willian F.; Bonfante, Estevam A.; Jimbo, Ryo; Hayashi, Mariko; Andersson, Martin; Alves, Gutemberg; Takamori, Esther R.; Beltrão, Paulo J.; Coelho, Paulo G.; Granjeiro, José M.

    2014-01-01

    Background It is known that physico/chemical alterations on biomaterial surfaces have the capability to modulate cellular behavior, affecting early tissue repair. Such surface modifications are aimed to improve early healing response and, clinically, offer the possibility to shorten the time from implant placement to functional loading. Since FAK and Src are intracellular proteins able to predict the quality of osteoblast adhesion, this study evaluated the osteoblast behavior in response to nanometer scale titanium surface texturing by monitoring FAK and Src phosphorylations. Methodology Four engineered titanium surfaces were used for the study: machined (M), dual acid-etched (DAA), resorbable media microblasted and acid-etched (MBAA), and acid-etch microblasted (AAMB). Surfaces were characterized by scanning electron microscopy, interferometry, atomic force microscopy, x-ray photoelectron spectroscopy and energy dispersive X-ray spectroscopy. Thereafter, those 4 samples were used to evaluate their cytotoxicity and interference on FAK and Src phosphorylations. Both Src and FAK were investigated by using specific antibody against specific phosphorylation sites. Principal Findings The results showed that both FAK and Src activations were differently modulated as a function of titanium surfaces physico/chemical configuration and protein adsorption. Conclusions It can be suggested that signaling pathways involving both FAK and Src could provide biomarkers to predict osteoblast adhesion onto different surfaces. PMID:24999733

  18. Immune complex stimulation of neutrophil apoptosis: investigating the involvement of oxidative and nonoxidative pathways.

    PubMed

    Ottonello, L; Frumento, G; Arduino, N; Dapino, P; Tortolina, G; Dallegri, F

    2001-01-15

    Neutrophils are involved in the pathogenesis of various inflammatory diseases. One of the mechanisms by which neutrophilic inflammation is generated is immune complex (IC) deposition in tissue. As the clearance of apoptotic neutrophils from inflamed sites is considered a crucial determinant for the resolution of inflammation, we investigated the effects of IC-induced neutrophil activation on apoptosis and the mechanisms regulating neutrophil survival. Our results show that IC stimulated apoptosis efficiently. The percentage of apoptotic neutrophils was reduced by the anti-FcgammaRII mAb IV.3, but not by anti-FcgammaRIII mAb 3G8. The spontaneous apoptosis was completely inhibited by the antioxidant compound catalase, which in turn prevented only partially the apoptosis in presence of IC. The oxidative metabolism triggered by IC was inhibited only blocking both FcgammaRII and FcgammaRIII. Neutrophils from patients with chronic granulomatous disease, congenitally incapable of producing oxidants, showed low level of spontaneous apoptosis, but underwent a nearly 3-fold increment in the apoptosis rate when incubated with IC. In conclusion, neutrophil apoptosis appears to be a process governed by multiple pathways, some of which are strictly ROS-dependent, others acting in a nonoxidative manner. In particular, the herein shown FcgammaRII-dependent, ROS-independent, signal-inducing neutrophil apoptosis may uncover new pharmacological targets for the promotion of cell removal from sites of inflammation, thereby favoring the resolution of the inflammatory process. PMID:11163533

  19. Stimulation of MC38 tumor growth by insulin analog X10 involves the serine synthesis pathway.

    PubMed

    Hvid, Henning; Fendt, Sarah-Maria; Blouin, Marie-José; Birman, Elena; Voisin, Gregory; Svendsen, Angela Manegold; Frank, Russell; Vander Heiden, Matthew G; Stephanopoulos, Gregory; Hansen, Bo Falck; Pollak, Michael

    2012-08-01

    Recent evidence suggests that type II diabetes is associated with increased risk and/or aggressive behavior of several cancers, including those arising from the colon. Concerns have been raised that endogenous hyperinsulinemia and/or exogenous insulin and insulin analogs might stimulate proliferation of neoplastic cells. However, the mechanisms underlying possible growth-promoting effects of insulin and insulin analogs in cancer cells in vivo, such as changes in gene expression, are incompletely described. We observed that administration of the insulin analog X10 significantly increased tumor growth and proliferation in a murine colon cancer model (MC38 cell allografts). Insulin and X10 altered gene expression in MC38 tumors in a similar fashion, but X10 was more potent in terms of the number of genes influenced and the magnitude of changes in gene expression. Many of the affected genes were annotated to metabolism, nutrient uptake, and protein synthesis. Strikingly, expression of genes encoding enzymes in the serine synthesis pathway, recently shown to be critical for neoplastic proliferation, was increased following treatment with insulin and X10. Using stable isotopic tracers and mass spectrometry, we confirmed that insulin and X10 increased glucose contribution to serine synthesis in MC38 cells. The data demonstrate that the tumor growth-promoting effects of insulin and X10 are associated with changes in expression of genes involved in cellular energy metabolism and reveal previously unrecognized effects of insulin and X10 on serine synthesis. PMID:22685267

  20. The major human AP endonuclease (Ape1) is involved in the nucleotide incision repair pathway

    PubMed Central

    Gros, Laurent; Ishchenko, Alexander A.; Ide, Hiroshi; Elder, Rhoderick H.; Saparbaev, Murat K.

    2004-01-01

    In nucleotide incision repair (NIR), an endonuclease nicks oxidatively damaged DNA in a DNA glycosylase-independent manner, providing the correct ends for DNA synthesis coupled to the repair of the remaining 5′-dangling modified nucleotide. This mechanistic feature is distinct from DNA glycosylase-mediated base excision repair. Here we report that Ape1, the major apurinic/apyrimidinic endonuclease in human cells, is the damage- specific endonuclease involved in NIR. We show that Ape1 incises DNA containing 5,6-dihydro-2′-deoxyuridine, 5,6-dihydrothymidine, 5-hydroxy-2′-deoxyuridine, alpha-2′-deoxyadenosine and alpha-thymidine adducts, generating 3′-hydroxyl and 5′-phosphate termini. The kinetic constants indicate that Ape1-catalysed NIR activity is highly efficient. The substrate specificity and protein conformation of Ape1 is modulated by MgCl2 concentrations, thus providing conditions under which NIR becomes a major activity in cell-free extracts. While the N-terminal region of Ape1 is not required for AP endonuclease function, we show that it regulates the NIR activity. The physiological relevance of the mammalian NIR pathway is discussed. PMID:14704345

  1. Nrf2/ARE Pathway Involved in Oxidative Stress Induced by Paraquat in Human Neural Progenitor Cells.

    PubMed

    Dou, Tingting; Yan, Mengling; Wang, Xinjin; Lu, Wen; Zhao, Lina; Lou, Dan; Wu, Chunhua; Chang, Xiuli; Zhou, Zhijun

    2016-01-01

    Compelling evidences have shown that diverse environmental insults arising during early life can either directly lead to a reduction in the number of dopaminergic neurons or cause an increased susceptibility to neurons degeneration with subsequent environmental insults or with aging alone. Oxidative stress is considered the main effect of neurotoxins exposure. In this study, we investigated the oxidative stress effect of Paraquat (PQ) on immortalized human embryonic neural progenitor cells by treating them with various concentrations of PQ. We show that PQ can decrease the activity of SOD and CAT but increase MDA and LDH level. Furthermore, the activities of Cyc and caspase-9 were found increased significantly at 10 μM of PQ treatment. The cytoplasmic Nrf2 protein expressions were upregulated at 10 μM but fell back at 100 μM. The nuclear Nrf2 protein expressions were upregulated as well as the downstream mRNA expressions of HO-1 and NQO1 in a dose-dependent manner. In addition, the proteins expression of PKC and CKII was also increased significantly even at 1 μM. The results suggested that Nrf2/ARE pathway is involved in mild to moderate PQ-induced oxidative stress which is evident from dampened Nrf2 activity and low expression of antioxidant genes in PQ induced oxidative damage. PMID:26649146

  2. The GABAergic Septohippocampal Pathway Is Directly Involved in Internal Processes Related to Operant Reward Learning

    PubMed Central

    Vega-Flores, Germán; Rubio, Sara E.; Jurado-Parras, M. Teresa; Gómez-Climent, María Ángeles; Hampe, Christiane S.; Manto, Mario; Soriano, Eduardo; Pascual, Marta; Gruart, Agnès; Delgado-García, José M.

    2014-01-01

    We studied the role of γ-aminobutyric acid (GABA)ergic septohippocampal projections in medial septum (MS) self-stimulation of behaving mice. Self-stimulation was evoked in wild-type (WT) mice using instrumental conditioning procedures and in J20 mutant mice, a type of mouse with a significant deficit in GABAergic septohippocampal projections. J20 mice showed a significant modification in hippocampal activities, including a different response for input/output curves and the paired-pulse test, a larger long-term potentiation (LTP), and a delayed acquisition and lower performance in the MS self-stimulation task. LTP evoked at the CA3–CA1 synapse further decreased self-stimulation performance in J20, but not in WT, mice. MS self-stimulation evoked a decrease in the amplitude of field excitatory postsynaptic potentials (fEPSPs) at the CA3–CA1 synapse in WT, but not in J20, mice. This self-stimulation-dependent decrease in the amplitude of fEPSPs was also observed in the presence of another positive reinforcer (food collected during an operant task) and was canceled by the local administration of an antibody-inhibiting glutamate decarboxylase 65 (GAD65). LTP evoked in the GAD65Ab-treated group was also larger than in controls. The hippocampus has a different susceptibility to septal GABAergic inputs depending on ongoing cognitive processes, and the GABAergic septohippocampal pathway is involved in consummatory processes related to operant rewards. PMID:23479403

  3. Gap Junctions in the Ventral Hippocampal-Medial Prefrontal Pathway Are Involved in Anxiety Regulation

    PubMed Central

    Schoenfeld, Timothy J.; Kloth, Alexander D.; Hsueh, Brian; Runkle, Matthew B.; Kane, Gary A.; Wang, Samuel S.-H.

    2014-01-01

    Anxiety disorders are highly prevalent but little is known about their underlying mechanisms. Gap junctions exist in brain regions important for anxiety regulation, such as the ventral hippocampus (vHIP) and mPFC, but their functions in these areas have not been investigated. Using pharmacological blockade of neuronal gap junctions combined with electrophysiological recordings, we found that gap junctions play a role in theta rhythm in the vHIP and mPFC of adult mice. Bilateral infusion of neuronal gap junction blockers into the vHIP decreased anxiety-like behavior on the elevated plus maze and open field. Similar anxiolytic effects were observed with unilateral infusion of these drugs into the vHIP combined with contralateral infusion into the mPFC. No change in anxious behavior was observed with gap junction blockade in the unilateral vHIP alone or in the bilateral dorsal HIP. Since physical exercise is known to reduce anxiety, we examined the effects of long-term running on the expression of the neuronal gap junction protein connexin-36 among inhibitory interneurons and found a reduction in the vHIP. Despite this change, we observed no alteration in theta frequency or power in long-term runners. Collectively, these findings suggest that neuronal gap junctions in the vHIP–mPFC pathway are important for theta rhythm and anxiety regulation under sedentary conditions but that additional mechanisms are likely involved in running-induced reduction in anxiety. PMID:25411496

  4. Involvement of an ABC transporter in a developmental pathway regulating hypocotyl cell elongation in the light

    PubMed Central

    Sidler, M; Hassa, P; Hasan, S; Ringli, C; Dudler, R

    1998-01-01

    In the dark, plant seedlings follow the skotomorphogenetic developmental program, which results in hypocotyl cell elongation. When the seedlings are exposed to light, a switch to photomorphogenetic development occurs, and hypocotyl cell elongation is inhibited. We have manipulated the expression of the AtPGP1 (for Arabidopsis thaliana P glycoprotein1) gene in transgenic Arabidopsis plants by using sense and antisense constructs. We show that within a certain light fluence rate window, overexpression of the AtPGP1 gene under the control of the cauliflower mosaic virus 35S promoter causes plants to develop longer hypocotyls, whereas expression of the gene in antisense orientation results in hypocotyls shorter than those occurring in the wild type. In the dark, hypocotyls of transgenic and wild-type plants are indistinguishable. Because the AtPGP1 gene encodes a member of the superfamily of ATP binding cassette-containing (ABC) transporters, these results imply that a transport process is involved in a hypocotyl cell elongation pathway active in the light. The AtPGP1 transporter is localized in the plasmalemma, as indicated by immunohistochemical techniques and biochemical membrane separation methods. Analysis of the AtPGP1 expression pattern by using reporter gene constructs and in situ hybridization shows that in wild-type seedlings, AtPGP1 is expressed in both the root and shoot apices. PMID:9761790

  5. The alternative respiratory pathway is involved in brassinosteroid-induced environmental stress tolerance in Nicotiana benthamiana

    PubMed Central

    Deng, Xing-Guang; Zhu, Tong; Zhang, Da-Wei; Lin, Hong-Hui

    2015-01-01

    Brassinosteroids (BRs), plant steroid hormones, play essential roles in modulating cell elongation, vascular differentiation, senescence, and stress responses. However, the mechanisms by which BRs regulate plant mitochondria and resistance to abiotic stress remain largely unclear. Mitochondrial alternative oxidase (AOX) is involved in the plant response to a variety of environmental stresses. In this report, the role of AOX in BR-induced tolerance against cold, polyethylene glycol (PEG), and high-light stresses was investigated. Exogenous applied brassinolide (BL, the most active BR) induced, while brassinazole (BRZ, a BR biosynthesis inhibitor) reduced alternative respiration and AOX1 expression in Nicotiana benthamiana. Chemical scavenging of H2O2 and virus-induced gene silencing (VIGS) of NbRBOHB compromised the BR-induced alternative respiratory pathway, and this result was further confirmed by NbAOX1 promoter analysis. Furthermore, inhibition of AOX activity by chemical treatment or a VIGS-based approach decreased plant resistance to environmental stresses and compromised BR-induced stress tolerance. Taken together, our results indicate that BR-induced AOX capability might contribute to the avoidance of superfluous reactive oxygen species accumulation and the protection of photosystems under stress conditions in N. benthamiana. PMID:26175355

  6. MicroRNAs Involved in Tumor Suppressor and Oncogene Pathways; Implications for Hepatobiliary Neoplasia

    PubMed Central

    Mott, Justin L.

    2009-01-01

    MicroRNAs are a class of small regulatory RNAs that function to modulate protein expression. This control allows for fine-tuning of the cellular phenotype, including regulation of proliferation, cell signaling, and apoptosis; not surprisingly, microRNAs contribute to liver cancer biology. Recent investigations in human liver cancers and tumor-derived cell lines have demonstrated decreased or increased expression of particular microRNAs in hepatobiliary cancer cells. Based on predicted and validated protein targets as well as functional consequences of altered expression, microRNAs with decreased expression in liver tumor cells may normally aid in limiting neoplastic transformation. Conversely, selected microRNAs that are upregulated in liver tumor cells can promote malignant features, contributing to carcinogenesis. In addition, microRNAs themselves are subject to regulated expression, including regulation by tumor suppressor and oncogene pathways. This review will focus on the expression and function of cancer-related microRNAs, including their intimate involvement in tumor suppressor and oncogene signaling networks relevant to hepatobiliary neoplasia. PMID:19585622

  7. Mineralocorticoid receptor is involved in the aldosterone pathway in human red blood cells

    PubMed Central

    Bordin, Luciana; Saccardi, Carlo; Donà, Gabriella; Sabbadin, Chiara; Andrisani, Alessandra; Ambrosini, Guido; Plebani, Mario; Brunati, Anna Maria; Ragazzi, Eugenio; Gizzo, Salvatore; Armanini, Decio

    2016-01-01

    We have recently demonstrated that excessive aldosterone (Aldo) secretion in primary aldosteronism (PA) is associated with red blood cells (RBC) senescence. These alterations were prevented/inhibited by cortisol (Cort) or canrenone (Can) raising the hypothesis that Aldo effects in RBC may be mediated by mineralocorticoid receptor (MR), though to date MR has never been demonstrated in human RBC. The aim of this multicenter comparative study was to investigate whether Aldo effects were mediated by MR in these a-nucleated cells. We included 12 healthy controls (HC) and 22 patients with PA. MR presence and activation were evaluated in RBC cytosol by glycerol gradient sedimentation, Western blotting, immuno-precipitation and radioimmunoassay. We demonstrated that RBC contained cytosolic MR, aggregated with HSP90 and other proteins to form multiprotein complex. Aldo induced MR to release from the complex and to form MR dimers which were quickly proteolyzed. Cort induced MR release but not dimers formation while Can was not able to induce MR release. In addition, RBC cytosol from PA patients contained significantly higher amounts of both MR fragments (p<0.0001) and Aldo (p<0.0001) concentrations. In conclusion, in RBC a genomic-like Aldo pathway is proposed involving MR activation, dimerization and proteolysis, but lacking nuclear transcription. In addition, dimers proteolysis may ensure a sort of Aldo scavenging from circulation by entrapping Aldo in MR fragments. PMID:27158328

  8. Mineralocorticoid receptor is involved in the aldosterone pathway in human red blood cells.

    PubMed

    Bordin, Luciana; Saccardi, Carlo; Donà, Gabriella; Sabbadin, Chiara; Andrisani, Alessandra; Ambrosini, Guido; Plebani, Mario; Brunati, Anna Maria; Ragazzi, Eugenio; Gizzo, Salvatore; Armanini, Decio

    2016-01-01

    We have recently demonstrated that excessive aldosterone (Aldo) secretion in primary aldosteronism (PA) is associated with red blood cells (RBC) senescence. These alterations were prevented/inhibited by cortisol (Cort) or canrenone (Can) raising the hypothesis that Aldo effects in RBC may be mediated by mineralocorticoid receptor (MR), though to date MR has never been demonstrated in human RBC. The aim of this multicenter comparative study was to investigate whether Aldo effects were mediated by MR in these a-nucleated cells. We included 12 healthy controls (HC) and 22 patients with PA. MR presence and activation were evaluated in RBC cytosol by glycerol gradient sedimentation, Western blotting, immuno-precipitation and radioimmunoassay. We demonstrated that RBC contained cytosolic MR, aggregated with HSP90 and other proteins to form multiprotein complex. Aldo induced MR to release from the complex and to form MR dimers which were quickly proteolyzed. Cort induced MR release but not dimers formation while Can was not able to induce MR release. In addition, RBC cytosol from PA patients contained significantly higher amounts of both MR fragments (p<0.0001) and Aldo (p<0.0001) concentrations. In conclusion, in RBC a genomic-like Aldo pathway is proposed involving MR activation, dimerization and proteolysis, but lacking nuclear transcription. In addition, dimers proteolysis may ensure a sort of Aldo scavenging from circulation by entrapping Aldo in MR fragments. PMID:27158328

  9. Influence of gold(I) complexes involving adenine derivatives on major drug-drug interaction pathway.

    PubMed

    Dvořák, Zdeněk; Novotná, Aneta; Vančo, Ján; Trávníček, Zdeněk

    2013-12-01

    A series of considerably anti-inflammatory active gold(I) mixed-ligand complexes, involving the benzyl-substituted derivatives of N6-benzyladenine (HLn) and triphenylphosphine (PPh3) as ligands and having the general formula [Au(Ln)(PPh3)]·xH2O (1-4; n=1-4 and x=0-1), was evaluated for the ability to influence the expression of CYP1A1/2 and CYP3A4 and transcriptional activity of glucocorticoid (GR) and aryl hydrocarbon (AhR) receptors in primary human hepatocytes and HepG2 cells. In both tests, evaluating the ability of the complexes to modulate the expression of CYP1A1, CYP1A2 and CYP3A4 in primary human hepatocytes and influence the transcriptional activity of AhR and GR in the reporter cell lines, no negative influence on the major drug-metabolizing cytochrome P450 isoenzymes and their signaling pathway (through GR and AhR receptors) was observed. These positive findings revealed another substantial evidence that could lead to utilization of the complexes as effective and relatively safe drugs for the treatment of hard-to-treat inflammation-related diseases, such as rheumatoid arthritis, comparable or even better than clinically used gold-containing drug Auranofin. PMID:24157406

  10. Evolution of threonine aldolases, a diverse family involved in the second pathway of glycine biosynthesis.

    PubMed

    Liu, Guangxiu; Zhang, Manxiao; Chen, Ximing; Zhang, Wei; Ding, Wei; Zhang, Qi

    2015-02-01

    Threonine aldolases (TAs) catalyze the interconversion of threonine and glycine plus acetaldehyde in a pyridoxal phosphate-dependent manner. This class of enzymes complements the primary glycine biosynthetic pathway catalyzed by serine hydroxymethyltransferase (SHMT), and was shown to be necessary for yeast glycine auxotrophy. Because the reverse reaction of TA involves carbon-carbon bond formation, resulting in a β-hydroxyl-α-amino acid with two adjacent chiral centers, TAs are of high interests in synthetic chemistry and bioengineering studies. Here, we report systematic phylogenetic analysis of TAs. Our results demonstrated that L-TAs and D-TAs that are specific for L- and D-threonine, respectively, are two phylogenetically unique families, and both enzymes are different from their closely related enzymes SHMTs and bacterial alanine racemases (ARs). Interestingly, L-TAs can be further grouped into two evolutionarily distinct families, which share low sequence similarity with each other but likely possess the same structural fold, suggesting a convergent evolution of these enzymes. The first L-TA family contains enzymes of both prokaryotic and eukaryotic origins, and is related to fungal ARs, whereas the second contains only prokaryotic L-TAs. Furthermore, we show that horizontal gene transfer may occur frequently during the evolution of both L-TA families. Our results indicate the complex, dynamic, and convergent evolution process of TAs and suggest an updated classification scheme for L-TAs. PMID:25644973

  11. Cholinergic receptor pathways involved in apoptosis, cell proliferation and neuronal differentiation

    PubMed Central

    Resende, Rodrigo R; Adhikari, Avishek

    2009-01-01

    Acetylcholine (ACh) has been shown to modulate neuronal differentiation during early development. Both muscarinic and nicotinic acetylcholine receptors (AChRs) regulate a wide variety of physiological responses, including apoptosis, cellular proliferation and neuronal differentiation. However, the intracellular mechanisms underlying these effects of AChR signaling are not fully understood. It is known that activation of AChRs increase cellular proliferation and neurogenesis and that regulation of intracellular calcium through AChRs may underlie the many functions of ACh. Intriguingly, activation of diverse signaling molecules such as Ras-mitogen-activated protein kinase, phosphatidylinositol 3-kinase-Akt, protein kinase C and c-Src is modulated by AChRs. Here we discuss the roles of ACh in neuronal differentiation, cell proliferation and apoptosis. We also discuss the pathways involved in these processes, as well as the effects of novel endogenous AChRs agonists and strategies to enhance neuronal-differentiation of stem and neural progenitor cells. Further understanding of the intracellular mechanisms underlying AChR signaling may provide insights for novel therapeutic strategies, as abnormal AChR activity is present in many diseases. PMID:19712465

  12. Tetrahydroxystilbene glucoside improves TNF-α-induced endothelial dysfunction: involvement of TGFβ/Smad pathway and inhibition of vimentin expression.

    PubMed

    Yao, Wenjuan; Gu, Chengjing; Shao, Haoran; Meng, Guoliang; Wang, Huiming; Jing, Xiang; Zhang, Wei

    2015-01-01

    Endothelial dysfunction plays an important role in the pathogenesis of atherogenesis. 2,3,5,4'-tetrahydroxystilbene-2-O-β-D-glucoside (TSG), an active component of the rhizome extract from Polygonum multiflorum (PM), exhibits significant anti-atherosclerotic activity. Here, we used human umbilical vein endothelial cells (HUVECs) induced by tumor necrosis factor-α (TNF-α) in vitro to investigate the cytoprotective effects of TSG on TNF-α-induced endothelial injury and the related mechanisms. Pretreatment with 50 and 100 μM TSG markedly attenuated TNF-α-induced loss of cell viability and release of lactate dehydrogenase (LDH) and inhibited TNF-α-induced cell apoptosis. The inhibition of vimentin expression was involved in the cytoprotection afforded by TSG. Using inhibitors for PI3K and TGFβ or siRNA for Akt and Smad2, we found that vimentin production in HUVECs is regulated by TGFβ/Smad signaling, but not by PI3K-Akt-mTOR signaling. Meanwhile, TSG inhibited both the expression of TGFβ1 and the phosphorylation of Smad2 and Smad3, and TSG suppressed the nuclear translocation of Smad4 induced by TNF-α. These results suggest that TSG protects HUVECs against TNF-α-induced cell damage by inhibiting vimentin expression via the interruption of the TGFβ/Smad signaling pathway. PMID:25571766

  13. The transcriptional regulatory repertoire of Corynebacterium glutamicum: reconstruction of the network controlling pathways involved in lysine and glutamate production.

    PubMed

    Brinkrolf, Karina; Schröder, Jasmin; Pühler, Alfred; Tauch, Andreas

    2010-09-01

    Corynebacterium glutamicum is one of the best studied organisms of the high G+C branch of Gram-positive bacteria and an emerging model system for the suborder Corynebacterineae. To gain insights into the regulatory gene composition and architecture of the transcriptional regulatory network of C. glutamicum, components of the transcriptional regulatory repertoire were intensively studied by many scientific groups in recent years. In this mini-review, we summarize the present knowledge about the deduced transcriptional regulatory repertoire of C. glutamicum and the current status of transcriptional regulatory network reconstruction with regard to the genome-wide detection of transcriptional regulations, coregulatory interactions and hierarchical cross-regulations. Moreover, we provide an overview of those regulators and their transcriptional regulations controlling genes involved in the conversion of the carbon sources glucose, fructose and sucrose into the industrially relevant products l-lysine and l-glutamate. This data will contribute to our understanding of l-lysine and l-glutamate production by C. glutamicum from the perspective of systems biology and may provide the basis for computational modeling of the respective biotechnologically important metabolic pathways. PMID:19963020

  14. A comparison of the genetic pathways involved in the pathogenesis of three types of colorectal cancer.

    PubMed

    Tomlinson, I; Ilyas, M; Johnson, V; Davies, A; Clark, G; Talbot, I; Bodmer, W

    1998-02-01

    Patterns of allele loss (loss of heterozygosity, LOH) have been studied in order to investigate the genetic pathways involved in the pathogenesis of three types of colorectal cancer (CRC): sporadic CRC without replication errors (RER-) (32 cases); sporadic RER+ CRC (23 cases); and ulcerative colitis-associated CRC (UCACRC) (16 cases). Each tumour was assessed for allele loss at ten microsatellite markers which map close to known or putative tumour-suppressor genes: APC (5q21-q22); DCC (18q21.1); 1p35-p36; p16 (9p21); 22q; 8p; E-cadherin (16q22.1); beta-catenin (3p22-p21.3); RB1 (13q14.1-q14.2); and HLA. Overall, high frequencies of allele loss (> 30 per cent) were found near DCC (42 per cent), p16 (38 per cent), 22q (37 per cent), 1p35-p36 (34 per cent) and APC (31 per cent), and low frequencies (< 20 per cent) near RB1 (16 per cent) and E-cadherin (13 per cent). LOH near beta-catenin, HLA, and on 8p occurred at frequencies between 20 and 30 per cent. The overall frequency of allele loss did not differ among the three tumour groups, but some variation was seen at individual loci. There was a significantly higher frequency of LOH at 1p35-36 in RER+ tumours compared to RER- tumours. Allele loss at this site was also associated with a more advanced Dukes' stage at presentation. In addition, RER- tumours showed a higher frequency of allele loss at p16 than RER+ tumours. No significant difference existed at any locus between the frequency of LOH in sporadic CRC and in UCACRC. Pairwise analysis showed a negative association between LOH at APC and DCC, and between LOH at chromosome 22p and p53 overexpression. Thus, there may be specific differences between the mutation spectra of RER+ and RER- CRCs, but there are large degrees of overlap among the underlying genetic pathways of these cancers and UCACRCs. PMID:9602705

  15. Mediators and molecular pathways involved in the regulation of neutrophil extracellular trap formation mediated by activated platelets.

    PubMed

    Carestia, Agostina; Kaufman, Tomás; Rivadeneyra, Leonardo; Landoni, Verónica Inés; Pozner, Roberto Gabriel; Negrotto, Soledad; D'Atri, Lina Paola; Gómez, Ricardo Martín; Schattner, Mirta

    2016-01-01

    In addition to being key elements in hemostasis and thrombosis, platelets amplify neutrophil function. We aimed to gain further insight into the stimuli, mediators, molecular pathways, and regulation of neutrophil extracellular trap formation mediated by human platelets. Platelets stimulated by lipopolysaccharide, a wall component of gram-negative bacteria, Pam3-cysteine-serine-lysine 4, a mimetic of lipopeptide from gram-positive bacteria, Escherichia coli, Staphylococcus aureus, or physiologic platelet agonists promoting neutrophil extracellular trap formation and myeloperoxidase-associated DNA activity under static and flow conditions. Although P-selectin or glycoprotein IIb/IIIa were not involved, platelet glycoprotein Ib, neutrophil cluster of differentiation 18, and the release of von Willebrand factor and platelet factor 4 seemed to be critical for the formation of neutrophil extracellular traps. The secretion of these molecules depended on thromboxane A(2) production triggered by lipopolysaccharide or Pam3-cysteine-serine-lysine 4 but not on high concentrations of thrombin. Accordingly, aspirin selectively inhibited platelet-mediated neutrophil extracellular trap generation. Signaling through extracellular signal-regulated kinase, phosphatidylinositol 3-kinase, and Src kinases, but not p38 or reduced nicotinamide adenine dinucleotide phosphate oxidase, was involved in platelet-triggered neutrophil extracellular trap release. Platelet-mediated neutrophil extracellular trap formation was inhibited by prostacyclin. Our results support a role for stimulated platelets in promoting neutrophil extracellular trap formation, reveal that an endothelium-derived molecule contributes to limiting neutrophil extracellular trap formation, and highlight platelet inhibition as a potential target for controlling neutrophil extracellular trap cell death. PMID:26320263

  16. Deletion mapping of the potyviral helper component-proteinase reveals two regions involved in RNA binding.

    PubMed

    Urcuqui-Inchima, S; Maia, I G; Arruda, P; Haenni, A L; Bernardi, F

    2000-03-01

    The Potyvirus helper component-proteinase (HC-Pro) binds nonspecifically to single-stranded nucleic acids with a preference for RNA. To delineate the regions of the protein responsible for RNA binding, deletions were introduced into the full-length Potato potyvirus Y HC-Pro gene carried by an Escherichia coli expression vector. The corresponding proteins were expressed as fusions with the maltose-binding protein, purified, and assayed for their RNA-binding capacity. The results obtained by UV cross-linking and Northwestern blot assays demonstrated that the N- and C-terminal regions of HC-Pro are dispensable for RNA binding. They also revealed the presence of two independent RNA-binding domains (designated A and B) located in the central part of HC-Pro. Domain B appears to contain a ribonucleoprotein (RNP) motif typical of a large family of RNA-binding proteins involved in several cellular processes. The possibility that domain B consists of an RNP domain is discussed and suggests that HC-Pro could constitute the first example of a plant viral protein belonging to the RNP-containing family of proteins. PMID:10683332

  17. Immunogenic calreticulin exposure occurs through a phylogenetically conserved stress pathway involving the chemokine CXCL8

    PubMed Central

    Sukkurwala, A Q; Martins, I; Wang, Y; Schlemmer, F; Ruckenstuhl, C; Durchschlag, M; Michaud, M; Senovilla, L; Sistigu, A; Ma, Y; Vacchelli, E; Sulpice, E; Gidrol, X; Zitvogel, L; Madeo, F; Galluzzi, L; Kepp, O; Kroemer, G

    2014-01-01

    The exposure of calreticulin (CRT) on the surface of stressed and dying cancer cells facilitates their uptake by dendritic cells and the subsequent presentation of tumor-associated antigens to T lymphocytes, hence stimulating an anticancer immune response. The chemotherapeutic agent mitoxantrone (MTX) can stimulate the peripheral relocation of CRT in both human and yeast cells, suggesting that the CRT exposure pathway is phylogenetically conserved. Here, we show that pheromones can act as physiological inducers of CRT exposure in yeast cells, thereby facilitating the formation of mating conjugates, and that a large-spectrum inhibitor of G protein-coupled receptors (which resemble the yeast pheromone receptor) prevents CRT exposure in human cancer cells exposed to MTX. An RNA interference screen as well as transcriptome analyses revealed that chemokines, in particular human CXCL8 (best known as interleukin-8) and its mouse ortholog Cxcl2, are involved in the immunogenic translocation of CRT to the outer leaflet of the plasma membrane. MTX stimulated the production of CXCL8 by human cancer cells in vitro and that of Cxcl2 by murine tumors in vivo. The knockdown of CXCL8/Cxcl2 receptors (CXCR1/Cxcr1 and Cxcr2) reduced MTX-induced CRT exposure in both human and murine cancer cells, as well as the capacity of the latter-on exposure to MTX-to elicit an anticancer immune response in vivo. Conversely, the addition of exogenous Cxcl2 increased the immunogenicity of dying cells in a CRT-dependent manner. Altogether, these results identify autocrine and paracrine chemokine signaling circuitries that modulate CRT exposure and the immunogenicity of cell death. PMID:23787997

  18. A Small Molecule Screen Exposes mTOR Signaling Pathway Involvement in Radiation-Induced Apoptosis.

    PubMed

    Sharlow, Elizabeth R; Leimgruber, Stephanie; Lira, Ana; McConnell, Michael J; Norambuena, Andrés; Bloom, George S; Epperly, Michael W; Greenberger, Joel S; Lazo, John S

    2016-05-20

    Individuals are at risk of exposure to acute ionizing radiation (IR) from a nuclear accident or terrorism, but we lack effective therapies to mitigate the lethal IR effects. In the current study, we exploited an optimized, cell-based, high throughput screening assay to interrogate a small molecule library comprising 3437 known pharmacologically active compounds for mitigation against IR-induced apoptosis. Thirty-three library compounds significantly reduced apoptosis when administered 1 h after 4 Gy IR. Two- or three-dimensional computational structural analyses of the compounds indicated only one or two chemical clusters with most of the compounds being unique structures. The mechanistic target of rapamycin complex 1 (mTORC1) inhibitor, rapamycin, was the most potent compound, and it mitigated apoptosis by 50% at 200 ± 50 pM. Other mTOR inhibitors, namely everolimus, AZD8055, and torin 1, also suppressed apoptosis, providing additional pharmacological evidence for mTOR pathway involvement in regulating cell death after IR. Everolimus and torin 1 treatment after IR decreased the S phase population and enforced both G1 and G2 phase arrest. This prorogation of cell cycle progression was accompanied by decreased IR-induced DNA damage measured by γH2AX phosphorylation at Ser139. RNA interference-mediated knockdown of the respective mTORC1 and mTORC2 subunits, Raptor or Rictor, also mitigated IR-induced apoptosis. Collectively, this study suggests a central role for the mTOR signaling in the cytotoxic response to IR and offers a useful platform to probe for additional agents. PMID:26938669

  19. DELLA proteins are common components of symbiotic rhizobial and mycorrhizal signalling pathways.

    PubMed

    Jin, Yue; Liu, Huan; Luo, Dexian; Yu, Nan; Dong, Wentao; Wang, Chao; Zhang, Xiaowei; Dai, Huiling; Yang, Jun; Wang, Ertao

    2016-01-01

    Legumes form symbiotic associations with either nitrogen-fixing bacteria or arbuscular mycorrhizal fungi. Formation of these two symbioses is regulated by a common set of signalling components that act downstream of recognition of rhizobia or mycorrhizae by host plants. Central to these pathways is the calcium and calmodulin-dependent protein kinase (CCaMK)-IPD3 complex which initiates nodule organogenesis following calcium oscillations in the host nucleus. However, downstream signalling events are not fully understood. Here we show that Medicago truncatula DELLA proteins, which are the central regulators of gibberellic acid signalling, positively regulate rhizobial symbiosis. Rhizobia colonization is impaired in della mutants and we provide evidence that DELLAs can promote CCaMK-IPD3 complex formation and increase the phosphorylation state of IPD3. DELLAs can also interact with NSP2-NSP1 and enhance the expression of Nod-factor-inducible genes in protoplasts. We show that DELLA is able to bridge a protein complex containing IPD3 and NSP2. Our results suggest a transcriptional framework for regulation of root nodule symbiosis. PMID:27514472

  20. AglQ Is a Novel Component of the Haloferax volcanii N-Glycosylation Pathway

    PubMed Central

    Arbiv, Adi; Yurist-Doutsch, Sophie; Guan, Ziqiang; Eichler, Jerry

    2013-01-01

    N-glycosylation is a post-translational modification performed by members of all three domains of life. Studies on the halophile Haloferax volcanii have offered insight into the archaeal version of this universal protein-processing event. In the present study, AglQ was identified as a novel component of the pathway responsible for the assembly and addition of a pentasaccharide to select Asn residues of Hfx. volcanii glycoproteins, such as the S-layer glycoprotein. In cells deleted of aglQ, both dolichol phosphate, the lipid carrier used in Hfx. volcanii N-glycosylation, and modified S-layer glycoprotein Asn residues only presented the first three pentasaccharide subunits, pointing to a role for AglQ in either preparing the third sugar for attachment of the fourth pentasaccharide subunit or processing the fourth sugar prior to its addition to the lipid-linked trisaccharide. To better define the precise role of AglQ, shown to be a soluble protein, bioinformatics tools were recruited to identify sequence or structural homologs of known function. Site-directed mutagenesis experiments guided by these predictions identified residues important for AglQ function. The results obtained point to AglQ acting as an isomerase in Hfx. volcanii N-glycosylation. PMID:24236216

  1. DELLA proteins are common components of symbiotic rhizobial and mycorrhizal signalling pathways

    PubMed Central

    Jin, Yue; Liu, Huan; Luo, Dexian; Yu, Nan; Dong, Wentao; Wang, Chao; Zhang, Xiaowei; Dai, Huiling; Yang, Jun; Wang, Ertao

    2016-01-01

    Legumes form symbiotic associations with either nitrogen-fixing bacteria or arbuscular mycorrhizal fungi. Formation of these two symbioses is regulated by a common set of signalling components that act downstream of recognition of rhizobia or mycorrhizae by host plants. Central to these pathways is the calcium and calmodulin-dependent protein kinase (CCaMK)–IPD3 complex which initiates nodule organogenesis following calcium oscillations in the host nucleus. However, downstream signalling events are not fully understood. Here we show that Medicago truncatula DELLA proteins, which are the central regulators of gibberellic acid signalling, positively regulate rhizobial symbiosis. Rhizobia colonization is impaired in della mutants and we provide evidence that DELLAs can promote CCaMK–IPD3 complex formation and increase the phosphorylation state of IPD3. DELLAs can also interact with NSP2–NSP1 and enhance the expression of Nod-factor-inducible genes in protoplasts. We show that DELLA is able to bridge a protein complex containing IPD3 and NSP2. Our results suggest a transcriptional framework for regulation of root nodule symbiosis. PMID:27514472

  2. Involvement of the flagellar assembly pathway in Vibrio alginolyticus adhesion under environmental stresses

    PubMed Central

    Wang, Lu; Huang, Lixing; Su, Yongquan; Qin, Yingxue; Kong, Wendi; Ma, Ying; Xu, Xiaojin; Lin, Mao; Zheng, Jiang; Yan, Qingpi

    2015-01-01

    Adhesion is an important virulence factor of Vibrio alginolyticus. This factor may be affected by environmental conditions; however, its molecular mechanism remains unclear. In our previous research, adhesion deficient strains were obtained by culturing V. alginolyticus under stresses including Cu, Pb, Hg, and low pH. With RNA-seq and bioinformatics analysis, we found that all of these stress treatments significantly affected the flagellar assembly pathway, which may play an important role in V. alginolyticus adhesion. Therefore, we hypothesized that the environmental stresses of the flagellar assembly pathway may be one way in which environmental conditions affect adhesion. To verify our hypothesis, a bioinformatics analysis, QPCR, RNAi, in vitro adhesion assay and motility assay were performed. Our results indicated that (1) the flagellar assembly pathway was sensitive to environmental stresses, (2) the flagellar assembly pathway played an important role in V. alginolyticus adhesion, and (3) motility is not the only way in which the flagellar assembly pathway affects adhesion. PMID:26322276

  3. Two-Component Signal Transduction Pathways Regulating Growth and Cell Cycle Progression in a Bacterium: A System-Level Analysis

    PubMed Central

    2005-01-01

    Two-component signal transduction systems, comprised of histidine kinases and their response regulator substrates, are the predominant means by which bacteria sense and respond to extracellular signals. These systems allow cells to adapt to prevailing conditions by modifying cellular physiology, including initiating programs of gene expression, catalyzing reactions, or modifying protein–protein interactions. These signaling pathways have also been demonstrated to play a role in coordinating bacterial cell cycle progression and development. Here we report a system-level investigation of two-component pathways in the model organism Caulobacter crescentus. First, by a comprehensive deletion analysis we show that at least 39 of the 106 two-component genes are required for cell cycle progression, growth, or morphogenesis. These include nine genes essential for growth or viability of the organism. We then use a systematic biochemical approach, called phosphotransfer profiling, to map the connectivity of histidine kinases and response regulators. Combining these genetic and biochemical approaches, we identify a new, highly conserved essential signaling pathway from the histidine kinase CenK to the response regulator CenR, which plays a critical role in controlling cell envelope biogenesis and structure. Depletion of either cenK or cenR leads to an unusual, severe blebbing of cell envelope material, whereas constitutive activation of the pathway compromises cell envelope integrity, resulting in cell lysis and death. We propose that the CenK–CenR pathway may be a suitable target for new antibiotic development, given previous successes in targeting the bacterial cell wall. Finally, the ability of our in vitro phosphotransfer profiling method to identify signaling pathways that operate in vivo takes advantage of an observation that histidine kinases are endowed with a global kinetic preference for their cognate response regulators. We propose that this system

  4. IDENTIFICATION OF THE ROLE OF APOPTOSIS PATHWAYS POTENTIALLY INVOLVED IN FORMALDEHYDE-INDUCED CARCINOGENESIS USING CDNA ARRAYS

    EPA Science Inventory

    Identification of the Role of Apoptosis Pathways Potentially Involved in Formaldehyde- Induced Carcinogenesis Using cDNA Arrays.

    Formaldehyde (FA) is a genotoxic chemical found in household, medicinal, and industrial products. Although the major source of human exposure is...

  5. Association of Polymorphisms in BDNF, MTHFR, and Genes Involved in the Dopaminergic Pathway with Memory in a Healthy Chinese Population

    ERIC Educational Resources Information Center

    Yeh, Ting-Kuang; Hu, Chung-Yi; Yeh, Ting-Chi; Lin, Pei-Jung; Wu, Chung-Hsin; Lee, Po-Lei; Chang, Chun-Yen

    2012-01-01

    The contribution of genetic factors to the memory is widely acknowledged. Research suggests that these factors include genes involved in the dopaminergic pathway, as well as the genes for brain-derived neurotrophic factor (BDNF) and methylenetetrahydrofolate reductase (MTHFR). The activity of the products of these genes is affected by single…

  6. Identifying components of the hair-cell interactome involved in cochlear amplification

    PubMed Central

    Zheng, Jing; Anderson, Charles T; Miller, Katharine K; Cheatham, MaryAnn; Dallos, Peter

    2009-01-01

    Background Although outer hair cells (OHCs) play a key role in cochlear amplification, it is not fully understood how they amplify sound signals by more than 100 fold. Two competing or possibly complementary mechanisms, stereocilia-based and somatic electromotility-based amplification, have been considered. Lacking knowledge about the exceptionally rich protein networks in the OHC plasma membrane, as well as related protein-protein interactions, limits our understanding of cochlear function. Therefore, we focused on finding protein partners for two important membrane proteins: Cadherin 23 (cdh23) and prestin. Cdh23 is one of the tip-link proteins involved in transducer function, a key component of mechanoelectrical transduction and stereocilia-based amplification. Prestin is a basolateral membrane protein responsible for OHC somatic electromotility. Results Using the membrane-based yeast two-hybrid system to screen a newly built cDNA library made predominantly from OHCs, we identified two completely different groups of potential protein partners using prestin and cdh23 as bait. These include both membrane bound and cytoplasmic proteins with 12 being de novo gene products with unknown function(s). In addition, some of these genes are closely associated with deafness loci, implying a potentially important role in hearing. The most abundant prey for prestin (38%) is composed of a group of proteins involved in electron transport, which may play a role in OHC survival. The most abundant group of cdh23 prey (55%) contains calcium-binding domains. Since calcium performs an important role in hair cell mechanoelectrical transduction and amplification, understanding the interactions between cdh23 and calcium-binding proteins should increase our knowledge of hair cell function at the molecular level. Conclusion The results of this study shed light on some protein networks in cochlear hair cells. Not only was a group of de novo genes closely associated with known deafness loci

  7. Components of the calcium-calcineurin signaling pathway in fungal cells and their potential as antifungal targets.

    PubMed

    Liu, Shuyuan; Hou, Yinglong; Liu, Weiguo; Lu, Chunyan; Wang, Weixin; Sun, Shujuan

    2015-04-01

    In recent years, the emergence of fungal resistance has become frequent, partly due to the widespread clinical use of fluconazole, which is minimally toxic and effective in the prevention and treatment of Candida albicans infections. The limited selection of antifungal drugs for clinical fungal infection therapy has prompted us to search for new antifungal drug targets. Calcium, which acts as the second messenger in both mammals and fungi, plays a direct role in controlling the expression patterns of its signaling systems and has important roles in cell survival. In addition, calcium and some of the components, mainly calcineurin, in the fungal calcium signaling pathway mediate fungal resistance to antifungal drugs. Therefore, an overview of the components of the fungal calcium-calcineurin signaling network and their potential roles as antifungal targets is urgently needed. The calcium-calcineurin signaling pathway consists of various channels, transporters, pumps, and other proteins or enzymes. Many transcriptional profiles have indicated that mutant strains that lack some of these components are sensitized to fluconazole or other antifungal drugs. In addition, many researchers have identified efficient compounds that exhibit antifungal activity by themselves or in combination with antifungal drugs by targeting some of the components in the fungal calcium-calcineurin signaling pathway. This targeting disrupts Ca(2+) homeostasis, which suggests that this pathway contains potential targets for the development of new antifungal drugs. PMID:25636321

  8. Components of the Calcium-Calcineurin Signaling Pathway in Fungal Cells and Their Potential as Antifungal Targets

    PubMed Central

    Liu, Shuyuan; Hou, Yinglong; Liu, Weiguo; Lu, Chunyan; Wang, Weixin

    2015-01-01

    In recent years, the emergence of fungal resistance has become frequent, partly due to the widespread clinical use of fluconazole, which is minimally toxic and effective in the prevention and treatment of Candida albicans infections. The limited selection of antifungal drugs for clinical fungal infection therapy has prompted us to search for new antifungal drug targets. Calcium, which acts as the second messenger in both mammals and fungi, plays a direct role in controlling the expression patterns of its signaling systems and has important roles in cell survival. In addition, calcium and some of the components, mainly calcineurin, in the fungal calcium signaling pathway mediate fungal resistance to antifungal drugs. Therefore, an overview of the components of the fungal calcium-calcineurin signaling network and their potential roles as antifungal targets is urgently needed. The calcium-calcineurin signaling pathway consists of various channels, transporters, pumps, and other proteins or enzymes. Many transcriptional profiles have indicated that mutant strains that lack some of these components are sensitized to fluconazole or other antifungal drugs. In addition, many researchers have identified efficient compounds that exhibit antifungal activity by themselves or in combination with antifungal drugs by targeting some of the components in the fungal calcium-calcineurin signaling pathway. This targeting disrupts Ca2+ homeostasis, which suggests that this pathway contains potential targets for the development of new antifungal drugs. PMID:25636321

  9. Modulation of WNT/β-catenin pathway in melanoma by biologically active components derived from plants.

    PubMed

    Gajos-Michniewicz, Anna; Czyz, Malgorzata

    2016-03-01

    Metastatic melanoma is an aggressive cancer, often resistant to treatment. Therefore, it is essential to determine the molecular mechanisms leading to melanoma or underlying resistance to therapy, and the response to targeted inhibition of the RAS/BRAF/MEK/ERK pathway was a good lesson in this respect. Aberrant WNT/β-catenin pathway is observed in melanoma, and the modulators of this signaling cascade have been under investigation in the context of therapy as well as chemoprevention. Several natural compounds were recognized as being capable of targeting elements of the WNT/β-catenin pathway in various cancers, however, only a few of them can modulate this pathway in melanoma. This review examines recent research on the role of the WNT/β-catenin pathway in tumor development and maintenance, as well as summarizes the current knowledge concerning the modulation of this pathway in melanoma by active compounds of natural origin. PMID:26851176

  10. Single-strand gap repair involves both RecF and RecBCD pathways.

    PubMed

    Pagès, Vincent

    2016-08-01

    Homologous recombination repairs discontinuities in DNA including single-strand gaps (SSGs) and double-strand breaks (DSBs). This commentary describes how the RecBCD and RecF pathways might be exchangeable for the repair of their respective DSB and SSG canonical substrates. In particular, I will discuss how the RecBCD pathway could engage in the repair of an SSG even when the latter is not associated with a DSB. PMID:26874520

  11. Steroidogenic pathways involved in androgen biosynthesis in eumenorrheic women and patients with polycystic ovary syndrome.

    PubMed

    Saito, Kazuki; Matsuzaki, Toshiya; Iwasa, Takeshi; Miyado, Mami; Saito, Hidekazu; Hasegawa, Tomonobu; Homma, Keiko; Inoue, Eisuke; Miyashiro, Yoshimichi; Kubota, Toshiro; Irahara, Minoru; Ogata, Tsutomu; Fukami, Maki

    2016-04-01

    The conventional Δ5 and Δ4 steroidogenic pathways mediate androgen production in females. While multiple non-conventional pathways to dihydrotestosterone (DHT) have recently been postulated in humans, the functional significance of these pathways remains to be elucidated. The aim of this study was to clarify the origin of androgens in healthy women and in patients with polycystic ovary syndrome (PCOS), a multifactorial disorder characterized by androgen overproduction. We measured 13 steroids in blood samples of 31 eumenorrheic females and 28 PCOS patients using liquid chromatography-tandem mass spectrometry and chemiluminescent enzyme immunoassay. We found that 17-hydroxy (17-OH) progesterone (17-OHP), androstenedione (Δ4A), testosterone, androstanedione, androsterone, and androstanediol levels were higher in the patient group than in the eumenorrheic group, while levels of other steroids were comparable between the two groups. In the eumenorrheic group, DHT levels were correlated with testosterone, androstanedione, and androstanediol. Quantitative correlations were also observed among 17-OH allopregnanolone, androsterone, androstanediol, and DHT, and among Δ4A, androstanedione, androsterone, and androstanediol. In the patient group, DHT levels were correlated with testosterone levels, but not with androstanedione or androstanediol levels. Δ4A and testosterone paralleled 17-OHP. Androstanedione, androsterone, androstanediol, and 17-OH allopregnanolone were quantitatively correlated. In both groups, multivariable linear regression analyses suggested relationships between androsterone and androstanedione, as well as between androsterone and 17-OH allopregnanolone. These results indicate that multiple androgen biosynthesis pathways are operating in eumenorrheic females and PCOS patients. In PCOS patients, excessive androgens are produced primarily via the conventional pathways, while two alternative pathways; i.e., an androstanedione-mediated pathway and a so

  12. Genes Involved in the Endoplasmic Reticulum N-Glycosylation Pathway of the Red Microalga Porphyridium sp.: A Bioinformatic Study

    PubMed Central

    Levy-Ontman, Oshrat; Fisher, Merav; Shotland, Yoram; Weinstein, Yacob; Tekoah, Yoram; Arad, Shoshana Malis

    2014-01-01

    N-glycosylation is one of the most important post-translational modifications that influence protein polymorphism, including protein structures and their functions. Although this important biological process has been extensively studied in mammals, only limited knowledge exists regarding glycosylation in algae. The current research is focused on the red microalga Porphyridium sp., which is a potentially valuable source for various applications, such as skin therapy, food, and pharmaceuticals. The enzymes involved in the biosynthesis and processing of N-glycans remain undefined in this species, and the mechanism(s) of their genetic regulation is completely unknown. In this study, we describe our pioneering attempt to understand the endoplasmic reticulum N-Glycosylation pathway in Porphyridium sp., using a bioinformatic approach. Homology searches, based on sequence similarities with genes encoding proteins involved in the ER N-glycosylation pathway (including their conserved parts) were conducted using the TBLASTN function on the algae DNA scaffold contigs database. This approach led to the identification of 24 encoded-genes implicated with the ER N-glycosylation pathway in Porphyridium sp. Homologs were found for almost all known N-glycosylation protein sequences in the ER pathway of Porphyridium sp.; thus, suggesting that the ER-pathway is conserved; as it is in other organisms (animals, plants, yeasts, etc.). PMID:24514561

  13. Genomic Copy Number Variation Affecting Genes Involved in the Cell Cycle Pathway: Implications for Somatic Mosaicism

    PubMed Central

    Iourov, Ivan Y.; Vorsanova, Svetlana G.; Zelenova, Maria A.; Korostelev, Sergei A.; Yurov, Yuri B.

    2015-01-01

    Somatic genome variations (mosaicism) seem to represent a common mechanism for human intercellular/interindividual diversity in health and disease. However, origins and mechanisms of somatic mosaicism remain a matter of conjecture. Recently, it has been hypothesized that zygotic genomic variation naturally occurring in humans is likely to predispose to nonheritable genetic changes (aneuploidy) acquired during the lifetime through affecting cell cycle regulation, genome stability maintenance, and related pathways. Here, we have evaluated genomic copy number variation (CNV) in genes implicated in the cell cycle pathway (according to Kyoto Encyclopedia of Genes and Genomes/KEGG) within a cohort of patients with intellectual disability, autism, and/or epilepsy, in which the phenotype was not associated with genomic rearrangements altering this pathway. Benign CNVs affecting 20 genes of the cell cycle pathway were detected in 161 out of 255 patients (71.6%). Among them, 62 individuals exhibited >2 CNVs affecting the cell cycle pathway. Taking into account the number of individuals demonstrating CNV of these genes, a support for this hypothesis appears to be presented. Accordingly, we speculate that further studies of CNV burden across the genes implicated in related pathways might clarify whether zygotic genomic variation generates somatic mosaicism in health and disease. PMID:26421275

  14. Processing and MHC class II presentation of exogenous soluble antigen involving a proteasome-dependent cytosolic pathway in CD40-activated B cells.

    PubMed

    Becker, Hans Jiro; Kondo, Eisei; Shimabukuro-Vornhagen, Alexander; Theurich, Sebastian; von Bergwelt-Baildon, Michael S

    2016-08-01

    Activated B cells have the capacity to present antigen and induce immune responses as potent antigen-presenting cells (APCs). As in other APCs, antigen presentation by B cells involves antigen internalization, antigen processing, and peptide loading onto MHC molecules. However, while the mechanism of antigen processing has been studied extensively in other APCs, this pathway remains elusive in B cells. The aim of this study was to investigate the MHC class II processing pathway in CD40-activated B cells (CD40Bs), as a model for activated, antigen-presenting B cells. Using CMV pp65 as a model antigen, we evaluated processing and presentation of the CD4 + T-cell epitope 509-523 (K509) by human CD40Bs in ELISPOT assays. As expected, stimulation of specific CD4 + T-cell clones was attenuated after pretreatment of CD40Bs with inhibitors of classic class II pathway components. However, proteasome inhibitors such as epoxomicin limited antigen presentation as well. This suggests that the antigen is processed in a non-classical, cytosolic MHC class II pathway. Further experiments with truncated protein variants revealed involvement of the proteasome in processing of the N and C extensions of the epitope. Access to the cytosol was shown to be size dependent. Epoxomicin sensitivity exclusively in CD40B cells, but not in dendritic cells, suggests a novel processing mechanism unique to this APC. Our data suggest that B cells process antigen using a distinct, non-classical class II pathway. PMID:26561366

  15. The Protein Dredd Is an Essential Component of the c-Jun N-terminal Kinase Pathway in the Drosophila Immune Response*

    PubMed Central

    Guntermann, Silvia; Foley, Edan

    2011-01-01

    The Drosophila immune deficiency (IMD) pathway mobilizes c-Jun N-terminal kinase (JNK), caspase, and nuclear factor-κB (NF-κB) modules to counter infection with Gram-negative bacteria. Dredd is an essential caspase in the IMD pathway, and it is widely established that NF-κB activation depends on Dredd. More recent cell culture studies suggested a role for Dredd in the activation of dJNK (Drosophila JNK). However, there are no epistatic or mechanistic data on the involvement of Dredd in dJNK activation. More importantly, there is no in vivo evidence to demonstrate a physiological requirement for Dredd in the IMD/dJNK pathway. We performed a comprehensive analysis of the role of Dredd in the IMD/dJNK pathway, and we demonstrated that Dredd is essential for the activation of IMD/dJNK in cell culture. We positioned Dredd activity at an early point of the IMD/dJNK pathway and uncovered a series of interactions between Dredd and additional proximal IMD pathway molecules. Mechanistically, we showed that the caspase activity inhibitor p35 blocked dJNK activation and the induction of dJNK-dependent genes in cell culture and in vivo. Most importantly, we demonstrated that dredd mutant flies are completely inhibited in their ability to activate dJNK or express dJNK-responsive target genes after bacterial infection in vivo. In conclusion, we established Dredd as an essential component of the IMD pathway required for the full activation of IMD/dJNK in cell culture and in vivo. Our data enhance our appreciation of Dredd-dependent IMD signal transduction events. PMID:21730059

  16. The Fas-associated death domain protein/caspase-8/c-FLIP signaling pathway is involved in TNF-induced activation of ERK

    SciTech Connect

    Lueschen, Silke; Falk, Markus; Scherer, Gudrun; Ussat, Sandra; Paulsen, Maren; Adam-Klages, Sabine . E-mail: sadam@email.uni-kiel.de

    2005-10-15

    The cytokine TNF activates multiple signaling pathways leading to cellular responses ranging from proliferation and survival to apoptosis. While most of these pathways have been elucidated in detail over the past few years, the molecular mechanism leading to the activation of the MAP kinases ERK remains ill defined and is controversially discussed. Therefore, we have analyzed TNF-induced ERK activation in various human and murine cell lines and show that it occurs in a cell-type-specific manner. In addition, we provide evidence for the involvement of the signaling components Fas-associated death domain protein (FADD), caspase-8, and c-FLIP in the pathway activating ERK in response to TNF. This conclusion is based on the following observations: (I) Overexpression of FADD, caspase-8, or a c-FLIP protein containing the death effector domains only leads to enhanced and prolonged ERK activation after TNF treatment. (II) TNF-induced ERK activation is strongly diminished in the absence of FADD. Interestingly, the enzymatic function of caspase-8 is not required for TNF-induced ERK activation. Additional evidence suggests a role for this pathway in the proliferative response of murine fibroblasts to TNF.

  17. Phylogenomic study of lipid genes involved in microalgal biofuel production-candidate gene mining and metabolic pathway analyses.

    PubMed

    Misra, Namrata; Panda, Prasanna Kumar; Parida, Bikram Kumar; Mishra, Barada Kanta

    2012-01-01

    Optimizing microalgal biofuel production using metabolic engineering tools requires an in-depth understanding of the structure-function relationship of genes involved in lipid biosynthetic pathway. In the present study, genome-wide identification and characterization of 398 putative genes involved in lipid biosynthesis in Arabidopsis thaliana Chlamydomonas reinhardtii, Volvox carteri, Ostreococcus lucimarinus, Ostreococcus tauri and Cyanidioschyzon merolae was undertaken on the basis of their conserved motif/domain organization and phylogenetic profile. The results indicated that the core lipid metabolic pathways in all the species are carried out by a comparable number of orthologous proteins. Although the fundamental gene organizations were observed to be invariantly conserved between microalgae and Arabidopsis genome, with increased order of genome complexity there seems to be an association with more number of genes involved in triacylglycerol (TAG) biosynthesis and catabolism. Further, phylogenomic analysis of the genes provided insights into the molecular evolution of lipid biosynthetic pathway in microalgae and confirm the close evolutionary proximity between the Streptophyte and Chlorophyte lineages. Together, these studies will improve our understanding of the global lipid metabolic pathway and contribute to the engineering of regulatory networks of algal strains for higher accumulation of oil. PMID:23032611

  18. Methylamine Utilization via the N-Methylglutamate Pathway in Methylobacterium extorquens PA1 Involves a Novel Flow of Carbon through C1 Assimilation and Dissimilation Pathways

    PubMed Central

    Nayak, Dipti D.

    2014-01-01

    Methylotrophs grow on reduced single-carbon compounds like methylamine as the sole source of carbon and energy. In Methylobacterium extorquens AM1, the best-studied aerobic methylotroph, a periplasmic methylamine dehydrogenase that catalyzes the primary oxidation of methylamine to formaldehyde has been examined in great detail. However, recent metagenomic data from natural ecosystems are revealing the abundance and importance of lesser-known routes, such as the N-methylglutamate pathway, for methylamine oxidation. In this study, we used M. extorquens PA1, a strain that is closely related to M. extorquens AM1 but is lacking methylamine dehydrogenase, to dissect the genetics and physiology of the ecologically relevant N-methylglutamate pathway for methylamine oxidation. Phenotypic analyses of mutants with null mutations in genes encoding enzymes of the N-methylglutamate pathway suggested that γ-glutamylmethylamide synthetase is essential for growth on methylamine as a carbon source but not as a nitrogen source. Furthermore, analysis of M. extorquens PA1 mutants with defects in methylotrophy-specific dissimilatory and assimilatory modules suggested that methylamine use via the N-methylglutamate pathway requires the tetrahydromethanopterin (H4MPT)-dependent formaldehyde oxidation pathway but not a complete tetrahydrofolate (H4F)-dependent formate assimilation pathway. Additionally, we present genetic evidence that formaldehyde-activating enzyme (FAE) homologs might be involved in methylotrophy. Null mutants of FAE and homologs revealed that FAE and FAE2 influence the growth rate and FAE3 influences the yield during the growth of M. extorquens PA1 on methylamine. PMID:25225269

  19. Signaling pathway of nitric oxide production induced by ginsenoside Rb1 in human aortic endothelial cells: a possible involvement of androgen receptor.

    PubMed

    Yu, Jing; Eto, Masato; Akishita, Masahiro; Kaneko, Akiyo; Ouchi, Yasuyoshi; Okabe, Tetsuro

    2007-02-16

    Ginsenosides have been shown to stimulate nitric oxide (NO) production in aortic endothelial cells. However, the signaling pathways involved have not been well studied in human aortic endothelial cells. The present study was designed to examine whether purified ginsenoside Rb1, a major active component of ginseng could actually induce NO production and to clarify the signaling pathway in human aortic endothelial cells. NO production was rapidly increased by Rb1. The rapid increase in NO production was abrogated by treatment with nitric oxide synthetase inhibitor, L-NAME. Rb1 stimulated rapid phosphorylation of Akt (Ser473), ERK1/2 (Thr202/Thr204) and eNOS (Ser1177). Rapid phosphorylation of eNOS (Ser1177) was prevented by SH-5, an Akt inhibitor or wortmannin, PI3-kinase inhibitor and partially attenuated by PD98059, an upstream inhibitor for ERK1/2. Interestingly, NO production and eNOS phosphorylation at Ser1177 by Rb1 were abolished by androgen receptor antagonist, nilutamide. The results suggest that PI3kinase/Akt and MEK/ERK pathways and androgen receptor are involved in the regulation of acute eNOS activation by Rb1 in human aortic endothelial cells. PMID:17196933

  20. Involvement of transcription factor Oct-1 in the regulation of JAK-STAT signaling pathway in cells of Burkitt lymphoma.

    PubMed

    Pankratova, E V; Stepchenko, A G; Krylova, I D; Portseva, T N; Georgieva, S G

    2016-05-01

    We studied the role of transcription factor Oct-1 in the regulation of expression of genes of the JAK-STAT signaling pathway in the Namalwa Burkitt's lymphoma cell line. Overexpression of Oct-1 isoforms (Oct-1A, Oct-1L, and Oct-1X) causes a decrease in the activity of four genes involved in the JAK-STAT signaling pathway-IFNAR2, STAT1, STAT2, and STAT4. As a result of our research, it was found that genes STAT2 and STAT4 are direct targets for Oct-1 protein. PMID:27417729

  1. Enzymes involved in a novel anaerobic cyclohexane carboxylic acid degradation pathway.

    PubMed

    Kung, Johannes W; Meier, Anne-Katrin; Mergelsberg, Mario; Boll, Matthias

    2014-10-01

    The anaerobic degradation of cyclohexane carboxylic acid (CHC) has so far been studied only in Rhodopseudomonas palustris, in which CHC is activated to cyclohexanoyl coenzyme A (cyclohexanoyl-CoA [CHCoA]) and then dehydrogenated to cyclohex-1-ene-1-carboxyl-CoA (CHeneCoA). This intermediate is further degraded by reactions of the R. palustris-specific benzoyl-CoA degradation pathway of aromatic compounds. However, CHeneCoA is not an intermediate in the degradation of aromatic compounds in all other known anaerobic bacteria; consequently, degradation of CHC was mostly unknown in anaerobic bacteria. We identified a previously unknown CHC degradation pathway in the Fe(III)-reducing Geobacter metallireducens by determining the following CHC-induced in vitro activities: (i) the activation of CHC to CHCoA by a succinyl-CoA:CHC CoA transferase, (ii) the 1,2-dehydrogenation of CHCoA to CHeneCoA by CHCoA dehydrogenase, and (iii) the unusual 1,4-dehydrogenation of CHeneCoA to cyclohex-1,5-diene-1-carboxyl-CoA. This last represents a previously unknown joint intermediate of the CHC and aromatic compound degradation pathway in bacteria other than R. palustris. The enzymes catalyzing the three reactions were purified and characterized as specific enzymes after heterologous expression of the encoding genes. Quantitative reverse transcription-PCR revealed that expression of these genes was highly induced during growth with CHC but not with benzoate. The newly identified CHC degradation pathway is suggested to be present in nearly all CHC-degrading anaerobic bacteria, including denitrifying, Fe(III)-reducing, sulfate-reducing, and fermenting bacteria. Remarkably, all three CHC degradation pathways always link CHC catabolism to the catabolic pathways of aromatic compounds. We propose that the capacity to use CHC as a carbon source evolved from already-existing aromatic compound degradation pathways. PMID:25112478

  2. Two Components of the RNA-Directed DNA Methylation Pathway Associate with MORC6 and Silence Loci Targeted by MORC6 in Arabidopsis

    PubMed Central

    Liu, Zhang-Wei; Zhou, Jin-Xing; Huang, Huan-Wei; Li, Yong-Qiang; Shao, Chang-Rong; Li, Lin; Cai, Tao; Chen, She

    2016-01-01

    The SU(VAR)3-9 homolog SUVH9 and the double-stranded RNA-binding protein IDN2 were thought to be components of an RNA-directed DNA methylation (RdDM) pathway in Arabidopsis. We previously found that SUVH9 interacts with MORC6 but how the interaction contributes to transcriptional silencing remains elusive. Here, our genetic analysis indicates that SUVH2 and SUVH9 can either act in the same pathway as MORC6 or act synergistically with MORC6 to mediate transcriptional silencing. Moreover, we demonstrate that IDN2 interacts with MORC6 and mediates the silencing of a subset of MORC6 target loci. Like SUVH2, SUVH9, and IDN2, other RdDM components including Pol IV, Pol V, RDR2, and DRM2 are also required for transcriptional silencing at a subset of MORC6 target loci. MORC6 was previously shown to mediate transcriptional silencing through heterochromatin condensation. We demonstrate that the SWI/SNF chromatin-remodeling complex components SWI3B, SWI3C, and SWI3D interact with MORC6 as well as with SUVH9 and then mediate transcriptional silencing. These results suggest that the RdDM components are involved not only in DNA methylation but also in MORC6-mediated heterochromatin condensation. This study illustrates how DNA methylation is linked to heterochromatin condensation and thereby enhances transcriptional silencing at methylated genomic regions. PMID:27171427

  3. Two Components of the RNA-Directed DNA Methylation Pathway Associate with MORC6 and Silence Loci Targeted by MORC6 in Arabidopsis.

    PubMed

    Liu, Zhang-Wei; Zhou, Jin-Xing; Huang, Huan-Wei; Li, Yong-Qiang; Shao, Chang-Rong; Li, Lin; Cai, Tao; Chen, She; He, Xin-Jian

    2016-05-01

    The SU(VAR)3-9 homolog SUVH9 and the double-stranded RNA-binding protein IDN2 were thought to be components of an RNA-directed DNA methylation (RdDM) pathway in Arabidopsis. We previously found that SUVH9 interacts with MORC6 but how the interaction contributes to transcriptional silencing remains elusive. Here, our genetic analysis indicates that SUVH2 and SUVH9 can either act in the same pathway as MORC6 or act synergistically with MORC6 to mediate transcriptional silencing. Moreover, we demonstrate that IDN2 interacts with MORC6 and mediates the silencing of a subset of MORC6 target loci. Like SUVH2, SUVH9, and IDN2, other RdDM components including Pol IV, Pol V, RDR2, and DRM2 are also required for transcriptional silencing at a subset of MORC6 target loci. MORC6 was previously shown to mediate transcriptional silencing through heterochromatin condensation. We demonstrate that the SWI/SNF chromatin-remodeling complex components SWI3B, SWI3C, and SWI3D interact with MORC6 as well as with SUVH9 and then mediate transcriptional silencing. These results suggest that the RdDM components are involved not only in DNA methylation but also in MORC6-mediated heterochromatin condensation. This study illustrates how DNA methylation is linked to heterochromatin condensation and thereby enhances transcriptional silencing at methylated genomic regions. PMID:27171427

  4. Uncovering genetic components involved in regulating early immune responses to porcine reproductive and respiratory syndrome (PRRS)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Porcine reproductive and respiratory syndrome virus (PRRSV)-infected pigs are susceptible to pneumonia and reproductive losses. Our goal is to identify the most significant pathways and genes regulating early responses during two pathologic acute PRRSV infections as compared to protective vaccinatio...

  5. Differential involvement of indole-3-acetic acid biosynthetic pathways in pathogenicity and epiphytic fitness of Erwinia herbicola pv. gypsophilae.

    PubMed

    Manulis, S; Haviv-Chesner, A; Brandl, M T; Lindow, S E; Barash, I

    1998-07-01

    Erwinia herbicola pv. gypsophilae (Ehg), which induces galls on Gypsophila paniculata, harbors two major pathways for indole-3-acetic acid (IAA) synthesis, the indole-3-acetamide (IAM) and indole-3-pyruvate (IPyA) routes, as well as cytokinin biosynthetic genes. Mutants were generated in which the various biosynthetic routes were disrupted separately or jointly in order to assess the contribution of IAA of various origins and cytokinins to pathogenicity and epiphytic fitness. Inactivation of the IAM pathway or cytokinin biosynthesis caused the largest reduction in gall size. Inactivation of the IPyA pathway caused a minor, nonsignificant decrease in pathogenicity. No further reduction in gall size was observed by the simultaneous inactivation of both IAA pathways only or in combination with that of cytokinin production. However, inactivation of the IPyA pathway caused a 14-fold reduction in the population of Ehg on bean plants. Inactivation of the IAM pathway or cytokinin production did not affect epiphytic fitness. While the apparent transcriptional activity of iaaM-inaZ fusion increased slightly in cells of Ehg on bean and gypsophila leaves, compared with that in culture, very high levels of induction were observed in cells injected into gypsophila stems. In contrast, moderate levels of induction of ipdC-inaZ in Ehg were observed on leaves of these plants and in gypsophila stems, when compared with that in culture. These results suggest that the IAM pathway is involved primarily in gall formation and support the main contribution of the IpyA pathway to the epiphytic fitness of this bacterial species. PMID:9650296

  6. The involvement of the JAK-STAT signaling pathway in chronic inflammatory skin disease atopic dermatitis

    PubMed Central

    Bao, Lei; Zhang, Huayi; Chan, Lawrence S

    2013-01-01

    Atopic dermatitis (AD), a common chronic inflammatory skin disease, is characterized by inflammatory cell skin infiltration. The JAK-STAT pathway has been shown to play an essential role in the dysregulation of immune responses in AD, including the exaggeration of Th2 cell response, the activation of eosinophils, the maturation of B cells, and the suppression of regulatory T cells (Tregs). In addition, the JAK-STAT pathway, activated by IL-4, also plays a critical role in the pathogenesis of AD by upregulating epidermal chemokines, pro-inflammatroy cytokines, and pro-angiogenic factors as well as by downregulating antimicrobial peptides (AMPs) and factors responsible for skin barrier function. In this review, we will highlight the recent advances in our understanding of the JAK-STAT pathway in the pathogenesis of AD. PMID:24069552

  7. S100B secretion is stimulated by IL-1beta in glial cultures and hippocampal slices of rats: Likely involvement of MAPK pathway.

    PubMed

    de Souza, Daniela F; Leite, Marina C; Quincozes-Santos, André; Nardin, Patrícia; Tortorelli, Lucas S; Rigo, Maurício M; Gottfried, Carmem; Leal, Rodrigo B; Gonçalves, Carlos-Alberto

    2009-01-01

    S100B is an astrocyte-derived cytokine implicated in the IL-1beta-triggered cytokine cycle in Alzheimer's disease. However, the secretion of S100B following stimulation by IL-1beta has not been directly demonstrated. We investigated S100B secretion in cortical primary astrocyte cultures, C6 glioma cells and acute hippocampal slices exposed to IL-1beta. S100B secretion was induced by IL-1beta in all preparations, involving MAPK pathway and, apparently, NF-small ka, CyrillicB signaling. Astrocytes and C6 cells exhibited different sensitivities to IL-1beta. These results suggest that IL-1beta-induced S100B secretion is a component of the neuroinflammatory response, which would support the involvement of S100B in the genesis of neurodegenerative diseases. PMID:19042033

  8. A pathway to bone: signaling molecules and transcription factors involved in chondrocyte development and maturation

    PubMed Central

    Kozhemyakina, Elena; Lassar, Andrew B.; Zelzer, Elazar

    2015-01-01

    Decades of work have identified the signaling pathways that regulate the differentiation of chondrocytes during bone formation, from their initial induction from mesenchymal progenitor cells to their terminal maturation into hypertrophic chondrocytes. Here, we review how multiple signaling molecules, mechanical signals and morphological cell features are integrated to activate a set of key transcription factors that determine and regulate the genetic program that induces chondrogenesis and chondrocyte differentiation. Moreover, we describe recent findings regarding the roles of several signaling pathways in modulating the proliferation and maturation of chondrocytes in the growth plate, which is the ‘engine’ of bone elongation. PMID:25715393

  9. Identification of Genes and Pathways Involved in Retinal Neovascularization by Microarray Analysis of Two Animal Models of Retinal Angiogenesis

    PubMed Central

    Xu, Lili; Penn, John S.; Boone, Braden; Dexheimer, Phillip J.

    2010-01-01

    Purpose. Comparative retinal gene expression analysis in two rodent models of oxygen-induced retinopathy (OIR) was performed to identify the genes and pathways involved in retinal neovascularization. Methods. Three independent experimental runs were conducted for each species, according to standard protocols for induction of OIR. Total retinal RNA was isolated at two time points, corresponding to the early response to relative hypoxia (P13 in mouse, P15 in rat) and to the later phase of maximum retinal neovascularization (P18 in mouse, P20 in rat) and was used to prepare labeled probes for hybridization. Gene expression was compared between normal and experimental conditions for each species at each time point. Probesets with a false-discovery rate of ≤0.05 were considered significantly different and were classified as cellular functions or biological pathways. Changes in expression of selected genes were confirmed by quantitative rtPCR. Results. At the early time point, there were changes in 43 genes in each species, with two in common. Increased expression of members of the VEGF and ephrin receptor signaling pathways were identified in both models. At the later time point, there were changes in 26 genes in the rat and in 1622 in the mouse, with 13 in common. Four pathways were identified in both models. Conclusions. Genes and pathways known to be involved in angiogenesis, as well as other biologically plausible genes and pathways, were identified. This work serves as a comprehensive resource for the study of retinal neovascularization and identification of potential rational targets for antiangiogenic therapy. PMID:19834031

  10. MULTI-SENSOR REPORTER CELL TECHNOLOGY TO ASSESS HAZARD INVOLVING ENDOCRINE SIGNALING PATHWAYS

    EPA Science Inventory

    Results will define an experimental approach that can be used in a high-throughput format to evaluate the response of hormone signaling pathways and networks to individual chemicals or mixtures. The assay also will have application across species and would significantly reduce...

  11. The PINK1-Parkin pathway is involved in the regulation of mitochondrial remodeling process

    SciTech Connect

    Park, Jeehye; Lee, Gina; Chung, Jongkyeong

    2009-01-16

    The two Parkinson's disease (PD) genes, PTEN-induced kinase 1 (PINK1) and parkin, are linked in a common pathway which affects mitochondrial integrity and function. However, it is still not known what this pathway does in the mitochondria. Therefore, we investigated its physiological function in Drosophila. Because Drosophila PINK1 and parkin mutants show changes in mitochondrial morphology in both indirect flight muscles and dopaminergic neurons, we here investigated whether the PINK1-Parkin pathway genetically interacts with the regulators of mitochondrial fusion and fission such as Drp1, which promotes mitochondrial fission, and Opa1 or Marf, which induces mitochondrial fusion. Surprisingly, DrosophilaPINK1 and parkin mutant phenotypes were markedly suppressed by overexpression of Drp1 or downregulation of Opa1 or Marf, indicating that the PINK1-Parkin pathway regulates mitochondrial remodeling process in the direction of promoting mitochondrial fission. Therefore, we strongly suggest that mitochondrial fusion and fission process could be a prominent therapeutic target for the treatment of PD.

  12. Clathrin-dependent pathways and the cytoskeleton network are involved in ceramide endocytosis by a parasitic protozoan, Giardia lamblia.

    PubMed

    Hernandez, Yunuen; Castillo, Cynthia; Roychowdhury, Sukla; Hehl, Adrian; Aley, Stephen B; Das, Siddhartha

    2007-01-01

    Although identified as an early-diverged protozoan, Giardia lamblia shares many similarities with higher eukaryotic cells, including an internal membrane system and cytoskeleton, as well as secretory pathways. However, unlike many other eukaryotes, Giardia does not synthesize lipids de novo, but rather depends on exogenous sources for both energy production and organelle or membrane biogenesis. It is not known how lipid molecules are taken up by this parasite and if endocytic pathways are involved in this process. In this investigation, we tested the hypothesis that highly regulated and selective lipid transport machinery is present in Giardia and necessary for the efficient internalization and intracellular targeting of ceramide molecules, the major sphingolipid precursor. Using metabolic and pathway inhibitors, we demonstrate that ceramide is internalized through endocytic pathways and is primarily targeted into perinuclear/endoplasmic reticulum membranes. Further investigations suggested that Giardia uses both clathrin-dependent pathways and the actin cytoskeleton for ceramide uptake, as well as microtubule filaments for intracellular localization and targeting. We speculate that this parasitic protozoan has evolved cytoskeletal and clathrin-dependent endocytic mechanisms for importing ceramide molecules from the cell exterior for the synthesis of membranes and vesicles during growth and differentiation. PMID:17087963

  13. Mvb12 Is a Novel Member of ESCRT-I Involved in Cargo Selection by the Multivesicular Body Pathway

    PubMed Central

    Oestreich, Andrea J.; Davies, Brian A.; Payne, Johanna A.

    2007-01-01

    The multivesicular body (MVB) sorting pathway impacts a variety of cellular functions in eukaryotic cells. Perhaps the best understood role for the MVB pathway is the degradation of transmembrane proteins within the lysosome. Regulation of cargo selection by this pathway is critically important for normal cell physiology, and recent advances in our understanding of this process have highlighted the endosomal sorting complexes required for transport (ESCRTs) as pivotal players in this reaction. To better understand the mechanisms of cargo selection during MVB sorting, we performed a genetic screen to identify novel factors required for cargo-specific selection by this pathway and identified the Mvb12 protein. Loss of Mvb12 function results in differential defects in the selection of MVB cargoes. A variety of analyses indicate that Mvb12 is a stable member of ESCRT-I, a heterologous complex involved in cargo selection by the MVB pathway. Phenotypes displayed upon loss of Mvb12 are distinct from those displayed by the previously described ESCRT-I subunits (vacuolar protein sorting 23, -28, and -37), suggesting a distinct function than these core subunits. These data support a model in which Mvb12 impacts the selection of MVB cargoes by modulating the cargo recognition capabilities of ESCRT-I. PMID:17151358

  14. Acute Psychological Stress Modulates the Expression of Enzymes Involved in the Kynurenine Pathway throughout Corticolimbic Circuits in Adult Male Rats

    PubMed Central

    Vecchiarelli, Haley A.; Gandhi, Chaitanya P.; Hill, Matthew N.

    2016-01-01

    Tryptophan is an essential dietary amino acid that is necessary for protein synthesis, but also serves as the precursor for serotonin. However, in addition to these biological functions, tryptophan also serves as a precursor for the kynurenine pathway, which has neurotoxic (quinolinic acid) and neuroprotective (kynurenic acid) metabolites. Glucocorticoid hormones and inflammatory mediators, both of which are increased by stress, have been shown to bias tryptophan along the kynurenine pathway and away from serotonin synthesis; however, to date, there is no published data regarding the effects of stress on enzymes regulating the kynurenine pathway in a regional manner throughout the brain. Herein, we examined the effects of an acute psychological stress (120 min restraint) on gene expression patterns of enzymes along the kynurenine pathway over a protracted time-course (1–24 h post-stress termination) within the amygdala, hippocampus, hypothalamus, and medial prefrontal cortex. Time-dependent changes in differential enzymes along the kynurenine metabolism pathway, particularly those involved in the production of quinolinic acid, were found within the amygdala, hypothalamus, and medial prefrontal cortex, with no changes seen in the hippocampus. These regional differences acutely may provide mechanistic insight into processes that become dysregulated chronically in stress-associated disorders. PMID:26819772

  15. Shared Genetic Factors Involved in Celiac Disease, Type 2 Diabetes and Anorexia Nervosa Suggest Common Molecular Pathways for Chronic Diseases

    PubMed Central

    Mostowy, Joanna; Montén, Caroline; Gudjonsdottir, Audur H.; Arnell, Henrik; Browaldh, Lars; Nilsson, Staffan; Agardh, Daniel

    2016-01-01

    Background and Objectives Genome-wide association studies (GWAS) have identified several genetic regions involved in immune-regulatory mechanisms to be associated with celiac disease. Previous GWAS also revealed an over-representation of genes involved in type 2 diabetes and anorexia nervosa associated with celiac disease, suggesting involvement of common metabolic pathways for development of these chronic diseases. The aim of this study was to extend these previous analyses to study the gene expression in the gut from children with active celiac disease. Material and Methods Thirty six target genes involved in type 2 diabetes and four genes associated with anorexia nervosa were investigated for gene expression in small intestinal biopsies from 144 children with celiac disease at median (range) age of 7.4 years (1.6–17.8) and from 154 disease controls at a median (range) age 11.4.years (1.4–18.3). Results A total of eleven of genes were differently expressed in celiac patients compared with disease controls of which CD36, CD38, FOXP1, SELL, PPARA, PPARG, AGT previously associated with type 2 diabetes and AKAP6, NTNG1 with anorexia nervosa remained significant after correction for multiple testing. Conclusion Shared genetic factors involved in celiac disease, type 2 diabetes and anorexia nervosa suggest common underlying molecular pathways for these diseases. PMID:27483138

  16. NIK is involved in constitutive activation of the alternative NF-{kappa}B pathway and proliferation of pancreatic cancer cells

    SciTech Connect

    Nishina, Takashi; Yamaguchi, Noritaka; Gohda, Jin; Semba, Kentaro; Inoue, Jun-ichiro

    2009-10-09

    Pancreatic cancer has one of the poorest prognoses among human neoplasms. Constitutive activation of NF-{kappa}B is frequently observed in pancreatic cancer cells and is involved in their malignancy. However, little is known about the molecular mechanism of this constitutive NF-{kappa}B activation. Here, we show that the alternative pathway is constitutively activated and NF-{kappa}B-inducing kinase (NIK), a mediator of the alternative pathway, is significantly expressed in pancreatic cancer cells. siRNA-mediated silencing of NIK expression followed by subcellular fractionation revealed that NIK is constitutively involved in the processing of p100 and nuclear transport of p52 and RelB in pancreatic cancer cells. In addition, NIK silencing significantly suppressed proliferation of pancreatic cancer cells. These results clearly indicate that NIK is involved in the constitutive activation of the alternative pathway and controls cell proliferation in pancreatic cancer cells. Therefore, NIK might be a novel target for the treatment of pancreatic cancer.

  17. Receptors and signaling pathways involved in proliferation and differentiation of Sertoli cells

    PubMed Central

    Lucas, Thaís FG; Nascimento, Aline R; Pisolato, Raisa; Pimenta, Maristela T; Lazari, Maria Fatima M; Porto, Catarina S

    2014-01-01

    The identification of the hormones and other factors regulating Sertoli cell survival, proliferation, and maturation in neonatal, peripubertal, and pubertal life remains one of the most critical questions in testicular biology. The regulation of Sertoli cell proliferation and differentiation is thought to be controlled by cell–cell junctions and a set of circulating and local hormones and growth factors. In this review, we will focus on receptors and intracellular signaling pathways activated by androgen, follicle-stimulating hormone, thyroid hormone, activin, retinoids, insulin, insulin-like growth factor, relaxin, and estrogen, with special emphasis on estrogen receptors. Estrogen receptors activate intracellular signaling pathways that converge on cell cycle and transcription factors and play a role in the regulation of Sertoli cell proliferation and differentiation. PMID:25225624

  18. Second hit in cervical carcinogenesis process: involvement of wnt/beta catenin pathway

    PubMed Central

    Perez-Plasencia, Carlos; Duenas-Gonzalez, Alfonso; Alatorre-Tavera, Brenda

    2008-01-01

    The Human papillomavirus plays an important role in the initiation and progression of cervical cancer. However, it is a necessary but not sufficient cause to develop invasive carcinoma; hence, other factors are required in the pathogenesis of this malignancy. In this review we explore the hypothesis of the deregulation of wnt/β-catenin signaling pathway as a "second hit" required to develop cervical cancer. PMID:18606007

  19. Stimulation of growth by proteorhodopsin phototrophy involves regulation of central metabolic pathways in marine planktonic bacteria

    PubMed Central

    Palovaara, Joakim; Akram, Neelam; Baltar, Federico; Bunse, Carina; Forsberg, Jeremy; Pedrós-Alió, Carlos; González, José M.; Pinhassi, Jarone

    2014-01-01

    Proteorhodopsin (PR) is present in half of surface ocean bacterioplankton, where its light-driven proton pumping provides energy to cells. Indeed, PR promotes growth or survival in different bacteria. However, the metabolic pathways mediating the light responses remain unknown. We analyzed growth of the PR-containing Dokdonia sp. MED134 (where light-stimulated growth had been found) in seawater with low concentrations of mixed [yeast extract and peptone (YEP)] or single (alanine, Ala) carbon compounds as models for rich and poor environments. We discovered changes in gene expression revealing a tightly regulated shift in central metabolic pathways between light and dark conditions. Bacteria showed relatively stronger light responses in Ala compared with YEP. Notably, carbon acquisition pathways shifted toward anaplerotic CO2 fixation in the light, contributing 31 ± 8% and 24 ± 6% of the carbon incorporated into biomass in Ala and YEP, respectively. Thus, MED134 was a facultative double mixotroph, i.e., photo- and chemotrophic for its energy source and using both bicarbonate and organic matter as carbon sources. Unexpectedly, relative expression of the glyoxylate shunt genes (isocitrate lyase and malate synthase) was >300-fold higher in the light—but only in Ala—contributing a more efficient use of carbon from organic compounds. We explored these findings in metagenomes and metatranscriptomes and observed similar prevalence of the glyoxylate shunt compared with PR genes and highest expression of the isocitrate lyase gene coinciding with highest solar irradiance. Thus, regulatory interactions between dissolved organic carbon quality and central metabolic pathways critically determine the fitness of surface ocean bacteria engaging in PR phototrophy. PMID:25136122

  20. Growth hormone activity in mitochondria depends on GH receptor Box 1 and involves caveolar pathway targeting

    SciTech Connect

    Perret-Vivancos, Cecile; Abbate, Aude; Ardail, Dominique; Raccurt, Mireille; Usson, Yves; Lobie, Peter E.; Morel, Gerard . E-mail: gerard.morel@univ-lyon1.fr

    2006-02-01

    Growth hormone (GH) binding to its receptor (GHR) initiates GH-dependent signal transduction and internalization pathways to generate the biological effects. The precise role and way of action of GH on mitochondrial function are not yet fully understood. We show here that GH can stimulate cellular oxygen consumption in CHO cells transfected with cDNA coding for the full-length GHR. By using different GHR cDNA constructs, we succeeded in determining the different parts of the GHR implicated in the mitochondrial response to GH. Polarography and two-photon excitation fluorescence microscopy analysis showed that the Box 1 of the GHR intracellular domain was required for an activation of the mitochondrial respiration in response to a GH exposure. However, confocal laser scanning microscopy demonstrated that cells lacking the GHR Box 1 could efficiently internalize the hormone. We demonstrated that internalization mediated either by clathrin-coated pits or by caveolae was able to regulate GH mitochondrial effect: these two pathways are both essential to obtain the GH stimulatory action on mitochondrial function. Moreover, electron microscopic and biochemical approaches allowed us to identify the caveolar pathway as essential for targeting GH and GHR to mitochondria.

  1. Immune-Mediated Metabolic Kynurenine Pathways Are Involved in the Postoperative Cognitive Dysfunction after Cardiopulmonary Bypass.

    PubMed

    Yi, Shuang Qiang; Yang, Mi; Duan, Kai Ming

    2015-10-01

    Postoperative cognitive dysfunction (POCD) after cardiopulmonary bypass is a serious complication that can lead to personality changes, memory loss, reduction in the ability to learn, and other central nervous system dysfunctions. In recent years, there have been improvements in measures to protect the brain during surgery, although the incidence of cognitive dysfunction after cardiac surgery remains high (33 to 83% short-term and 20 to 60% long-term cognitive dysfunction). Despite the large amount of basic and clinical research on the incidence of POCD, its exact pathogenesis and complexity are not clear. Many studies have shown that the kynurenine pathway (KP) and cognitive function in humans are closely related. Some reports also show that the imbalance of some metabolites of the KP such as kynurenic acid and quinolinic acid (QUIN), which act in dynamic equilibrium under physiologic conditions, have effects on the central nervous system and can significantly affect cognitive function. Further studies have shown that inflammatory mediators may act on key enzymes of the KP causing KP-induced disorders. Severe inflammatory reaction occurs in patients undergoing cardiopulmonary bypass, which triggers metabolic pathways that are closely related to changes in cognitive function. In this review, we summarize that inflammation-induced metabolic kynurenine (KYN) pathway disorders are likely to have an important role in incidence of POCD after CPB surgery. PMID:25893921

  2. Disruption of apoptosis pathways involved in zebrafish gonad differentiation by 17α-ethinylestradiol and fadrozole exposures.

    PubMed

    Luzio, Ana; Matos, Manuela; Santos, Dércia; Fontaínhas-Fernandes, António A; Monteiro, Sandra M; Coimbra, Ana M

    2016-08-01

    Zebrafish (Danio rerio) sex determination seems to involve genetic factors (GSD) but also environmental factors (ESD), such as endocrine disrupting chemicals (EDCs) that are known to mimic endogenous hormones and disrupt gonad differentiation. Apoptosis has also been proposed to play a crucial role in zebrafish gonad differentiation. Nevertheless, the interactions between EDCs and apoptosis have received little attention. Thus, this study aimed to assess if and which apoptotic pathways are involved in zebrafish gonad differentiation and how EDCs may interfere with this process. With these purposes, zebrafish were exposed to 17α-ethinylestradiol (EE2, 4ng/L) and fadrozole (Fad, 50μg/L) from 2h to 35days post-fertilization (dpf). Afterwards, a gene expression analysis by qRT-PCR and a stereological analysis, based on systematic sampling and protein immunohistochemistry, were performed. The death receptors (FAS; TRADD), anti-apoptotic (BCL-2; MDM2), pro-apoptotic (CASP-2 and -6) and cell proliferation (BIRC5/survivin; JUN) genes and proteins were evaluated. In general, apoptosis was inhibited in females through the involvement of anti-apoptotic pathways, while in males apoptosis seemed to be crucial to the failure of the "juvenile ovary" development and the induction of testes transformation. The JUN protein was shown to be necessary in juvenile ovaries, while the BIRC5 protein seemed to be involved in zebrafish spermatogenesis. Both EDCs, EE2 and Fad, increased the apoptosis stimulus in zebrafish gonad. It was noticed that the few females that were resistant to Fad-induced sex reversal had increased anti-apoptotic factor levels, while males exposed to EE2 showed increased pro-apoptotic genes/proteins and were more advanced in gonad differentiation. Overall, our findings show that apoptosis pathways are involved in zebrafish gonad differentiation and that EDCs can disrupt this process. PMID:27337697

  3. Involvement of the Hippo pathway in regeneration and fibrogenesis after ischaemic acute kidney injury: YAP is the key effector.

    PubMed

    Xu, Jing; Li, Pei-Xue; Wu, Jun; Gao, Yi-Jun; Yin, Meng-Xin; Lin, Ye; Yang, Ming; Chen, Dong-Ping; Sun, Hai-Peng; Liu, Zeng-Bo; Gu, Xiang-Chen; Huang, Hong-Ling; Fu, Li-Li; Hu, Hui-Min; He, Liang-Liang; Wu, Wen-Qing; Fei, Zhao-Liang; Ji, Hong-Bin; Zhang, Lei; Mei, Chang-Lin

    2016-03-01

    Renal tubule cells can recover after they undergo AKI (acute kidney injury). An incomplete repair of renal tubules can result in progressive fibrotic CKD (chronic kidney disease). Studies have revealed the relationship between tubular epithelial cells and kidney fibrogenesis. However, the underlying mechanism remains unclear. Hippo pathway components were evaluated in complete/incomplete repair of I/R (ischaemia/reperfusion) AKI rat models, HK-2 cells and AKI human renal biopsy samples. We found that the expression levels of the Hippo pathway components changed dynamically during kidney regeneration and fibrogenesis in rat models of I/R-induced AKI and human renal biopsy samples. The transcription cofactor YAP (Yes-associated protein) might be a key effector of renal regeneration and fibrogenesis. Our results showed further that YAP might elicit both beneficial and detrimental effects on I/R AKI. After I/R injury occurred, YAP could promote the repair of the injured epithelia. The constant YAP increase and activation might be related to interstitial fibrosis and abnormal renal tubule differentiation. These results indicate that the proper modulation of the Hippo pathway, specifically the transcription cofactor YAP, during repair might be a potent therapeutic target in AKI-CKD transition after I/R injury. PMID:26574480

  4. Involvement of the Hippo pathway in regeneration and fibrogenesis after ischaemic acute kidney injury: YAP is the key effector

    PubMed Central

    Xu, Jing; Li, Pei-Xue; Wu, Jun; Gao, Yi-Jun; Yin, Meng-Xin; Lin, Ye; Yang, Ming; Chen, Dong-Ping; Sun, Hai-Peng; Liu, Zeng-Bo; Gu, Xiang-Chen; Huang, Hong-Ling; Fu, Li-Li; Hu, Hui-Min; He, Liang-Liang; Wu, Wen-Qing; Fei, Zhao-Liang; Ji, Hong-Bin

    2015-01-01

    Renal tubule cells can recover after they undergo AKI (acute kidney injury). An incomplete repair of renal tubules can result in progressive fibrotic CKD (chronic kidney disease). Studies have revealed the relationship between tubular epithelial cells and kidney fibrogenesis. However, the underlying mechanism remains unclear. Hippo pathway components were evaluated in complete/incomplete repair of I/R (ischaemia/reperfusion) AKI rat models, HK-2 cells and AKI human renal biopsy samples. We found that the expression levels of the Hippo pathway components changed dynamically during kidney regeneration and fibrogenesis in rat models of I/R-induced AKI and human renal biopsy samples. The transcription cofactor YAP (Yes-associated protein) might be a key effector of renal regeneration and fibrogenesis. Our results showed further that YAP might elicit both beneficial and detrimental effects on I/R AKI. After I/R injury occurred, YAP could promote the repair of the injured epithelia. The constant YAP increase and activation might be related to interstitial fibrosis and abnormal renal tubule differentiation. These results indicate that the proper modulation of the Hippo pathway, specifically the transcription cofactor YAP, during repair might be a potent therapeutic target in AKI–CKD transition after I/R injury. PMID:26574480

  5. Inhibition of Melanogenesis by the Pyridinyl Imidazole Class of Compounds: Possible Involvement of the Wnt/β-Catenin Signaling Pathway

    PubMed Central

    Bellei, Barbara; Pitisci, Angela; Izzo, Enzo; Picardo, Mauro

    2012-01-01

    While investigating the role of p38 MAPK in regulating melanogenesis, we found that pyridinyl imidazole inhibitors class compounds as well as the analog compound SB202474, which does not inhibit p38 MAPK, suppressed both α-MSH-induced melanogenesis and spontaneous melanin synthesis. In this study, we demonstrated that the inhibitory activity of the pyridinyl imidazoles correlates with inhibition of the canonical Wnt/β-catenin pathway activity. Imidazole-treated cells showed a reduction in the level of Tcf/Lef target genes involved in the β-catenin signaling network, including ubiquitous genes such as Axin2, Lef1, and Wisp1 as well as cell lineage-restricted genes such as microphthalmia-associated transcription factor and dopachrome tautomerase. Although over-expression of the Wnt signaling pathway effector β-catenin slightly restored the melanogenic program, the lack of complete reversion suggested that the imidazoles interfered with β-catenin-dependent transcriptional activity rather than with β-catenin expression. Accordingly, we did not observe any significant change in β-catenin protein expression. The independence of p38 MAPK activity from the repression of Wnt/β-catenin signaling pathway was confirmed by small interfering RNA knockdown of p38 MAPK expression, which by contrast, stimulated β-catenin-driven gene expression. Our data demonstrate that the small molecule pyridinyl imidazoles possess two distinct and opposite mechanisms that modulate β-catenin dependent transcription: a p38 inhibition-dependent effect that stimulates the Wnt pathway by increasing β-catenin protein expression and an off-target mechanism that inhibits the pathway by repressing β-catenin protein functionality. The p38-independent effect seems to be dominant and, at least in B16-F0 cells, results in a strong block of the Wnt/β-catenin signaling pathway. PMID:22427932

  6. Involvement of the SLIT/ROBO pathway in follicle development in the fetal ovary.

    PubMed

    Dickinson, Rachel E; Hryhorskyj, Lynn; Tremewan, Hannah; Hogg, Kirsten; Thomson, Axel A; McNeilly, Alan S; Duncan, W Colin

    2010-02-01

    In humans and domestic mammals, pivotal processes in ovary development, including primordial follicle assembly, occur prenatally. These events are essential for determining fertility in adult life; however, they remain poorly understood at the mechanistic level. In mammals, the SLITs (SLIT1, SLIT2 and SLIT3) and their ROBO (ROBO1, ROBO2, ROBO3/RIG-1 and ROBO4/MAGIC ROBO) receptors regulate neural, leukocyte, vascular smooth muscle cell and endothelial cell migration. In addition, the SLIT/ROBO pathway has functional roles in embryonic development and in the adult ovary by inhibiting cell migration and promoting apoptosis. We therefore characterised follicle formation and investigated the expression and localisation of the ROBO/SLIT pathway in the ovine fetal ovary. Using RT-PCR, we identified SLIT2, SLIT3, ROBO1, ROBO2 and ROBO4 in sheep ovaries harvested across gestation. The real-time quantitative PCR results implied that ROBO2 expression and ROBO4 expression were elevated during the early stages of follicle formation and stayed abundant during primordial follicle maturation (P<0.05). Immunohistochemistry examination demonstrated that ROBO1 was localised to the pre-granulosa cells, while ROBO2, ROBO4 and SLIT2 were expressed in the oocytes of the developing primordial follicle. This indicates that in the fetal ovary, SLIT-ROBO signalling may require an autocrine and paracrine interaction. Furthermore, at the time of increased SLIT-ROBO expression, there was a significant reduction in the number of proliferating oocytes in the developing ovary (P<0.0001). Overall, these results suggest, for the first time, that the SLIT-ROBO pathway is expressed at the time of follicle formation during fetal ovary development. PMID:19900988

  7. Involvement of glycolysis/gluconeogenesis and signaling regulatory pathways in Saccharomyces cerevisiae biofilms during fermentation

    PubMed Central

    Li, Zhenjian; Chen, Yong; Liu, Dong; Zhao, Nan; Cheng, Hao; Ren, Hengfei; Guo, Ting; Niu, Huanqing; Zhuang, Wei; Wu, Jinglan; Ying, Hanjie

    2015-01-01

    Compared to free (free-living) cells, biofilm cells show increased resistance and stability to high-pressure fermentation conditions, although the reasons underlying these phenomena remain unclear. Here, we investigated biofilm formation with immobilized Saccharomyces cerevisiae cells grown on fiber surfaces during the process of ethanol fermentation. The development of biofilm colonies was visualized by fluorescent labeling and confocal microscopy. RNA from yeast cells at three different biofilm development periods was extracted and sequenced by high-throughput sequencing. We quantitated gene expression differences between biofilm cells and free cells and found that 2098, 1556, and 927 genes were significantly differentially expressed, respectively. We also validated the expression of previously reported genes and identified novel genes and pathways under the control of this system. Statistical analysis revealed that biofilm genes show significant gene expression changes principally in the initial period of biofilm formation compared to later periods. Carbohydrate metabolism, amino acid metabolism, signal transduction, and oxidoreductase activity were needed for biofilm formation. In contrast to previous findings, we observed some differential expression performances of FLO family genes, indicating that cell aggregation in our immobilized fermentation system was possibly independent of flocculation. Cyclic AMP-protein kinase A and mitogen-activated protein kinase pathways regulated signal transduction pathways during yeast biofilm formation. We found that carbohydrate metabolism, especially glycolysis/gluconeogenesis, played a key role in the development of S. cerevisiae biofilms. This work provides an important dataset for future studies aimed at gaining insight into the regulatory mechanisms of immobilized cells in biofilms, as well as for optimizing bioprocessing applications with S. cerevisiae. PMID:25755652

  8. Identification and Pathway Analysis of microRNAs with No Previous Involvement in Breast Cancer

    PubMed Central

    Rebollar-Vega, Rosa; Quintanar-Jurado, Valeria; Maffuz-Aziz, Antonio; Jimenez-Sanchez, Gerardo; Bautista-Piña, Veronica; Arellano-Llamas, Rocio; Hidalgo-Miranda, Alfredo

    2012-01-01

    microRNA expression signatures can differentiate normal and breast cancer tissues and can define specific clinico-pathological phenotypes in breast tumors. In order to further evaluate the microRNA expression profile in breast cancer, we analyzed the expression of 667 microRNAs in 29 tumors and 21 adjacent normal tissues using TaqMan Low-density arrays. 130 miRNAs showed significant differential expression (adjusted P value = 0.05, Fold Change = 2) in breast tumors compared to the normal adjacent tissue. Importantly, the role of 43 of these microRNAs has not been previously reported in breast cancer, including several evolutionary conserved microRNA*, showing similar expression rates to that of their corresponding leading strand. The expression of 14 microRNAs was replicated in an independent set of 55 tumors. Bioinformatic analysis of mRNA targets of the altered miRNAs, identified oncogenes like ERBB2, YY1, several MAP kinases, and known tumor-suppressors like FOXA1 and SMAD4. Pathway analysis identified that some biological process which are important in breast carcinogenesis are affected by the altered microRNA expression, including signaling through MAP kinases and TP53 pathways, as well as biological processes like cell death and communication, focal adhesion and ERBB2-ERBB3 signaling. Our data identified the altered expression of several microRNAs whose aberrant expression might have an important impact on cancer-related cellular pathways and whose role in breast cancer has not been previously described. PMID:22438871

  9. Accommodation of aminoacyl-tRNA into the ribosome involves reversible excursions along multiple pathways

    PubMed Central

    Whitford, Paul C.; Geggier, Peter; Altman, Roger B.; Blanchard, Scott C.; Onuchic, José N.; Sanbonmatsu, Karissa Y.

    2010-01-01

    The ribosome is a massive ribonucleoprotein complex (∼2.4 MDa) that utilizes large-scale structural fluctuations to produce unidirectional protein synthesis. Accommodation is a key conformational change during transfer RNA (tRNA) selection that allows movement of tRNA into the ribosome. Here, we address the structure–function relationship that governs accommodation using all-atom molecular simulations and single-molecule fluorescence resonance energy transfer (smFRET). Simulations that employ an all-atom, structure-based (Gō-like) model illuminate the interplay between configurational entropy and effective enthalpy during the accommodation process. This delicate balance leads to spontaneous reversible accommodation attempts, which are corroborated by smFRET measurements. The dynamics about the endpoints of accommodation (the A/T and A/A conformations) obtained from structure-based simulations are validated by multiple 100–200 ns explicit-solvent simulations (3.2 million atoms for a cumulative 1.4 μs), and previous crystallographic analysis. We find that the configurational entropy of the 3′-CCA end of aminoacyl-tRNA resists accommodation, leading to a multistep accommodation process that encompasses a distribution of parallel pathways. The calculated mechanism is robust across simulation methods and protocols, suggesting that the structure of the accommodation corridor imposes stringent limitations on the accessible pathways. The identified mechanism and observed parallel pathways establish an atomistic framework for interpreting a large body of biochemical data and demonstrate that conformational changes during translation occur through a stochastic trial-and-error process, rather than in concerted lock-step motions. PMID:20427512

  10. Accommodation of aminoacyl-tRNA into the ribosome involves reversible excursions along multiple pathways.

    PubMed

    Whitford, Paul C; Geggier, Peter; Altman, Roger B; Blanchard, Scott C; Onuchic, José N; Sanbonmatsu, Karissa Y

    2010-06-01

    The ribosome is a massive ribonucleoprotein complex ( approximately 2.4 MDa) that utilizes large-scale structural fluctuations to produce unidirectional protein synthesis. Accommodation is a key conformational change during transfer RNA (tRNA) selection that allows movement of tRNA into the ribosome. Here, we address the structure-function relationship that governs accommodation using all-atom molecular simulations and single-molecule fluorescence resonance energy transfer (smFRET). Simulations that employ an all-atom, structure-based (Gō-like) model illuminate the interplay between configurational entropy and effective enthalpy during the accommodation process. This delicate balance leads to spontaneous reversible accommodation attempts, which are corroborated by smFRET measurements. The dynamics about the endpoints of accommodation (the A/T and A/A conformations) obtained from structure-based simulations are validated by multiple 100-200 ns explicit-solvent simulations (3.2 million atoms for a cumulative 1.4 micros), and previous crystallographic analysis. We find that the configurational entropy of the 3'-CCA end of aminoacyl-tRNA resists accommodation, leading to a multistep accommodation process that encompasses a distribution of parallel pathways. The calculated mechanism is robust across simulation methods and protocols, suggesting that the structure of the accommodation corridor imposes stringent limitations on the accessible pathways. The identified mechanism and observed parallel pathways establish an atomistic framework for interpreting a large body of biochemical data and demonstrate that conformational changes during translation occur through a stochastic trial-and-error process, rather than in concerted lock-step motions. PMID:20427512