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Sample records for pathways regulate tnf-a

  1. Multiple pathways regulate shoot branching

    PubMed Central

    Rameau, Catherine; Bertheloot, Jessica; Leduc, Nathalie; Andrieu, Bruno; Foucher, Fabrice; Sakr, Soulaiman

    2015-01-01

    Shoot branching patterns result from the spatio-temporal regulation of axillary bud outgrowth. Numerous endogenous, developmental and environmental factors are integrated at the bud and plant levels to determine numbers of growing shoots. Multiple pathways that converge to common integrators are most probably involved. We propose several pathways involving not only the classical hormones auxin, cytokinins and strigolactones, but also other signals with a strong influence on shoot branching such as gibberellins, sugars or molecular actors of plant phase transition. We also deal with recent findings about the molecular mechanisms and the pathway involved in the response to shade as an example of an environmental signal controlling branching. We propose the TEOSINTE BRANCHED1, CYCLOIDEA, PCF transcription factor TB1/BRC1 and the polar auxin transport stream in the stem as possible integrators of these pathways. We finally discuss how modeling can help to represent this highly dynamic system by articulating knowledges and hypothesis and calculating the phenotype properties they imply. PMID:25628627

  2. Informatics approaches to understanding TGFβ pathway regulation

    PubMed Central

    Kahlem, Pascal; Newfeld, Stuart J.

    2009-01-01

    Summary In recent years, informatics studies have predicted several new ways in which the transforming growth factor β (TGFβ) signaling pathway can be post-translationally regulated. Subsequently, many of these predictions were experimentally validated. These approaches include phylogenetic predictions for the phosphorylation, sumoylation and ubiquitylation of pathway components, as well as kinetic models of endocytosis, phosphorylation and nucleo-cytoplasmic shuttling. We review these studies and provide a brief `how to' guide for phylogenetics. Our hope is to stimulate experimental tests of informatics-based predictions for TGFβ signaling, as well as for other signaling pathways, and to expand the number of developmental pathways that are being analyzed computationally. PMID:19855015

  3. Light regulation of metabolic pathways in fungi.

    PubMed

    Tisch, Doris; Schmoll, Monika

    2010-02-01

    Light represents a major carrier of information in nature. The molecular machineries translating its electromagnetic energy (photons) into the chemical language of cells transmit vital signals for adjustment of virtually every living organism to its habitat. Fungi react to illumination in various ways, and we found that they initiate considerable adaptations in their metabolic pathways upon growth in light or after perception of a light pulse. Alterations in response to light have predominantly been observed in carotenoid metabolism, polysaccharide and carbohydrate metabolism, fatty acid metabolism, nucleotide and nucleoside metabolism, and in regulation of production of secondary metabolites. Transcription of genes is initiated within minutes, abundance and activity of metabolic enzymes are adjusted, and subsequently, levels of metabolites are altered to cope with the harmful effects of light or to prepare for reproduction, which is dependent on light in many cases. This review aims to give an overview on metabolic pathways impacted by light and to illustrate the physiological significance of light for fungi. We provide a basis for assessment whether a given metabolic pathway might be subject to regulation by light and how these properties can be exploited for improvement of biotechnological processes.

  4. WNK signalling pathways in blood pressure regulation.

    PubMed

    Murthy, Meena; Kurz, Thimo; O'Shaughnessy, Kevin M

    2017-04-01

    Hypertension (high blood pressure) is a major public health problem affecting more than a billion people worldwide with complications, including stroke, heart failure and kidney failure. The regulation of blood pressure is multifactorial reflecting genetic susceptibility, in utero environment and external factors such as obesity and salt intake. In keeping with Arthur Guyton's hypothesis, the kidney plays a key role in blood pressure control and data from clinical studies; physiology and genetics have shown that hypertension is driven a failure of the kidney to excrete excess salt at normal levels of blood pressure. There is a number of rare Mendelian blood pressure syndromes, which have shed light on the molecular mechanisms involved in dysregulated ion transport in the distal kidney. One in particular is Familial hyperkalemic hypertension (FHHt), an autosomal dominant monogenic form of hypertension characterised by high blood pressure, hyperkalemia, hyperchloremic metabolic acidosis, and hypercalciuria. The clinical signs of FHHt are treated by low doses of thiazide diuretic, and it mirrors Gitelman syndrome which features the inverse phenotype of hypotension, hypokalemic metabolic alkalosis, and hypocalciuria. Gitelman syndrome is caused by loss of function mutations in the thiazide-sensitive Na/Cl cotransporter (NCC); however, FHHt patients do not have mutations in the SCL12A3 locus encoding NCC. Instead, mutations have been identified in genes that have revealed a key signalling pathway that regulates NCC and several other key transporters and ion channels in the kidney that are critical for BP regulation. This is the WNK kinase signalling pathway that is the subject of this review.

  5. [Enrichment of extracellular DNA from the cultivation medium of human peripheral blood mononuclears with genomic CpG rich fragments results in increased cell production of IL-6 and TNF-a via activation of the NF-kB signaling pathway].

    PubMed

    Speranskii, A I; Kostyuk, S V; Kalashnikova, E A; Veiko, N N

    2016-03-01

    Previously, it was found that blood plasma extracellular DNA (ecDNA) of patients with rheumatoid arthritis (RA) is enriched with CpG-rich genomic DNA fragments, which contain TLR9 ligands (Veiko et al., 2006). In this study we have demonstrated that ecDNA of a RA patient and model fragments added to a cultivation medium of peripheral blood mononuclear cells (PBMC) of healthy donors stimulate expression of genes for the TLR9-MyD88-NF-kB signaling pathway; this leads to a significant increase in concentrations of the proinflammatory cytokines IL-6 and TNF-a in the cultivation medium. Human genomic DNA non-enriched with the CpG sequences did not stimulate IL-6 and TNF-a synthesis in PBMC. A scheme explaining the potential role ecDNA in the induction and maintenance of increased levels of the proinflammatory cytokines under conditions damaging the human cells has been proposed.

  6. [Pathways of flowering regulation in plants].

    PubMed

    Liu, Yongping; Yang, Jing; Yang, Mingfeng

    2015-11-01

    Flowering, the floral transition from vegetative growth to reproductive growth, is induced by diverse endogenous and exogenous cues, such as photoperiod, temperature, hormones and age. Precise flowering time is critical to plant growth and evolution of species. The numerous renewal molecular and genetic results have revealed five flowering time pathways, including classical photoperiod pathway, vernalization pathway, autonomous pathway, gibberellins (GA) pathway and newly identified age pathway. These pathways take on relatively independent role, and involve extensive crosstalks and feedback loops. This review describes the complicated regulatory network of this floral transition to understand the molecular mechanism of flowering and provide references for further research in more plants.

  7. Master regulators, regulatory networks, and pathways of glioblastoma subtypes.

    PubMed

    Bozdag, Serdar; Li, Aiguo; Baysan, Mehmet; Fine, Howard A

    2014-01-01

    Glioblastoma multiforme (GBM) is the most common malignant brain tumor. GBM samples are classified into subtypes based on their transcriptomic and epigenetic profiles. Despite numerous studies to better characterize GBM biology, a comprehensive study to identify GBM subtype- specific master regulators, gene regulatory networks, and pathways is missing. Here, we used FastMEDUSA to compute master regulators and gene regulatory networks for each GBM subtype. We also ran Gene Set Enrichment Analysis and Ingenuity Pathway Analysis on GBM expression dataset from The Cancer Genome Atlas Project to compute GBM- and GBM subtype-specific pathways. Our analysis was able to recover some of the known master regulators and pathways in GBM as well as some putative novel regulators and pathways, which will aide in our understanding of the unique biology of GBM subtypes.

  8. Signaling pathways involved in MDSC regulation.

    PubMed

    Trikha, Prashant; Carson, William E

    2014-08-01

    The immune system has evolved mechanisms to protect the host from the deleterious effects of inflammation. The generation of immune suppressive cells like myeloid derived suppressor cells (MDSCs) that can counteract T cell responses represents one such strategy. There is an accumulation of immature myeloid cells or MDSCs in bone marrow (BM) and lymphoid organs under pathological conditions such as cancer. MDSCs represent a population of heterogeneous myeloid cells comprising of macrophages, granulocytes and dendritic cells that are at early stages of development. Although, the precise signaling pathways and molecular mechanisms that lead to MDSC generation and expansion in cancer remains to be elucidated. It is widely believed that perturbation of signaling pathways involved during normal hematopoietic and myeloid development under pathological conditions such as tumorogenesis contributes to the development of suppressive myeloid cells. In this review we discuss the role played by key signaling pathways such as PI3K, Ras, Jak/Stat and TGFb during myeloid development and how their deregulation under pathological conditions can lead to the generation of suppressive myeloid cells or MDSCs. Targeting these pathways should help in elucidating mechanisms that lead to the expansion of MDSCs in cancer and point to methods for eliminating these cells from the tumor microenvironment.

  9. WW domain interactions regulate the Hippo tumor suppressor pathway

    PubMed Central

    Salah, Z; Aqeilan, R I

    2011-01-01

    The Hippo kinase pathway is emerging as a conserved signaling pathway that is essential for organ growth and tumorigenesis in Drosophila and mammalians. Although the signaling of the core kinases is relatively well understood, less is known about the upstream inputs, downstream outputs and regulation of the whole cascade. Enrichment of the Hippo pathway components with WW domains and their cognate proline-rich interacting motifs provides a versatile platform for further understanding the mechanisms that regulate organ growth and tumorigenesis. Here, we review recently discovered mechanisms of WW domain-mediated interactions that contribute to the regulation of the Hippo signaling pathway in tumorigenesis. We further discuss new insights and future directions on the emerging role of such regulation. PMID:21677687

  10. WW domain interactions regulate the Hippo tumor suppressor pathway.

    PubMed

    Salah, Z; Aqeilan, R I

    2011-06-16

    The Hippo kinase pathway is emerging as a conserved signaling pathway that is essential for organ growth and tumorigenesis in Drosophila and mammalians. Although the signaling of the core kinases is relatively well understood, less is known about the upstream inputs, downstream outputs and regulation of the whole cascade. Enrichment of the Hippo pathway components with WW domains and their cognate proline-rich interacting motifs provides a versatile platform for further understanding the mechanisms that regulate organ growth and tumorigenesis. Here, we review recently discovered mechanisms of WW domain-mediated interactions that contribute to the regulation of the Hippo signaling pathway in tumorigenesis. We further discuss new insights and future directions on the emerging role of such regulation.

  11. The Hippo pathway regulates hematopoiesis in Drosophila melanogaster.

    PubMed

    Milton, Claire C; Grusche, Felix A; Degoutin, Joffrey L; Yu, Eefang; Dai, Qi; Lai, Eric C; Harvey, Kieran F

    2014-11-17

    The Salvador-Warts-Hippo (Hippo) pathway is an evolutionarily conserved regulator of organ growth and cell fate. It performs these functions in epithelial and neural tissues of both insects and mammals, as well as in mammalian organs such as the liver and heart. Despite rapid advances in Hippo pathway research, a definitive role for this pathway in hematopoiesis has remained enigmatic. The hematopoietic compartments of Drosophila melanogaster and mammals possess several conserved features. D. melanogaster possess three types of hematopoietic cells that most closely resemble mammalian myeloid cells: plasmatocytes (macrophage-like cells), crystal cells (involved in wound healing), and lamellocytes (which encapsulate parasites). The proteins that control differentiation of these cells also control important blood lineage decisions in mammals. Here, we define the Hippo pathway as a key mediator of hematopoiesis by showing that it controls differentiation and proliferation of the two major types of D. melanogaster blood cells, plasmatocytes and crystal cells. In animals lacking the downstream Hippo pathway kinase Warts, lymph gland cells overproliferated, differentiated prematurely, and often adopted a mixed lineage fate. The Hippo pathway regulated crystal cell numbers by both cell-autonomous and non-cell-autonomous mechanisms. Yorkie and its partner transcription factor Scalloped were found to regulate transcription of the Runx family transcription factor Lozenge, which is a key regulator of crystal cell fate. Further, Yorkie or Scalloped hyperactivation induced ectopic crystal cells in a non-cell-autonomous and Notch-pathway-dependent fashion.

  12. Mevalonate Pathway Regulates Cell Size Homeostasis and Proteostasis through Autophagy

    PubMed Central

    Miettinen, Teemu P.; Björklund, Mikael

    2015-01-01

    Summary Balance between cell growth and proliferation determines cell size homeostasis, but little is known about how metabolic pathways are involved in the maintenance of this balance. Here, we perform a screen with a library of clinically used drug molecules for their effects on cell size. We find that statins, inhibitors of the mevalonate pathway, reduce cell proliferation and increase cell size and cellular protein density in various cell types, including primary human cells. Mevalonate pathway effects on cell size and protein density are mediated through geranylgeranylation of the small GTPase RAB11, which is required for basal autophagic flux. Our results identify the mevalonate pathway as a metabolic regulator of autophagy and expose a paradox in the regulation of cell size and proteostasis, where inhibition of an anabolic pathway can cause an increase in cell size and cellular protein density. PMID:26686643

  13. PIKfyve Regulation of Endosome-Linked Pathways

    PubMed Central

    de Lartigue, Jane; Polson, Hannah; Feldman, Morri; Shokat, Kevan; Tooze, Sharon A; Urbé, Sylvie; Clague, Michael J

    2009-01-01

    The phosphoinositide 5-kinase (PIKfyve) is a critical enzyme for the synthesis of PtdIns(3,5)P2, that has been implicated in various trafficking events associated with the endocytic pathway. We have now directly compared the effects of siRNA-mediated knockdown of PIKfyve in HeLa cells with a specific pharmacological inhibitor of enzyme activity. Both approaches induce changes in the distribution of CI-M6PR and trans-Golgi network (TGN)-46 proteins, which cycles between endosomes and TGN, leading to their accumulation in dispersed punctae, whilst the TGN marker golgin-245 retains a perinuclear disposition. Trafficking of CD8-CI-M6PR (retromer-dependent) and CD8-Furin (retromer-independent) chimeras from the cell surface to the TGN is delayed following drug administration, as is the transport of the Shiga toxin B-subunit. siRNA knockdown of PIKfyve produced no defect in epidermal growth factor receptor (EGFR) degradation, unless combined with knockdown of its activator molecule Vac14, suggesting that a low threshold of PtdIns(3,5)P2 is necessary and sufficient for this pathway. Accordingly pharmacological inhibition of PIKfyve results in a profound block to the lysosomal degradation of activated epidermal growth factor (EGF) and Met receptors. Immunofluorescence revealed EGF receptors to be trapped in the interior of a swollen endosomal compartment. In cells starved of amino acids, PIKfyve inhibition leads to the accumulation of the lipidated form of GFP-LC3, a marker of autophagosomal structures, which can be visualized as fluorescent punctae. We suggest that PIKfyve inhibition may render the late endosome/lysosome compartment refractory to fusion with both autophagosomes and with EGFR-containing multivesicular bodies. PMID:19582903

  14. Profiling of molecular pathways regulated by microRNA 601.

    PubMed

    Ohdaira, Hiroaki; Nakagawa, Hiroki; Yoshida, Kenichi

    2009-12-01

    MicroRNAs (miRNAs) have been implicated in complex vertebrate developmental and pathological systems as a versatile class of molecules involved in the regulation of various biological processes and molecular pathways. To elucidate the role of miRNAs in human somatic cells, an understanding of the molecular framework regulated by individual miRNA is essential. In this study, we examined the effect of hsa-miR-601 on gene expression changes in human lung cancer cells A549. To achieve this, DNA microarray and global pathway analyses were performed on hsa-miR-601 introduced cells for two successive days. Gene ontology analysis revealed that the effect of hsa-miR-601 over-represented the negative regulation of translation/translational initiation, whereas GenMAPP analysis revealed that several characteristic pathways were changed in hsa-miR-601 introduced A549 cells compared to control short RNA introduced cells. Among them, up-regulation of actin cytoskeleton and down-regulation of Fas-induced apoptosis pathway occurred on two successive days after hsa-miR-601 introduction. Using a luciferase reporter assay, we also showed that hsa-miR-601 specifically repressed nuclear factor-kappaB (NF-kappaB) transcription factor-dependent reporter expression, a key component of the immune-oncogenesis pathway. These findings suggest that hsa-miR-601 could affect a variety of signaling pathways accompanying orchestrated gene expression changes. Our results argue that individual miRNAs affect complex regulation of cellular signaling pathways.

  15. Neurotrophin signalling pathways regulating neuronal apoptosis.

    PubMed

    Miller, F D; Kaplan, D R

    2001-07-01

    Recent evidence indicates that naturally occurring neuronal death in mammals is regulated by the interplay between receptor-mediated prosurvival and proapoptotic signals. The neurotrophins, a family of growth factors best known for their positive effects on neuronal biology, have now been shown to mediate both positive and negative survival signals, by signalling through the Trk and p75 neurotrophin receptors, respectively. The mechanisms whereby these two neurotrophin receptors interact to determine neuronal survival have been difficult to decipher, largely because both can signal independently or coincidentally, depending upon the cell or developmental context. Nonetheless, the past several years have seen significant advances in our understanding of this receptor signalling system. In this review, we focus on the proapoptotic actions of the p75 neurotrophin receptor (p75NTR), and on the interplay between Trk and p75NTR that determines neuronal survival.

  16. The N-end rule pathway and regulation by proteolysis

    PubMed Central

    Varshavsky, Alexander

    2011-01-01

    The N-end rule relates the regulation of the in vivo half-life of a protein to the identity of its N-terminal residue. Degradation signals (degrons) that are targeted by the N-end rule pathway include a set called N-degrons. The main determinant of an N-degron is a destabilizing N-terminal residue of a protein. In eukaryotes, the N-end rule pathway is a part of the ubiquitin system and consists of two branches, the Ac/N-end rule and the Arg/N-end rule pathways. The Ac/N-end rule pathway targets proteins containing Nα-terminally acetylated (Nt-acetylated) residues. The Arg/N-end rule pathway recognizes unacetylated N-terminal residues and involves N-terminal arginylation. Together, these branches target for degradation a majority of cellular proteins. For example, more than 80% of human proteins are cotranslationally Nt-acetylated. Thus, most proteins harbor a specific degradation signal, termed AcN-degron, from the moment of their birth. Specific N-end rule pathways are also present in prokaryotes and in mitochondria. Enzymes that produce N-degrons include methionine-aminopeptidases, caspases, calpains, Nt-acetylases, Nt-amidases, arginyl-transferases, and leucyl-transferases. Regulated degradation of specific proteins by the N-end rule pathway mediates a legion of physiological functions, including the sensing of heme, oxygen, and nitric oxide; selective elimination of misfolded proteins; the regulation of DNA repair, segregation, and condensation; the signaling by G proteins; the regulation of peptide import, fat metabolism, viral and bacterial infections, apoptosis, meiosis, spermatogenesis, neurogenesis, and cardiovascular development; and the functioning of adult organs, including the pancreas and the brain. Discovered 25 years ago, this pathway continues to be a fount of biological insights. PMID:21633985

  17. Engineering dynamic pathway regulation using stress-response promoters.

    PubMed

    Dahl, Robert H; Zhang, Fuzhong; Alonso-Gutierrez, Jorge; Baidoo, Edward; Batth, Tanveer S; Redding-Johanson, Alyssa M; Petzold, Christopher J; Mukhopadhyay, Aindrila; Lee, Taek Soon; Adams, Paul D; Keasling, Jay D

    2013-11-01

    Heterologous pathways used in metabolic engineering may produce intermediates toxic to the cell. Dynamic control of pathway enzymes could prevent the accumulation of these metabolites, but such a strategy requires sensors, which are largely unknown, that can detect and respond to the metabolite. Here we applied whole-genome transcript arrays to identify promoters that respond to the accumulation of toxic intermediates, and then used these promoters to control accumulation of the intermediate and improve the final titers of a desired product. We apply this approach to regulate farnesyl pyrophosphate (FPP) production in the isoprenoid biosynthetic pathway in Escherichia coli. This strategy improved production of amorphadiene, the final product, by twofold over that from inducible or constitutive promoters, eliminated the need for expensive inducers, reduced acetate accumulation and improved growth. We extended this approach to another toxic intermediate to demonstrate the broad utility of identifying novel sensor-regulator systems for dynamic regulation.

  18. Stress responses during ageing: molecular pathways regulating protein homeostasis.

    PubMed

    Kyriakakis, Emmanouil; Princz, Andrea; Tavernarakis, Nektarios

    2015-01-01

    The ageing process is characterized by deterioration of physiological function accompanied by frailty and ageing-associated diseases. The most broadly and well-studied pathways influencing ageing are the insulin/insulin-like growth factor 1 signaling pathway and the dietary restriction pathway. Recent studies in diverse organisms have also delineated emerging pathways, which collectively or independently contribute to ageing. Among them the proteostatic-stress-response networks, inextricably affect normal ageing by maintaining or restoring protein homeostasis to preserve proper cellular and organismal function. In this chapter, we survey the involvement of heat stress and endoplasmic reticulum stress responses in the regulation of longevity, placing emphasis on the cross talk between different response mechanisms and their systemic effects. We further discuss novel insights relevant to the molecular pathways mediating these stress responses that may facilitate the development of innovative interventions targeting age-related pathologies such as diabetes, cancer, cardiovascular and neurodegenerative diseases.

  19. Bacterial Transcriptional Regulators for Degradation Pathways of Aromatic Compounds

    PubMed Central

    Tropel, David; van der Meer, Jan Roelof

    2004-01-01

    Human activities have resulted in the release and introduction into the environment of a plethora of aromatic chemicals. The interest in discovering how bacteria are dealing with hazardous environmental pollutants has driven a large research community and has resulted in important biochemical, genetic, and physiological knowledge about the degradation capacities of microorganisms and their application in bioremediation, green chemistry, or production of pharmacy synthons. In addition, regulation of catabolic pathway expression has attracted the interest of numerous different groups, and several catabolic pathway regulators have been exemplary for understanding transcription control mechanisms. More recently, information about regulatory systems has been used to construct whole-cell living bioreporters that are used to measure the quality of the aqueous, soil, and air environment. The topic of biodegradation is relatively coherent, and this review presents a coherent overview of the regulatory systems involved in the transcriptional control of catabolic pathways. This review summarizes the different regulatory systems involved in biodegradation pathways of aromatic compounds linking them to other known protein families. Specific attention has been paid to describing the genetic organization of the regulatory genes, promoters, and target operon(s) and to discussing present knowledge about signaling molecules, DNA binding properties, and operator characteristics, and evidence from regulatory mutants. For each regulator family, this information is combined with recently obtained protein structural information to arrive at a possible mechanism of transcription activation. This demonstrates the diversity of control mechanisms existing in catabolic pathways. PMID:15353566

  20. Eosinophil granule cationic proteins regulate the classical pathway of complement.

    PubMed Central

    Weiler, J M; Edens, R E; Bell, C S; Gleich, G J

    1995-01-01

    Major basic protein, the primary constituent of eosinophil granules, regulates the alternative and classical pathways of complement. Major basic protein and other eosinophil granule cationic proteins, which are important in mediating tissue damage in allergic disease, regulate the alternative pathway by interfering with C3b interaction with factor B to assemble an alternative pathway C3 convertase. In the present study, eosinophil peroxidase, eosinophil cationic protein and eosinophil-derived neurotoxin, as well as major basic protein, were examined for capacity to regulate the classical pathway. Eosinophil peroxidase, eosinophil cationic protein and major basic protein inhibited formation of cell-bound classical pathway C3 convertase (EAC1,4b,2a), causing 50% inhibition of complement-mediated lysis at about 0.19, 0.75 and 0.5 micrograms/10(7) cellular intermediates, respectively. Eosinophil-derived neurotoxin had no activity on this pathway of complement. The eosinophil granule proteins were examined for activity on the formation of the membrane attack complex. Major basic protein and eosinophil cationic protein had no activity on terminal lysis. In contrast, eosinophil peroxidase inhibited lysis of EAC1,4b,2a,3b,5b, but had only minimal activity on later events in complement lysis. These polycations were then examined to determine the site(s) at which they regulated the early classical pathway. Eosinophil granule polycationic proteins: (1) reduced the Zmax at all time points but had only minimal effect on the Tmax during the formation of the classical pathway C3 convertase (EAC1,4b,2a); (2) inhibited formation of EAC1,4b,2a proportional to C4 but independent of C2 concentration; (3) inhibited fluid phase formation of C1,4b,2a, as reflected by a decrease in C1-induced consumption of C2 over time; and (4) inhibited C1 activity over time without a direct effect on either C4 or C2. These observations suggest that polycations regulate the early classical pathway by

  1. Regulation of methanol utilisation pathway genes in yeasts

    PubMed Central

    Hartner, Franz S; Glieder, Anton

    2006-01-01

    Methylotrophic yeasts such as Candida boidinii, Hansenula polymorpha, Pichia methanolica and Pichia pastoris are an emerging group of eukaryotic hosts for recombinant protein production with an ever increasing number of applications during the last 30 years. Their applications are linked to the use of strong methanol-inducible promoters derived from genes of the methanol utilisation pathway. These promoters are tightly regulated, highly repressed in presence of non-limiting concentrations of glucose in the medium and strongly induced if methanol is used as carbon source. Several factors involved in this tight control and their regulatory effects have been described so far. This review summarises available data about the regulation of promoters from methanol utilisation pathway genes. Furthermore, the role of cis and trans acting factors (e.g. transcription factors, glucose processing enzymes) in the expression of methanol utilisation pathway genes is reviewed both in the context of the native cell environment as well as in heterologous hosts. PMID:17169150

  2. Control and regulation of pathways via negative feedback

    PubMed Central

    2017-01-01

    The biochemical networks found in living organisms include a huge variety of control mechanisms at multiple levels of organization. While the mechanistic and molecular details of many of these control mechanisms are understood, their exact role in driving cellular behaviour is not. For example, yeast glycolysis has been studied for almost 80 years but it is only recently that we have come to understand the systemic role of the multitude of feedback and feed-forward controls that exist in this pathway. In this article, control theory is discussed as an approach to dissect the control logic of complex pathways. One of the key issues is distinguishing between the terms control and regulation and how these concepts are applied to regulated enzymes such as phosphofructokinase. In doing so, one of the paradoxes in metabolic regulation can be resolved where enzymes such as phosphofructokinase have little control but, nevertheless, possess significant regulatory influence. PMID:28202588

  3. Transcriptional Regulation of Carbohydrate Utilization Pathways in the Bifidobacterium Genus.

    PubMed

    Khoroshkin, Matvei S; Leyn, Semen A; Van Sinderen, Douwe; Rodionov, Dmitry A

    2016-01-01

    Bifidobacteria, which represent common commensals of mammalian gut, are believed to have positive effects on human health. The influence of certain non-digestible carbohydrates (and their use as so-called prebiotics) on growth and metabolic activity of bifidobacteria is of increasing interest; however, mechanisms of transcriptional control of carbohydrate metabolism are poorly understood in these species. We used a comparative genomics approach to reconstruct carbohydrate utilization pathways and transcriptional regulons in 10 Bifidobacterium genomes. Analysis of regulatory gene regions revealed candidate DNA motifs and reconstructed regulons for 268 transcription factors from the LacI, ROK, DeoR, AraC, GntR, and TetR families that form 64 orthologous groups of regulators. Most of the reconstructed regulons are local and control specific catabolic pathways for host- and diet-derived glycans and monosaccharides. Mosaic distributions of many of these local regulators across Bifidobacterium species correlate with distribution of corresponding catabolic pathways. In contrast, the maltose, galactose, sucrose, and fructose regulons, as well as a novel global LacI-family regulator that is predicted to control the central carbohydrate metabolism and arabinose catabolism genes, are universally present in all 10 studied bifidobacteria. A novel group of TetR-family regulators presumably controls the glucoside and galactoside utilization pathways. Paralogs of the ribose repressor RbsR control the pyrimidine nucleoside utilization genes. Multiple paralogs of the maltose regulator MalR co-regulate large sets of genes involved in maltodextrin utilization. The inferred metabolic regulons provide new insights on diverse carbohydrate utilization networks in bifidobacteria that can be employed in metabolic modeling, phenotype prediction and the rational development of novel prebiotics.

  4. Transcriptional Regulation of Carbohydrate Utilization Pathways in the Bifidobacterium Genus

    PubMed Central

    Khoroshkin, Matvei S.; Leyn, Semen A.; Van Sinderen, Douwe; Rodionov, Dmitry A.

    2016-01-01

    Bifidobacteria, which represent common commensals of mammalian gut, are believed to have positive effects on human health. The influence of certain non-digestible carbohydrates (and their use as so-called prebiotics) on growth and metabolic activity of bifidobacteria is of increasing interest; however, mechanisms of transcriptional control of carbohydrate metabolism are poorly understood in these species. We used a comparative genomics approach to reconstruct carbohydrate utilization pathways and transcriptional regulons in 10 Bifidobacterium genomes. Analysis of regulatory gene regions revealed candidate DNA motifs and reconstructed regulons for 268 transcription factors from the LacI, ROK, DeoR, AraC, GntR, and TetR families that form 64 orthologous groups of regulators. Most of the reconstructed regulons are local and control specific catabolic pathways for host- and diet-derived glycans and monosaccharides. Mosaic distributions of many of these local regulators across Bifidobacterium species correlate with distribution of corresponding catabolic pathways. In contrast, the maltose, galactose, sucrose, and fructose regulons, as well as a novel global LacI-family regulator that is predicted to control the central carbohydrate metabolism and arabinose catabolism genes, are universally present in all 10 studied bifidobacteria. A novel group of TetR-family regulators presumably controls the glucoside and galactoside utilization pathways. Paralogs of the ribose repressor RbsR control the pyrimidine nucleoside utilization genes. Multiple paralogs of the maltose regulator MalR co-regulate large sets of genes involved in maltodextrin utilization. The inferred metabolic regulons provide new insights on diverse carbohydrate utilization networks in bifidobacteria that can be employed in metabolic modeling, phenotype prediction and the rational development of novel prebiotics. PMID:26903998

  5. Estrogen receptors regulate innate immune cells and signaling pathways.

    PubMed

    Kovats, Susan

    2015-04-01

    Humans show strong sex differences in immunity to infection and autoimmunity, suggesting sex hormones modulate immune responses. Indeed, receptors for estrogens (ERs) regulate cells and pathways in the innate and adaptive immune system, as well as immune cell development. ERs are ligand-dependent transcription factors that mediate long-range chromatin interactions and form complexes at gene regulatory elements, thus promoting epigenetic changes and transcription. ERs also participate in membrane-initiated steroid signaling to generate rapid responses. Estradiol and ER activity show profound dose- and context-dependent effects on innate immune signaling pathways and myeloid cell development. While estradiol most often promotes the production of type I interferon, innate pathways leading to pro-inflammatory cytokine production may be enhanced or dampened by ER activity. Regulation of innate immune cells and signaling by ERs may contribute to the reported sex differences in innate immune pathways. Here we review the recent literature and highlight several molecular mechanisms by which ERs regulate the development or functional responses of innate immune cells.

  6. Signal Transduction Pathways that Regulate CAB Gene Expression

    SciTech Connect

    Chory, Joanne

    2004-12-31

    The process of chloroplast differentiation, involves the coordinate regulation of many nuclear and chloroplast genes. The cues for the initiation of this developmental program are both extrinsic (e.g., light) and intrinsic (cell-type and plastid signals). During this project period, we utilized a molecular genetic approach to select for Arabidopsis mutants that did not respond properly to environmental light conditions, as well as mutants that were unable to perceive plastid damage. These latter mutants, called gun mutants, define two retrograde signaling pathways that regulate nuclear gene expression in response to chloroplasts. A major finding was to identify a signal from chloroplasts that regulates nuclear gene transcription. This signal is the build-up of Mg-Protoporphyrin IX, a key intermediate of the chlorophyll biosynthetic pathway. The signaling pathways downstream of this signal are currently being studied. Completion of this project has provided an increased understanding of the input signals and retrograde signaling pathways that control nuclear gene expression in response to the functional state of chloroplasts. These studies should ultimately influence our abilities to manipulate plant growth and development, and will aid in the understanding of the developmental control of photosynthesis.

  7. Signal Transduction Pathways that Regulate CAB Gene Expression

    SciTech Connect

    Chory, Joanne

    2006-01-16

    The process of chloroplast differentiation, involves the coordinate regulation of many nuclear and chloroplast genes. The cues for the initiation of this developmental program are both extrinsic (e.g., light) and intrinsic (cell-type and plastid signals). During this project period, we utilized a molecular genetic approach to select for Arabidopsis mutants that did not respond properly to environmental light conditions, as well as mutants that were unable to perceive plastid damage. These latter mutants, called gun mutants, define two retrograde signaling pathways that regulate nuclear gene expression in response to chloroplasts. A major finding was to identify a signal from chloroplasts that regulates nuclear gene transcription. This signal is the build-up of Mg-Protoporphyrin IX, a key intermediate of the chlorophyll biosynthetic pathway. The signaling pathways downstream of this signal are currently being studied. Completion of this project has provided an increased understanding of the input signals and retrograde signaling pathways that control nuclear gene expression in response to the functional state of chloroplasts. These studies should ultimately influence our abilities to manipulate plant growth and development, and will aid in the understanding of the developmental control of photosynthesis.

  8. YAP regulates neuronal differentiation through Sonic hedgehog signaling pathway

    SciTech Connect

    Lin, Yi-Ting; Ding, Jing-Ya; Li, Ming-Yang; Yeh, Tien-Shun; Wang, Tsu-Wei; Yu, Jenn-Yah

    2012-09-10

    Tight regulation of cell numbers by controlling cell proliferation and apoptosis is important during development. Recently, the Hippo pathway has been shown to regulate tissue growth and organ size in Drosophila. In mammalian cells, it also affects cell proliferation and differentiation in various tissues, including the nervous system. Interplay of several signaling cascades, such as Notch, Wnt, and Sonic Hedgehog (Shh) pathways, control cell proliferation during neuronal differentiation. However, it remains unclear whether the Hippo pathway coordinates with other signaling cascades in regulating neuronal differentiation. Here, we used P19 cells, a mouse embryonic carcinoma cell line, as a model to study roles of YAP, a core component of the Hippo pathway, in neuronal differentiation. P19 cells can be induced to differentiate into neurons by expressing a neural bHLH transcription factor gene Ascl1. Our results showed that YAP promoted cell proliferation and inhibited neuronal differentiation. Expression of Yap activated Shh but not Wnt or Notch signaling activity during neuronal differentiation. Furthermore, expression of Yap increased the expression of Patched homolog 1 (Ptch1), a downstream target of the Shh signaling. Knockdown of Gli2, a transcription factor of the Shh pathway, promoted neuronal differentiation even when Yap was over-expressed. We further demonstrated that over-expression of Yap inhibited neuronal differentiation in primary mouse cortical progenitors and Gli2 knockdown rescued the differentiation defect in Yap over-expressing cells. In conclusion, our study reveals that Shh signaling acts downstream of YAP in regulating neuronal differentiation. -- Highlights: Black-Right-Pointing-Pointer YAP promotes cell proliferation and inhibits neuronal differentiation in P19 cells. Black-Right-Pointing-Pointer YAP promotes Sonic hedgehog signaling activity during neuronal differentiation. Black-Right-Pointing-Pointer Knockdown of Gli2 rescues the Yap

  9. Protein Kinases of the Hippo Pathway: Regulation and Substrates

    PubMed Central

    Avruch, Joseph; Zhou, Dawang; Fitamant, Julien; Bardeesy, Nabeel; Mou, Fan; Barrufet, Laura Regué

    2012-01-01

    The “Hippo” signaling pathway has emerged as a major regulator of cell proliferation and survival in metazoans. The pathway, as delineated by genetic and biochemical studies in Drosophila, consists of a kinase cascade regulated by cell-cell contact and cell polarity that inhibits the transcriptional coactivator Yorkie and its proliferative, anti-differentiation, antiapoptotic transcriptional program. The core pathway components are the GC kinase Hippo, which phosphorylates the noncatalytic polypeptide Mats/Mob1 and, with the assistance of the scaffold protein Salvador, phosphorylates the ndr-family kinase Lats. In turn phospho-Lats, after binding to phospho-Mats, autoactivates and phosphorylates Yorkie, resulting in its nuclear exit. Hippo also uses the scaffold protein Furry and a different Mob protein to control another ndr-like kinase, the morphogenetic regulator Tricornered. Architecturally homologous kinase cascades consisting of a GC kinase, a Mob protein, a scaffolding polypeptide and an ndr-like kinase are well described in yeast; in S. cerevisiae e.g., the MEN pathway promotes mitotic exit whereas the RAM network, using a different GC kinase, Mob protein, scaffold and ndr-like kinase, regulates cell polarity and morphogenesis. In mammals, the Hippo orthologues Mst1 and Mst2 utilize the Salvador ortholog WW45/Sav1 and other scaffolds to regulate the kinases Lats1/Lats2 and ndr1/ndr2. As in Drosophila, murine Mst1/Mst2, in a redundant manner, negatively regulate the Yorkie ortholog YAP in the epithelial cells of the liver and gut; loss of both Mst1 and Mst2 results in hyperproliferation and tumorigenesis that can be largely negated by reduction or elimination of YAP. Despite this conservation, considerable diversification in pathway composition and regulation is already evident; in skin e.g., YAP phosphorylation is independent of Mst1Mst2 and Lats1Lats2. Moreover, in lymphoid cells, Mst1/Mst2, under the control of the Rap1 GTPase and independent of YAP

  10. The deubiquitinating enzyme USP1 regulates the Fanconi anemia pathway.

    PubMed

    Nijman, Sebastian M B; Huang, Tony T; Dirac, Annette M G; Brummelkamp, Thijn R; Kerkhoven, Ron M; D'Andrea, Alan D; Bernards, René

    2005-02-04

    Protein ubiquitination and deubiquitination are dynamic processes implicated in the regulation of numerous cellular pathways. Monoubiquitination of the Fanconi anemia (FA) protein FANCD2 appears to be critical in the repair of DNA damage because many of the proteins that are mutated in FA are required for FANCD2 ubiquitination. By screening a gene family RNAi library, we identify the deubiquitinating enzyme USP1 as a novel component of the Fanconi anemia pathway. Inhibition of USP1 leads to hyperaccumulation of monoubiquitinated FANCD2. Furthermore, USP1 physically associates with FANCD2, and the proteins colocalize in chromatin after DNA damage. Finally, analysis of crosslinker-induced chromosomal aberrations in USP1 knockdown cells suggests a role in DNA repair. We propose that USP1 deubiquitinates FANCD2 when cells exit S phase or recommence cycling after a DNA damage insult and may play a critical role in the FA pathway by recycling FANCD2.

  11. The mevalonate pathway regulates primitive streak formation via protein farnesylation

    PubMed Central

    Okamoto-Uchida, Yoshimi; Yu, Ruoxing; Miyamura, Norio; Arima, Norie; Ishigami-Yuasa, Mari; Kagechika, Hiroyuki; Yoshida, Suguru; Hosoya, Takamitsu; Nawa, Makiko; Kasama, Takeshi; Asaoka, Yoichi; Alois, Reiner Wimmer; Elling, Ulrich; Penninger, Josef M.; Nishina, Sachiko; Azuma, Noriyuki; Nishina, Hiroshi

    2016-01-01

    The primitive streak in peri-implantation embryos forms the mesoderm and endoderm and controls cell differentiation. The metabolic cues regulating primitive streak formation remain largely unknown. Here we utilised a mouse embryonic stem (ES) cell differentiation system and a library of well-characterised drugs to identify these metabolic factors. We found that statins, which inhibit the mevalonate metabolic pathway, suppressed primitive streak formation in vitro and in vivo. Using metabolomics and pharmacologic approaches we identified the downstream signalling pathway of mevalonate and revealed that primitive streak formation requires protein farnesylation but not cholesterol synthesis. A tagging-via-substrate approach revealed that nuclear lamin B1 and small G proteins were farnesylated in embryoid bodies and important for primitive streak gene expression. In conclusion, protein farnesylation driven by the mevalonate pathway is a metabolic cue essential for primitive streak formation. PMID:27883036

  12. BPIFB6 Regulates Secretory Pathway Trafficking and Enterovirus Replication

    PubMed Central

    Morosky, Stefanie; Lennemann, Nicholas J.

    2016-01-01

    ABSTRACT Bactericidal/permeability-increasing protein (BPI) fold-containing family B, member 3 (BPIFB3) is an endoplasmic reticulum (ER)-localized host factor that negatively regulates coxsackievirus B (CVB) replication through its control of the autophagic pathway. Here, we show that another member of the BPIFB family, BPIFB6, functions as a positive regulator of CVB, and other enterovirus, replication by controlling secretory pathway trafficking and Golgi complex morphology. We show that similar to BPIFB3, BPIFB6 localizes exclusively to the ER, where it associates with other members of the BPIFB family. However, in contrast to our findings that RNA interference (RNAi)-mediated silencing of BPIFB3 greatly enhances CVB replication, we show that silencing of BPIFB6 expression dramatically suppresses enterovirus replication in a pan-viral manner. Mechanistically, we show that loss of BPIFB6 expression induces pronounced alterations in retrograde and anterograde trafficking, which correlate with dramatic fragmentation of the Golgi complex. Taken together, these data implicate BPIFB6 as a key regulator of secretory pathway trafficking and viral replication and suggest that members of the BPIFB family participate in diverse host cell functions to regulate virus infections. IMPORTANCE Enterovirus infections are associated with a number of severe pathologies, such as aseptic meningitis, dilated cardiomyopathy, type I diabetes, paralysis, and even death. These viruses, which include coxsackievirus B (CVB), poliovirus (PV), and enterovirus 71 (EV71), co-opt the host cell secretory pathway, which controls the transport of proteins from the endoplasmic reticulum to the Golgi complex, to facilitate their replication. Here we report on the identification of a novel regulator of the secretory pathway, bactericidal/permeability-increasing protein (BPI) fold-containing family B, member 6 (BPIFB6), whose expression is required for enterovirus replication. We show that loss of

  13. A divergent canonical WNT-signaling pathway regulates microtubule dynamics

    PubMed Central

    Ciani, Lorenza; Krylova, Olga; Smalley, Matthew J.; Dale, Trevor C.; Salinas, Patricia C.

    2004-01-01

    Dishevelled (DVL) is associated with axonal microtubules and regulates microtubule stability through the inhibition of the serine/threonine kinase, glycogen synthase kinase 3β (GSK-3β). In the canonical WNT pathway, the negative regulator Axin forms a complex with β-catenin and GSK-3β, resulting in β-catenin degradation. Inhibition of GSK-3β by DVL increases β-catenin stability and TCF transcriptional activation. Here, we show that Axin associates with microtubules and unexpectedly stabilizes microtubules through DVL. In turn, DVL stabilizes microtubules by inhibiting GSK-3β through a transcription- and β-catenin–independent pathway. More importantly, axonal microtubules are stabilized after DVL localizes to axons. Increased microtubule stability is correlated with a decrease in GSK-3β–mediated phosphorylation of MAP-1B. We propose a model in which Axin, through DVL, stabilizes microtubules by inhibiting a pool of GSK-3β, resulting in local changes in the phosphorylation of cellular targets. Our data indicate a bifurcation in the so-called canonical WNT-signaling pathway to regulate microtubule stability. PMID:14734535

  14. p53 regulates the mevalonate pathway in human glioblastoma multiforme

    PubMed Central

    Laezza, C; D'Alessandro, A; Di Croce, L; Picardi, P; Ciaglia, E; Pisanti, S; Malfitano, A M; Comegna, M; Faraonio, R; Gazzerro, P; Bifulco, M

    2015-01-01

    The mevalonate (MVA) pathway is an important metabolic pathway implicated in multiple aspects of tumorigenesis. In this study, we provided evidence that p53 induces the expression of a group of enzymes of the MVA pathway including 3′-hydroxy-3′-methylglutaryl-coenzyme A reductase, MVA kinase, farnesyl diphosphate synthase and farnesyl diphosphate farnesyl transferase 1, in the human glioblastoma multiforme cell line, U343 cells, and in normal human astrocytes, NHAs. Genetic and pharmacologic perturbation of p53 directly influences the expression of these genes. Furthermore, p53 is recruited to the gene promoters in designated p53-responsive elements, thereby increasing their transcription. Such effect was abolished by site-directed mutagenesis in the p53-responsive element of promoter of the genes. These findings highlight another aspect of p53 functions unrelated to tumor suppression and suggest p53 as a novel regulator of the MVA pathway providing insight into the role of this pathway in cancer progression. PMID:26469958

  15. New insights into transduction pathways that regulate boar sperm function.

    PubMed

    Hurtado de Llera, A; Martin-Hidalgo, D; Gil, M C; Garcia-Marin, L J; Bragado, M J

    2016-01-01

    Detailed molecular mechanisms mediating signal transduction cascades that regulate boar sperm function involving Ser/Thr and tyrosine phosphorylation of proteins have been reviewed previously. Therefore, this review will focus in those kinase pathways identified recently (<10 years) in boar spermatozoa that regulate different functional spermatozoa processes. AMP-activated protein kinase (AMPK) is a cell energy sensor kinase that was first identified in mammalian spermatozoa in 2012, and since then it has emerged as an essential regulator of boar sperm function. Signaling pathways leading to AMPK activation in boar sperm are highlighted in this review (PKA, CaMKKα/β, and PKC as well as Ca(2+) and cAMP messengers as upstream regulators). Interestingly, stimuli considered as cell stress (hyperosmotic stress, inhibition of mitochondrial activity, absence of intracellular Ca(2+)) markedly activate AMPK in boar spermatozoa. Moreover, AMPK plays a remarkable and necessary regulatory role in mammalian sperm function, controlling essential boar sperm functional processes such as motility, viability, mitochondrial membrane potential, organization and fluidity of plasma membrane, and outer acrosome membrane integrity. These mentioned processes are all required under fluctuating environment of spermatozoa when transiting through the female reproductive tract to achieve fertilization. An applied role of AMPK in artificial insemination techniques is also suggested as during boar seminal doses preservation at 17 °C, physiological levels of AMPK activity markedly increase (maximum on Day 7) and result essential to maintain the aforementioned fundamental sperm processes. Moreover, regulation of sperm function exerted by the glycogen synthase kinase 3 and Src family kinase pathways is summarized.

  16. Targeting TNF: a therapeutic strategy for Alzheimer's disease.

    PubMed

    Cheng, Xin; Shen, Yong; Li, Rena

    2014-11-01

    Tumor necrosis factor (TNF), a ligand cytokine, is involved in systemic inflammation. Apart from the well-known pharmacological effects of TNF inhibitors on autoimmune disorders, interest in the effects of TNF in neurodegenerative disorders such as Alzheimer disease (AD) is increasing. TNF and its type 1 receptor (TNFRI) are not only involved in AD-related brain neuroinflammation, but also contribute to amyloidogenesis via β-secretase regulation, suggesting TNF as a promising candidate for future AD therapy. Although the potential adverse effects of TNF-based AD therapies have been of concerns, here we summarize recent discoveries relating to TNF and TNFRI-mediated signal transduction as potential therapeutic targets in AD pathology and clinical investigations.

  17. [Central regulation of body temperature by RANKL/RANK pathway].

    PubMed

    Hanada, Reiko; Penninger, Josef M

    2011-08-01

    Receptor-activator of NF-κB ligand (RANKL) and its specific receptor RANK are key regulators of bone remodeling, lymph node formation, establishment of the thymic microenviroment, mammary gland development during pregnancy, bone metastasis in cancer and sex-hormone, progestin, -driven breast cancer. RANKL and RANK are also expressed in the central nervous systems (CNS) especially existed in the main region of thermoregulation. Central RANKL injection to the rodents induces fever via PGE(2)/EP3R pathway. This pathway is related with inflammation related fever. On the other hand, female mice with RANK gene deletion in neuron and astrocytes show increased their basal body temperature at the dark phase, which suggests RANKL/RANK system also regulates physiological thremoregulation in female. Not only in rodents but also in human, two children with a homozygous RANK mutation exhibit an abrogated fever response in pneumonia compare with the age-matched children with pneumonia. Thus, the central RANKL/RANK pathway has an important role for thermoregulation.

  18. Copper as a key regulator of cell signalling pathways.

    PubMed

    Grubman, Alexandra; White, Anthony R

    2014-05-22

    Copper is an essential element in many biological processes. The critical functions associated with copper have resulted from evolutionary harnessing of its potent redox activity. This same property also places copper in a unique role as a key modulator of cell signal transduction pathways. These pathways are the complex sequence of molecular interactions that drive all cellular mechanisms and are often associated with the interplay of key enzymes including kinases and phosphatases but also including intracellular changes in pools of smaller molecules. A growing body of evidence is beginning to delineate the how, when and where of copper-mediated control over cell signal transduction. This has been driven by research demonstrating critical changes to copper homeostasis in many disorders including cancer and neurodegeneration and therapeutic potential through control of disease-associated cell signalling changes by modulation of copper-protein interactions. This timely review brings together for the first time the diverse actions of copper as a key regulator of cell signalling pathways and discusses the potential strategies for controlling disease-associated signalling processes using copper modulators. It is hoped that this review will provide a valuable insight into copper as a key signal regulator and stimulate further research to promote our understanding of copper in disease and therapy.

  19. Dicer-dependent pathways regulate chondrocyte proliferation and differentiation.

    PubMed

    Kobayashi, Tatsuya; Lu, Jun; Cobb, Bradley S; Rodda, Stephen J; McMahon, Andrew P; Schipani, Ernestina; Merkenschlager, Matthias; Kronenberg, Henry M

    2008-02-12

    Small noncoding RNAs, microRNAs (miRNAs), bind to messenger RNAs through base pairing to suppress gene expression. Despite accumulating evidence that miRNAs play critical roles in various biological processes across diverse organisms, their roles in mammalian skeletal development have not been demonstrated. Here, we show that Dicer, an essential component for biogenesis of miRNAs, is essential for normal skeletal development. Dicer-null growth plates show a progressive reduction in the proliferating pool of chondrocytes, leading to severe skeletal growth defects and premature death of mice. The reduction of proliferating chondrocytes in Dicer-null growth plates is caused by two distinct mechanisms: decreased chondrocyte proliferation and accelerated differentiation into postmitotic hypertrophic chondrocytes. These defects appear to be caused by mechanisms downstream or independent of the Ihh-PTHrP signaling pathway, a pivotal signaling system that regulates chondrocyte proliferation and differentiation. Microarray analysis of Dicer-null chondrocytes showed limited expression changes in miRNA-target genes, suggesting that, in the majority of cases, chondrocytic miRNAs do not directly regulate target RNA abundance. Our results demonstrate the critical role of the Dicer-dependent pathway in the regulation of chondrocyte proliferation and differentiation during skeletal development.

  20. Autopalmitoylation of TEAD Proteins Regulates Transcriptional Output of Hippo Pathway

    PubMed Central

    Chan, PuiYee; Han, Xiao; Zheng, Baohui; DeRan, Michael; Yu, Jianzhong; Jarugumilli, Gopala K.; Deng, Hua; Pan, Duojia; Luo, Xuelian; Wu, Xu

    2016-01-01

    TEA domain (TEAD) transcription factors bind to the co-activator YAP/TAZ, and regulate the transcriptional output of Hippo pathway, playing critical roles in organ size control and tumorigenesis. Protein S-palmitoylation attaches fatty acid (palmitate) to cysteine residues, and regulates protein trafficking, membrane localization and signaling activities. Using activity-based chemical probes, we discovered that human TEADs possess intrinsic palmitoylating enzyme-like activities, and undergo autopalmitoylation at evolutionarily conserved cysteine residues under physiological conditions. We determined the crystal structures of lipid-bound TEADs, and found that the lipid chain of palmitate inserts into a conserved deep hydrophobic pocket. Strikingly, palmitoylation is required for TEAD’s binding to YAP/TAZ, but dispensable for the binding to Vgll4 tumor suppressor. In addition, palmitoylation does not alter TEAD’s localization. Moreover, TEAD palmitoylation-deficient mutants impaired TAZ-mediated muscle differentiation in vitro, and Yorkie-mediated tissue overgrowth in Drosophila in vivo. Our study directly linked autopalmitoylation to the transcriptional regulation of Hippo pathway. PMID:26900866

  1. Epigenetic high regulation of ATAD2 regulates the Hh pathway in human hepatocellular carcinoma.

    PubMed

    Wu, Gang; Lu, Xiaojun; Wang, Yawei; He, Hui; Meng, Xiangyu; Xia, Shuguan; Zhen, Kunming; Liu, Yongfeng

    2014-07-01

    ATAD2 is associated with many cellular progresses such as cell growth, differentiation and apoptosis. Some studies suggest ATAD2 is highly expressed in cancer cells. In our previous studies, we found that ATAD2 is highly expressed in HCC tissues, compared with adjacent normal tissues, and patients with high expression of ATAD2 had a poorer prognosis. Moreover, we found mir-372 can regulate the expression of ATAD2 in HCC cell lines. We also detected a relationship between the mRNA expression of ATAD2 and Ptch1 by gene microarray. Here, we completed the function studies of ATAD2 in vivo and in vitro, and tested whether ATAD2 could regulate the Hh pathway. ATAD2 and Hh pathway protein expressions in 80 HCC specimens were examined by immunohistochemistry (IHC). The mRNA expression of ATAD2 and Hh pathway members in paired-HCC tissues and cell lines were, respectively, analyzed using quantitative PCR. ATAD2‑RNAi was transduced into HCCLM3 and Huh7 cells, using a lentiviral vector. The effect of ATAD2 in HCC cell lines on cell cycle and apoptosis were evaluated by flow cytometry. Tumorigenicity experiments in nude mice were performed to test the function of ATAD2 in vivo. Pharmacological regulation of Hh signaling was performed to test the relation between the ATAD2 and Hh pathways and C-myc. We found that ATAD2 and Ptch1 were both highly expressed in HCC tissues, compared with paired normal hepatic tissues. In addition, we found that ATAD2 could affect the expression of the Hh pathway by PCR and western blot anaysis in HCC cell lines, by observing the outcome before and after transfection. We speculate that ATAD2 cooperates with the MYC gene to regulate the expression of SMO and Gli, activating the Hh pathway and inducing an active feedback of the Hh pathway.

  2. Regulation of ribosomal DNA amplification by the TOR pathway

    PubMed Central

    Jack, Carmen V.; Cruz, Cristina; Hull, Ryan M.; Keller, Markus A.; Ralser, Markus; Houseley, Jonathan

    2015-01-01

    Repeated regions are widespread in eukaryotic genomes, and key functional elements such as the ribosomal DNA tend to be formed of high copy repeated sequences organized in tandem arrays. In general, high copy repeats are remarkably stable, but a number of organisms display rapid ribosomal DNA amplification at specific times or under specific conditions. Here we demonstrate that target of rapamycin (TOR) signaling stimulates ribosomal DNA amplification in budding yeast, linking external nutrient availability to ribosomal DNA copy number. We show that ribosomal DNA amplification is regulated by three histone deacetylases: Sir2, Hst3, and Hst4. These enzymes control homologous recombination-dependent and nonhomologous recombination-dependent amplification pathways that act in concert to mediate rapid, directional ribosomal DNA copy number change. Amplification is completely repressed by rapamycin, an inhibitor of the nutrient-responsive TOR pathway; this effect is separable from growth rate and is mediated directly through Sir2, Hst3, and Hst4. Caloric restriction is known to up-regulate expression of nicotinamidase Pnc1, an enzyme that enhances Sir2, Hst3, and Hst4 activity. In contrast, normal glucose concentrations stretch the ribosome synthesis capacity of cells with low ribosomal DNA copy number, and we find that these cells show a previously unrecognized transcriptional response to caloric excess by reducing PNC1 expression. PNC1 down-regulation forms a key element in the control of ribosomal DNA amplification as overexpression of PNC1 substantially reduces ribosomal DNA amplification rate. Our results reveal how a signaling pathway can orchestrate specific genome changes and demonstrate that the copy number of repetitive DNA can be altered to suit environmental conditions. PMID:26195783

  3. Metabolic control of signalling pathways and metabolic auto-regulation.

    PubMed

    Lorendeau, Doriane; Christen, Stefan; Rinaldi, Gianmarco; Fendt, Sarah-Maria

    2015-08-01

    Metabolic alterations have emerged as an important hallmark in the development of various diseases. Thus, understanding the complex interplay of metabolism with other cellular processes such as cell signalling is critical to rationally control and modulate cellular physiology. Here, we review in the context of mammalian target of rapamycin, AMP-activated protein kinase and p53, the orchestrated interplay between metabolism and cellular signalling as well as transcriptional regulation. Moreover, we discuss recent discoveries in auto-regulation of metabolism (i.e. how metabolic parameters such as metabolite levels activate or inhibit enzymes and thus metabolic pathways). Finally, we review functional consequences of post-translational modification on metabolic enzyme abundance and/or activities.

  4. Bioinformatic dissecting of TP53 regulation pathway underlying butyrate-induced histone modification in epigenetic regulation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Butyrate affects cell proliferation, differentiation and motility. Butyrate inhibits histone deacetylase (HDAC) activities and induces cell cycle arrest and apoptosis. TP53 is one of the most active upstream regulators discovered by IPA in our RNA sequencing data set. The TP53 signaling pathway pl...

  5. Multiple pathways regulate minisatellite stability during stationary phase in yeast.

    PubMed

    Kelly, Maire K; Brosnan, Laura; Jauert, Peter A; Dunham, Maitreya J; Kirkpatrick, David T

    2012-10-01

    Alterations in minisatellite DNA repeat tracts in humans have been correlated with a number of serious disorders, including cancer. Despite their importance for human health, the genetic factors that influence minisatellite stability are not well understood. Previously, we identified mutations in the Saccharomyces cerevisiae zinc homeostasis genes ZRT1 and ZAP1 that significantly increase the frequency of minisatellite alteration specifically during stationary phase. In this work, we identified mutants of END3, PKC1, and RAD27 that increase minisatellite instability during stationary phase. Genetic analysis reveals that these genes, along with ZRT1 and ZAP1, comprise multiple pathways regulating minisatellite stability during stationary phase. Minisatellite alterations generated by perturbation of any of these pathways occur via homologous recombination. We present evidence that suggests formation of ssDNA or ssDNA breaks may play a primary role in stationary phase instability. Finally, we examined the roles of these pathways in the stability of a human minisatellite tract associated with the HRAS1 oncogene and found that loss of RAD27, but not END3 or PKC1, destabilizes the HRAS1 minisatellite in stationary phase yeast. This result indicates that the genetic control of stationary phase minisatellite stability is dependent on the sequence composition of the minisatellite itself.

  6. Strigolactone regulates shoot development through a core signalling pathway

    PubMed Central

    Müller, Dörte

    2016-01-01

    ABSTRACT Strigolactones are a recently identified class of hormone that regulate multiple aspects of plant development. The DWARF14 (D14) α/β fold protein has been identified as a strigolactone receptor, which can act through the SCFMAX2 ubiquitin ligase, but the universality of this mechanism is not clear. Multiple proteins have been suggested as targets for strigolactone signalling, including both direct proteolytic targets of SCFMAX2, and downstream targets. However, the relevance and importance of these proteins to strigolactone signalling in many cases has not been fully established. Here we assess the contribution of these targets to strigolactone signalling in adult shoot developmental responses. We find that all examined strigolactone responses are regulated by SCFMAX2 and D14, and not by other D14-like proteins. We further show that all examined strigolactone responses likely depend on degradation of SMXL proteins in the SMXL6 clade, and not on the other proposed proteolytic targets BES1 or DELLAs. Taken together, our results suggest that in the adult shoot, the dominant mode of strigolactone signalling is D14-initiated, MAX2-mediated degradation of SMXL6-related proteins. We confirm that the BRANCHED1 transcription factor and the PIN-FORMED1 auxin efflux carrier are plausible downstream targets of this pathway in the regulation of shoot branching, and show that BRC1 likely acts in parallel to PIN1. PMID:27793831

  7. The Spectrin cytoskeleton regulates the Hippo signalling pathway

    PubMed Central

    Fletcher, Georgina C; Elbediwy, Ahmed; Khanal, Ichha; Ribeiro, Paulo S; Tapon, Nic; Thompson, Barry J

    2015-01-01

    The Spectrin cytoskeleton is known to be polarised in epithelial cells, yet its role remains poorly understood. Here, we show that the Spectrin cytoskeleton controls Hippo signalling. In the developing Drosophila wing and eye, loss of apical Spectrins (alpha/beta-heavy dimers) produces tissue overgrowth and mis-regulation of Hippo target genes, similar to loss of Crumbs (Crb) or the FERM-domain protein Expanded (Ex). Apical beta-heavy Spectrin binds to Ex and co-localises with it at the apical membrane to antagonise Yki activity. Interestingly, in both the ovarian follicular epithelium and intestinal epithelium of Drosophila, apical Spectrins and Crb are dispensable for repression of Yki, while basolateral Spectrins (alpha/beta dimers) are essential. Finally, the Spectrin cytoskeleton is required to regulate the localisation of the Hippo pathway effector YAP in response to cell density human epithelial cells. Our findings identify both apical and basolateral Spectrins as regulators of Hippo signalling and suggest Spectrins as potential mechanosensors. PMID:25712476

  8. SIRT1 regulates MAPK pathways in vitiligo skin: insight into the molecular pathways of cell survival

    PubMed Central

    Becatti, Matteo; Fiorillo, Claudia; Barygina, Victoria; Cecchi, Cristina; Lotti, Torello; Prignano, Francesca; Silvestro, Agrippino; Nassi, Paolo; Taddei, Niccolò

    2014-01-01

    Vitiligo is an acquired and progressive hypomelanotic disease that manifests as circumscribed depigmented patches on the skin. The aetiology of vitiligo remains unclear, but recent experimental data underline the interactions between melanocytes and other typical skin cells, particularly keratinocytes. Our previous results indicate that keratinocytes from perilesional skin show the features of damaged cells. Sirtuins (silent mating type information regulation 2 homolog) 1, well-known modulators of lifespan in many species, have a role in gene repression, metabolic control, apoptosis and cell survival, DNA repair, development, inflammation, neuroprotection and healthy ageing. In the literature there is no evidence for SIRT1 signalling in vitiligo and its possible involvement in disease progression. Here, biopsies were taken from the perilesional skin of 16 patients suffering from non-segmental vitiligo and SIRT1 signalling was investigated in these cells. For the first time, a new SIRT1/Akt, also known as Protein Kinase B (PKB)/mitogen-activated protein kinase (MAPK) signalling has been revealed in vitiligo. SIRT1 regulates MAPK pathway via Akt-apoptosis signal-regulating kinase-1 and down-regulates pro-apoptotic molecules, leading to decreased oxidative stress and apoptotic cell death in perilesional vitiligo keratinocytes. We therefore propose SIRT1 activation as a novel way of protecting perilesional vitiligo keratinocytes from damage. PMID:24410795

  9. Adverse outcome pathways: From research to regulation scientific workshop report.

    PubMed

    Kleinstreuer, Nicole C; Sullivan, Kristie; Allen, David; Edwards, Stephen; Mendrick, Donna L; Embry, Michelle; Matheson, Joanna; Rowlands, J Craig; Munn, Sharon; Maull, Elizabeth; Casey, Warren

    2016-04-01

    An adverse outcome pathway (AOP) helps to organize existing knowledge on chemical mode of action, starting with a molecular initiating event such as receptor binding, continuing through key events, and ending with an adverse outcome such as reproductive impairment. AOPs can help identify knowledge gaps where more research is needed to understand the underlying mechanisms, aid in chemical hazard characterization, and guide the development of new testing approaches that use fewer or no animals. A September 2014 workshop in Bethesda, Maryland considered how the AOP concept could improve regulatory assessments of chemical toxicity. Scientists from 21 countries, representing industry, academia, regulatory agencies, and special interest groups, attended the workshop, titled Adverse Outcome Pathways: From Research to Regulation. Workshop plenary presentations were followed by breakout sessions that considered regulatory acceptance of AOPs and AOP-based tools, criteria for building confidence in an AOP for regulatory use, and requirements to build quantitative AOPs and AOP networks. Discussions during the closing session emphasized a need to increase transparent and inclusive collaboration, especially with disciplines outside of toxicology. Additionally, to increase impact, working groups should be established to systematically prioritize and develop AOPs. Multiple collaborative projects and follow-up activities resulted from the workshop.

  10. TOR and RAS pathways regulate desiccation tolerance in Saccharomyces cerevisiae

    PubMed Central

    Welch, Aaron Z.; Gibney, Patrick A.; Botstein, David; Koshland, Douglas E.

    2013-01-01

    Tolerance to desiccation in cultures of Saccharomyces cerevisiae is inducible; only one in a million cells from an exponential culture survive desiccation compared with one in five cells in stationary phase. Here we exploit the desiccation sensitivity of exponentially dividing cells to understand the stresses imposed by desiccation and their stress response pathways. We found that induction of desiccation tolerance is cell autonomous and that there is an inverse correlation between desiccation tolerance and growth rate in glucose-, ammonia-, or phosphate-limited continuous cultures. A transient heat shock induces a 5000–fold increase in desiccation tolerance, whereas hyper-ionic, -reductive, -oxidative, or -osmotic stress induced much less. Furthermore, we provide evidence that the Sch9p-regulated branch of the TOR and Ras-cAMP pathway inhibits desiccation tolerance by inhibiting the stress response transcription factors Gis1p, Msn2p, and Msn4p and by activating Sfp1p, a ribosome biogenesis transcription factor. Among 41 mutants defective in ribosome biogenesis, a subset defective in 60S showed a dramatic increase in desiccation tolerance independent of growth rate. We suggest that reduction of a specific intermediate in 60S biogenesis, resulting from conditions such as heat shock and nutrient deprivation, increases desiccation tolerance. PMID:23171550

  11. Pathways regulating modality-specific axonal regeneration in peripheral nerve.

    PubMed

    Wood, Matthew D; Mackinnon, Susan E

    2015-03-01

    Following peripheral nerve injury, the distal nerve is primed for regenerating axons by generating a permissive environment replete with glial cells, cytokines, and neurotrophic factors to encourage axonal growth. However, increasing evidence demonstrates that regenerating axons within peripheral nerves still encounter axonal-growth inhibitors, such as chondroitin sulfate proteoglycans. Given the generally poor clinical outcomes following peripheral nerve injury and reconstruction, the use of pharmacological therapies to augment axonal regeneration and overcome inhibitory signals has gained considerable interest. Joshi et al. (2014) have provided evidence for preferential or modality-specific (motor versus sensory) axonal growth and regeneration due to inhibitory signaling from Rho-associated kinase (ROCK) pathway regulation. By providing inhibition to the ROCK signaling pathway through Y-27632, they demonstrate that motor neurons regenerating their axons are impacted to a greater extent compared to sensory neurons. In light of this evidence, we briefly review the literature regarding modality-specific axonal regeneration to provide context to their findings. We also describe potential and novel barriers, such as senescent Schwann cells, which provide additional axonal-growth inhibitory factors for future consideration following peripheral nerve injury.

  12. DUB3 Deubiquitylating Enzymes Regulate Hippo Pathway Activity by Regulating the Stability of ITCH, LATS and AMOT Proteins.

    PubMed

    Nguyen, Hung Thanh; Kugler, Jan-Michael; Cohen, Stephen M

    2017-01-01

    The YAP and TAZ transcriptional coactivators promote oncogenic transformation. Elevated YAP/TAZ activity has been documented in human tumors. YAP and TAZ are negatively regulated by the Hippo tumor suppressor pathway. The activity and stability of several Hippo pathway components, including YAP/TAZ, is regulated by ubiquitin mediated protein turnover and several ubiquitin ligase complexes have been implicated in human cancer. However, little is known about the deubiquitylating enzymes that counteract these ubiquitin ligases in regulation of the Hippo pathway. Here we identify the DUB3 family deubiquitylating enzymes as regulators of Hippo pathway activity. We provide evidence that DUB3 proteins regulate YAP/TAZ activity by controlling the stability of the E3 ligase ITCH, the LATS kinases and the AMOT family proteins. As a novel Hippo pathway regulator, DUB3 has the potential to act a tumor suppressor by limiting YAP activity.

  13. DUB3 Deubiquitylating Enzymes Regulate Hippo Pathway Activity by Regulating the Stability of ITCH, LATS and AMOT Proteins

    PubMed Central

    2017-01-01

    The YAP and TAZ transcriptional coactivators promote oncogenic transformation. Elevated YAP/TAZ activity has been documented in human tumors. YAP and TAZ are negatively regulated by the Hippo tumor suppressor pathway. The activity and stability of several Hippo pathway components, including YAP/TAZ, is regulated by ubiquitin mediated protein turnover and several ubiquitin ligase complexes have been implicated in human cancer. However, little is known about the deubiquitylating enzymes that counteract these ubiquitin ligases in regulation of the Hippo pathway. Here we identify the DUB3 family deubiquitylating enzymes as regulators of Hippo pathway activity. We provide evidence that DUB3 proteins regulate YAP/TAZ activity by controlling the stability of the E3 ligase ITCH, the LATS kinases and the AMOT family proteins. As a novel Hippo pathway regulator, DUB3 has the potential to act a tumor suppressor by limiting YAP activity. PMID:28061504

  14. Cross-regulation of signaling pathways: An example of nuclear hormone receptors and the canonical Wnt pathway

    SciTech Connect

    Beildeck, Marcy E.; Gelmann, Edward P.; Byers, Stephen W.

    2010-07-01

    Predicting the potential physiological outcome(s) of any given molecular pathway is complex because of cross-talk with other pathways. This is particularly evident in the case of the nuclear hormone receptor and canonical Wnt pathways, which regulate cell growth and proliferation, differentiation, apoptosis, and metastatic potential in numerous tissues. These pathways are known to intersect at many levels: in the intracellular space, at the membrane, in the cytoplasm, and within the nucleus. The outcomes of these interactions are important in the control of stem cell differentiation and maintenance, feedback loops, and regulating oncogenic potential. The aim of this review is to demonstrate the importance of considering pathway cross-talk when predicting functional outcomes of signaling, using nuclear hormone receptor/canonical Wnt pathway cross-talk as an example.

  15. Notch Signaling Pathway Regulates Progesterone Secretion in Murine Luteal Cells.

    PubMed

    Wang, Jing; Liu, Shuangmei; Peng, Lichao; Dong, Qiming; Bao, Riqiang; Lv, Qiulan; Tang, Min; Hu, Chuan; Li, Gang; Liang, Shangdong; Zhang, Chunping

    2015-10-01

    Notch signaling is an evolutionarily conserved pathway, which involves in various cell life activities. Other studies and our report showed that the Notch signaling plays very important role in follicle development in mammalian ovaries. In luteal cells, Notch ligand, delta-like ligand 4, is involved in normal luteal vasculature. In this study, murine luteal cells were cultured in vitro and treated with Notch signaling inhibitors, L-658,458 and N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycinet-butyl ester (DAPT). We found that L-658,458 and DAPT treatment decrease basal and human chorionic gonadotropin (hCG)-stimulated progesterone secretion. On the contrary, overexpression of intracellular domain of Notch3 increased basal and hCG-stimulated progesterone secretion. Further studies demonstrated that Notch signaling regulated the expression of steroidogenic acute regulatory protein and CYP11A, 2 key enzymes for progesterone synthesis. In conclusion, Notch signaling plays important role in regulating progesterone secretion in murine luteal cells.

  16. Transcriptional regulation of neurodevelopmental and metabolic pathways by NPAS3.

    PubMed

    Sha, L; MacIntyre, L; Machell, J A; Kelly, M P; Porteous, D J; Brandon, N J; Muir, W J; Blackwood, D H; Watson, D G; Clapcote, S J; Pickard, B S

    2012-03-01

    The basic helix-loop-helix PAS (Per, Arnt, Sim) domain transcription factor gene NPAS3 is a replicated genetic risk factor for psychiatric disorders. A knockout (KO) mouse model exhibits behavioral and adult neurogenesis deficits consistent with human illness. To define the location and mechanism of NPAS3 etiopathology, we combined immunofluorescent, transcriptomic and metabonomic approaches. Intense Npas3 immunoreactivity was observed in the hippocampal subgranular zone-the site of adult neurogenesis--but was restricted to maturing, rather than proliferating, neuronal precursor cells. Microarray analysis of a HEK293 cell line over-expressing NPAS3 showed that transcriptional targets varied according to circadian rhythm context and C-terminal deletion. The most highly up-regulated NPAS3 target gene, VGF, encodes secretory peptides with established roles in neurogenesis, depression and schizophrenia. VGF was just one of many NPAS3 target genes also regulated by the SOX family of transcription factors, suggesting an overlap in neurodevelopmental function. The parallel repression of multiple glycolysis genes by NPAS3 reveals a second role in the regulation of glucose metabolism. Comparison of wild-type and Npas3 KO metabolite composition using high-resolution mass spectrometry confirmed these transcriptional findings. KO brain tissue contained significantly altered levels of NAD(+), glycolysis metabolites (such as dihydroxyacetone phosphate and fructose-1,6-bisphosphate), pentose phosphate pathway components and Kreb's cycle intermediates (succinate and α-ketoglutarate). The dual neurodevelopmental and metabolic aspects of NPAS3 activity described here increase our understanding of mental illness etiology, and may provide a mechanism for innate and medication-induced susceptibility to diabetes commonly reported in psychiatric patients.

  17. MicroRNAs: New Regulators of Toll-Like Receptor Signalling Pathways

    PubMed Central

    He, Xiaobing; Jing, Zhizhong; Cheng, Guofeng

    2014-01-01

    Toll-like receptors (TLRs), a critical family of pattern recognition receptors (PRRs), are responsible for the innate immune responses via signalling pathways to provide effective host defence against pathogen infections. However, TLR-signalling pathways are also likely to stringently regulate tissue maintenance and homeostasis by elaborate modulatory mechanisms. MicroRNAs (miRNAs) have emerged as key regulators and as an essential part of the networks involved in regulating TLR-signalling pathways. In this review, we highlight our understanding of the regulation of miRNA expression profiles by TLR-signalling pathways and the regulation of TLR-signalling pathways by miRNAs. We focus on the roles of miRNAs in regulating TLR-signalling pathways by targeting multiple molecules, including TLRs themselves, their associated signalling proteins and regulatory molecules, and transcription factors and functional cytokines induced by them, at multiple levels. PMID:24772440

  18. 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...

  19. Trim69 regulates zebrafish brain development by ap-1 pathway

    PubMed Central

    Han, Ruiqin; Wang, Renxian; Zhao, Qing; Han, Yongqing; Zong, Shudong; Miao, Shiying; Song, Wei; Wang, Linfang

    2016-01-01

    Proteins belonging to the TRIM family have been implicated in a variety of cellular processes such as apoptosis, differentiation, neurogenesis, muscular physiology and innate immune responses. Trim69, previously identified as a novel gene cloned from a human testis cDNA library, has a homologous gene in zebrafish and this study focused on investigating the function of trim69 in zebrafish neurogenesis. Trim69 was found to be expressed in zebrafish embryo brain at the early stages. Knockdown of trim69 led to deformed brain development, obvious signs of apoptosis present in the head, and decreased expression of neuronal differentiation and stem cell markers. This phenotype was rescued upon co-injection of human mRNA together along with the trim69 knockdown. Results of this study also showed an interaction between TRIM69 and c-Jun in human cells, and upon TRIM69 knock down c-Jun expression subsequently increased, whereas the over-expression of TRIM69 led to the down-regulation of c-Jun. Additionally, knockdown both c-Jun and trim69 can rescue the deformed brain, evident cellular apoptosis in the head and decreased expression of neuronal differentiation and stem cell markers. Overall, our results support a role for trim69 in the development of the zebrafish brain through ap-1 pathway. PMID:27050765

  20. Dynamic optimization identifies optimal programmes for pathway regulation in prokaryotes.

    PubMed

    Bartl, Martin; Kötzing, Martin; Schuster, Stefan; Li, Pu; Kaleta, Christoph

    2013-01-01

    To survive in fluctuating environmental conditions, microorganisms must be able to quickly react to environmental challenges by upregulating the expression of genes encoding metabolic pathways. Here we show that protein abundance and protein synthesis capacity are key factors that determine the optimal strategy for the activation of a metabolic pathway. If protein abundance relative to protein synthesis capacity increases, the strategies shift from the simultaneous activation of all enzymes to the sequential activation of groups of enzymes and finally to a sequential activation of individual enzymes along the pathway. In the case of pathways with large differences in protein abundance, even more complex pathway activation strategies with a delayed activation of low abundance enzymes and an accelerated activation of high abundance enzymes are optimal. We confirm the existence of these pathway activation strategies as well as their dependence on our proposed constraints for a large number of metabolic pathways in several hundred prokaryotes.

  1. Altered Pathway Analyzer: A gene expression dataset analysis tool for identification and prioritization of differentially regulated and network rewired pathways

    PubMed Central

    Kaushik, Abhinav; Ali, Shakir; Gupta, Dinesh

    2017-01-01

    Gene connection rewiring is an essential feature of gene network dynamics. Apart from its normal functional role, it may also lead to dysregulated functional states by disturbing pathway homeostasis. Very few computational tools measure rewiring within gene co-expression and its corresponding regulatory networks in order to identify and prioritize altered pathways which may or may not be differentially regulated. We have developed Altered Pathway Analyzer (APA), a microarray dataset analysis tool for identification and prioritization of altered pathways, including those which are differentially regulated by TFs, by quantifying rewired sub-network topology. Moreover, APA also helps in re-prioritization of APA shortlisted altered pathways enriched with context-specific genes. We performed APA analysis of simulated datasets and p53 status NCI-60 cell line microarray data to demonstrate potential of APA for identification of several case-specific altered pathways. APA analysis reveals several altered pathways not detected by other tools evaluated by us. APA analysis of unrelated prostate cancer datasets identifies sample-specific as well as conserved altered biological processes, mainly associated with lipid metabolism, cellular differentiation and proliferation. APA is designed as a cross platform tool which may be transparently customized to perform pathway analysis in different gene expression datasets. APA is freely available at http://bioinfo.icgeb.res.in/APA. PMID:28084397

  2. Dynamic regulation of genetic pathways and targets during aging in Caenorhabditis elegans.

    PubMed

    He, Kan; Zhou, Tao; Shao, Jiaofang; Ren, Xiaoliang; Zhao, Zhongying; Liu, Dahai

    2014-03-01

    Numerous genetic targets and some individual pathways associated with aging have been identified using the worm model. However, less is known about the genetic mechanisms of aging in genome wide, particularly at the level of multiple pathways as well as the regulatory networks during aging. Here, we employed the gene expression datasets of three time points during aging in Caenorhabditis elegans (C. elegans) and performed the approach of gene set enrichment analysis (GSEA) on each dataset between adjacent stages. As a result, multiple genetic pathways and targets were identified as significantly down- or up-regulated. Among them, 5 truly aging-dependent signaling pathways including MAPK signaling pathway, mTOR signaling pathway, Wnt signaling pathway, TGF-beta signaling pathway and ErbB signaling pathway as well as 12 significantly associated genes were identified with dynamic expression pattern during aging. On the other hand, the continued declines in the regulation of several metabolic pathways have been demonstrated to display age-related changes. Furthermore, the reconstructed regulatory networks based on three of aging related Chromatin immunoprecipitation experiments followed by sequencing (ChIP-seq) datasets and the expression matrices of 154 involved genes in above signaling pathways provide new insights into aging at the multiple pathways level. The combination of multiple genetic pathways and targets needs to be taken into consideration in future studies of aging, in which the dynamic regulation would be uncovered.

  3. Nrf2-driven TERT regulates pentose phosphate pathway in glioblastoma

    PubMed Central

    Ahmad, F; Dixit, D; Sharma, V; Kumar, A; Joshi, S D; Sarkar, C; Sen, E

    2016-01-01

    Given the involvement of telomerase activation and dysregulated metabolism in glioma progression, the connection between these two critical players was investigated. Pharmacological inhibition of human Telomerase reverse transcriptase (hTERT) by Costunolide induced glioma cell apoptosis in a reactive oxygen species (ROS)-dependent manner. Costunolide induced an ROS-dependent increase in p53 abrogated telomerase activity. Costunolide decreased Nrf2 level; and ectopic Nrf2 expression decreased Costunolide-induced ROS generation. While TERT knock-down abrogated Nrf2 levels, overexpression of Nrf2 increased TERT expression. Inhibition of hTERT either by Costunolide, or by siRNA or dominant-negative hTERT (DN-hTERT) abrogated (i) expression of Glucose-6-phosphate dehydrogenase (G6PD) and Transketolase (TKT) – two major nodes in the pentose phosphate (PPP) pathway; and (ii) phosphorylation of glycogen synthase (GS). hTERT knock-down decreased TKT activity and increased glycogen accumulation. Interestingly, siRNA-mediated knock-down of TKT elevated glycogen accumulation. Coherent with the in vitro findings, Costunolide reduced tumor burden in heterotypic xenograft glioma mouse model. Costunolide-treated tumors exhibited diminished TKT activity, heightened glycogen accumulation, and increased senescence. Importantly, glioblastoma multiforme (GBM) patient tumors bearing TERT promoter mutations (C228T and C250T) known to be associated with increased telomerase activity; exhibited elevated Nrf2 and TKT expression and decreased glycogen accumulation. Taken together, our findings highlight the previously unknown (i) role of telomerase in the regulation of PPP and glycogen accumulation and (ii) the involvement of Nrf2-TERT loop in maintaining oxidative defense responses in glioma cells. PMID:27148686

  4. Molecular Pathways: Dietary Regulation of Stemness and Tumor Initiation by the PPAR-δ Pathway.

    PubMed

    Beyaz, Semir; Yilmaz, Ömer H

    2016-12-01

    Peroxisome proliferator-activated receptor delta (PPAR-δ) is a nuclear receptor transcription factor that regulates gene expression during development and disease states, such as cancer. However, the precise role of PPAR-δ during tumorigenesis is not well understood. Recent data suggest that PPAR-δ may have context-specific oncogenic and tumor-suppressive roles depending on the tissue, cell-type, or diet-induced physiology in question. For example, in the intestine, pro-obesity diets, such as a high-fat diet (HFD), are associated with increased colorectal cancer incidence. Interestingly, many of the effects of an HFD in the stem and progenitor cell compartment are driven by a robust PPAR-δ program and contribute to the early steps of intestinal tumorigenesis. Importantly, the PPAR-δ pathway or its downstream mediators may serve as therapeutic intervention points or biomarkers in colon cancer that arise in patients who are obese. Although potent PPAR-δ agonists and antagonists exist, their clinical utility may be enhanced by uncovering how PPAR-δ mediates tumorigenesis in diverse tissues and cell types as well as in response to diet. Clin Cancer Res; 22(23); 5636-41. ©2016 AACR.

  5. Schisandra Lignan Extract Protects against Carbon Tetrachloride-Induced Liver Injury in Mice by Inhibiting Oxidative Stress and Regulating the NF-κB and JNK Signaling Pathways

    PubMed Central

    Chen, Qingshan; Zhan, Qi; Li, Ying; Sun, Sen; Zhao, Liang

    2017-01-01

    Schisandra chinensis (S. chinensis) is a traditional Chinese herbal medicine widely used for the treatment of liver disease, whose main active components are lignans. However, the action mechanisms of the lignans in S. chinensis remain unclear. This study aimed to investigate the protective effect and related molecular mechanism of Schisandra lignan extract (SLE) against carbon tetrachloride- (CCl4-) induced acute liver injury in mice. Different doses of SLE at 50, 100, and 200 mg/kg were administered daily by gavage for 5 days before CCl4 treatment. The results showed that SLE significantly decreased the activities of serum ALT/AST and reduced liver pathologic changes induced by CCl4. Pretreatment with SLE not only decreased the content of MDA but increased SOD, GSH, and GSH-Px activities in the liver, suggesting that SLE attenuated CCl4-induced oxidative stress. The expression levels of inflammatory cytokines TNF-a, IL-1β, and IL-6 were decreased after oral administration of SLE, probably because lignans inhibited the NF-κB activity. Additionally, SLE also inhibited hepatocyte apoptosis by suppressing JNK activation and regulating Bcl-2/Bax signaling pathways. In conclusion, these results suggested that SLE prevented CCl4-induced liver injury through a combination of antioxidative stress, anti-inflammation, and antihepatocyte apoptosis and alleviated inflammation and apoptosis by regulating the NF-κB, JNK, and Bcl-2/Bax signaling pathways. PMID:28246539

  6. Strigolactones' ability to regulate root development may be executed by induction of the ethylene pathway.

    PubMed

    Koltai, Hinanit

    2011-07-01

    The newly defined phytohormones strigolactones (SLs) were recently shown to act as regulators of root development. Their positive effect on root-hair (RH) elongation enabled examination of their cross talk with auxin and ethylene. Analysis of wild-type plants and hormone-signaling mutants combined with hormonal treatments suggested that SLs and ethylene regulate RH elongation via a common regulatory pathway, in which ethylene is epistatic to SLs. The SL and auxin hormonal pathways were suggested to converge for regulation of RH elongation; this convergence was suggested to be mediated via the ethylene pathway, and to include regulation of auxin transport.

  7. Intracellular pathways regulating ciliary beating of rat brain ependymal cells

    PubMed Central

    Nguyen, Thien; Chin, Wei-Chun; O’Brien, Jennifer A; Verdugo, Pedro; Berger, Albert J

    2001-01-01

    The mammalian brain ventricles are lined with ciliated ependymal cells. As yet little is known about the mechanisms by which neurotransmitters regulate cilia beat frequency (CBF). Application of 5-HT to ependymal cells in cultured rat brainstem slices caused CBF to increase. 5-HT had an EC50 of 30 μM and at 100 μM attained a near-maximal CBF increase of 52.7 ± 4.1 % (mean ± s.d.) (n= 8). Bathing slices in Ca2+-free solution markedly reduced the 5-HT-mediated increase in CBF. Fluorescence measurements revealed that 5-HT caused a marked transient elevation in cytosolic Ca2+ ([Ca2+]c) that then slowly decreased to a plateau level. Analysis showed that the [Ca2+]c transient was due to release of Ca2+ from inositol 1,4,5-trisphosphate (IP3)-sensitive stores; the plateau was probably due to extracellular Ca2+ influx through Ca2+ release-activated Ca2+ (CRAC) channels. Application of ATP caused a sustained decrease in CBF. ATP had an EC50 of about 50 μM and 100 μM ATP resulted in a maximal 57.5 ± 6.5 % (n= 12) decrease in CBF. The ATP-induced decrease in CBF was unaffected by lowering extracellular [Ca2+], and no changes in [Ca2+]c were observed. Exposure of ependymal cells to forskolin caused a decrease in CBF. Ciliated ependymal cells loaded with caged cAMP exhibited a 54.3 ± 7.5 % (n= 9) decrease in CBF following uncaging. These results suggest that ATP reduces CBF by a Ca2+-independent cAMP-mediated pathway. Application of 5-HT and adenosine-5′-O-3-thiotriphosphate (ATP-γ-S) to acutely isolated ciliated ependymal cells resulted in CBF responses similar to those of ependymal cells in cultured slices suggesting that these neurotransmitters act directly on these cells. The opposite response of ciliated ependymal cells to 5-HT and ATP provides a novel mechanism for their active involvement in central nervous system signalling. PMID:11179397

  8. Regulation of cross-talk in yeast MAPK signaling pathways.

    PubMed

    Saito, Haruo

    2010-12-01

    MAP kinase (MAPK) modules are conserved three-kinase cascades that serve central roles in intracellular signal transduction in eukaryotic cells. MAPK pathways of different inputs and outputs use overlapping sets of signaling components. In yeast, for example, three MAPK pathways (pheromone response, filamentous growth response, and osmostress adaptation) all use the same Ste11 MAPK kinase kinase (MAPKKK). How undesirable leakage of signal, or cross-talk, is prevented between these pathways has been a subject of intensive study. This review discusses recent findings from yeast that indicate that there is no single mechanism, but that a combination of four general strategies (docking interactions, scaffold proteins, cross-pathway inhibition, and kinetic insulation) are utilized for the prevention of cross-talk between any two MAPK modules.

  9. Signaling pathways regulating cartilage growth plate formation and activity.

    PubMed

    Samsa, William E; Zhou, Xin; Zhou, Guang

    2017-02-01

    The growth plate is a highly specialized and dynamic cartilage structure that serves many essential functions in skeleton patterning, growth and endochondral ossification in developing vertebrates. Major signaling pathways initiated by classical morphogens and by other systemic and tissue-specific factors are intimately involved in key aspects of growth plate development. As a corollary of these essential functions, disturbances in these pathways due to mutations or environmental factors lead to severe skeleton disorders. Here, we review these pathways and the most recent progress made in understanding their roles in chondrocyte differentiation in growth plate development and activity. Furthermore, we discuss newly uncovered pathways involved in growth plate formation, including mTOR, the circadian clock, and the COP9 signalosome.

  10. Logical Analysis of Regulation of Interleukin-12 Expression Pathway Regulation During HCV Infection.

    PubMed

    Farooqi, Zia-Ur-Rehman; Tareen, Samar H K; Ahmed, Jamil; Zaidi, Najam-Us-Sahar S

    2016-01-01

    Hepatitis C virus (HCV) triggers coordinated innate and adaptive response in host cell. HCV genome and proteins of the replicating virus are recognized as non-self-antigens by host cell to activate Toll Like Receptors (TLRs). Activated TLRs ultimately express cytokines, which can clear virus either by activating interferon (IFN), protein kinase C (PKC) and RNA Lase system or through activation of cytotoxic T-lymphocytes. Interleukin-12 (IL-12) is a potent antiviral cytokine, capable of clearing HCV by bridging both innate and adaptive antiviral immune response. Activation of TLR-4 on macrophages surface induces expression of IL-12 via NF-κB and AP-1 transcriptional pathway. After expression, IL- 12 releases IFN-γ, which activates anti-HCV cytotoxic lymphocytes. Conversely, in chronic HCV infection downregulation of IL-12 has been reported instead of by number of studies. Keeping in view of the above mentioned facts, this study was designed to evaluate HCV-core mediated down-regulation of IL-12 transcriptional pathway by employing a logical modeling approach based on the Ren´e Thomas formalism. The logical parameters of entities were estimated by using SMBioNet. The Logical model represents all possible dynamics of protein expression involved during course of HCV pathology. Results demonstrated that at chronic stage of infection, though TLR-4 was constantly active but yet it failed to express the NF-κB, AP-1, IL-12 and IFN-γ. This mechanism was indicative of incorporation of core mediated changes in IL-12 regulatory pathway. Moreover, results also indicate that HCV adopts different trajectories to accomplish the persistence of chronic phase of infection. It also implicated that human immune system tries to clear HCV but core is capable of inducing system oscillations to evade the immunity.

  11. Meaningful relationships: the regulation of the Ras/Raf/MEK/ERK pathway by protein interactions.

    PubMed Central

    Kolch, W

    2000-01-01

    The Ras/Raf/MEK (mitogen-activated protein kinase/ERK kinase)/ERK (extracellular-signal-regulated kinase) pathway is at the heart of signalling networks that govern proliferation, differentiation and cell survival. Although the basic regulatory steps have been elucidated, many features of this pathway are only beginning to emerge. This review focuses on the role of protein-protein interactions in the regulation of this pathway, and how they contribute to co-ordinate activation steps, subcellular redistribution, substrate phosphorylation and cross-talk with other signalling pathways. PMID:11023813

  12. Carbohydrate metabolism in Archaea: current insights into unusual enzymes and pathways and their regulation.

    PubMed

    Bräsen, Christopher; Esser, Dominik; Rauch, Bernadette; Siebers, Bettina

    2014-03-01

    The metabolism of Archaea, the third domain of life, resembles in its complexity those of Bacteria and lower Eukarya. However, this metabolic complexity in Archaea is accompanied by the absence of many "classical" pathways, particularly in central carbohydrate metabolism. Instead, Archaea are characterized by the presence of unique, modified variants of classical pathways such as the Embden-Meyerhof-Parnas (EMP) pathway and the Entner-Doudoroff (ED) pathway. The pentose phosphate pathway is only partly present (if at all), and pentose degradation also significantly differs from that known for bacterial model organisms. These modifications are accompanied by the invention of "new," unusual enzymes which cause fundamental consequences for the underlying regulatory principles, and classical allosteric regulation sites well established in Bacteria and Eukarya are lost. The aim of this review is to present the current understanding of central carbohydrate metabolic pathways and their regulation in Archaea. In order to give an overview of their complexity, pathway modifications are discussed with respect to unusual archaeal biocatalysts, their structural and mechanistic characteristics, and their regulatory properties in comparison to their classic counterparts from Bacteria and Eukarya. Furthermore, an overview focusing on hexose metabolic, i.e., glycolytic as well as gluconeogenic, pathways identified in archaeal model organisms is given. Their energy gain is discussed, and new insights into different levels of regulation that have been observed so far, including the transcript and protein levels (e.g., gene regulation, known transcription regulators, and posttranslational modification via reversible protein phosphorylation), are presented.

  13. Adverse Outcome Pathways: From Research to Regulation - Scientific Workshop Report

    EPA Science Inventory

    An adverse outcome pathway (AOP) organizes existing knowledge on chemical mode of action, starting with a molecular initiating event such as receptor binding, continuing through key events, and ending with an adverse outcome such as reproductive impairment. AOPs can help identify...

  14. Post-transcriptional regulation of interferons and their signaling pathways.

    PubMed

    Savan, Ram

    2014-05-01

    Interferons (IFNs) are low molecular weight cell-derived proteins that include the type I, II, and III IFN families. IFNs are critical for an optimal immune response during microbial infections while dysregulated expression can lead to autoimmune diseases. Given its role in disease, it is important to understand cellular mechanisms of IFN regulation. 3' untranslated regions (3' UTRs) have emerged as potent regulators of mRNA and protein dosage and are controlled through multiple regulatory elements including adenylate uridylate (AU)-rich elements (AREs) and microRNA (miRNA) recognition elements. These AREs are targeted by RNA-binding proteins (ARE-BPs) for degradation and/or stabilization through an ARE-mediated decay process. miRNA are endogenous, single-stranded RNA molecules ~22 nucleotides in length that regulate mRNA translation through the miRNA-induced silencing complex. IFN transcripts, like other labile mRNAs, harbor AREs in their 3' UTRs that dictate the turnover of mRNA. This review is a survey of the literature related to IFN regulation by miRNA, ARE-BPs, and how these complexes interact dynamically on the 3' UTR. Additionally, downstream effects of these post-transcriptional regulators on the immune response will be discussed. Review topics include past studies, current understanding, and future challenges in the study of post-transcriptional regulation affecting IFN responses.

  15. Post-Transcriptional Regulation of Interferons and Their Signaling Pathways

    PubMed Central

    2014-01-01

    Interferons (IFNs) are low molecular weight cell-derived proteins that include the type I, II, and III IFN families. IFNs are critical for an optimal immune response during microbial infections while dysregulated expression can lead to autoimmune diseases. Given its role in disease, it is important to understand cellular mechanisms of IFN regulation. 3′ untranslated regions (3′ UTRs) have emerged as potent regulators of mRNA and protein dosage and are controlled through multiple regulatory elements including adenylate uridylate (AU)-rich elements (AREs) and microRNA (miRNA) recognition elements. These AREs are targeted by RNA-binding proteins (ARE-BPs) for degradation and/or stabilization through an ARE-mediated decay process. miRNA are endogenous, single-stranded RNA molecules ∼22 nucleotides in length that regulate mRNA translation through the miRNA-induced silencing complex. IFN transcripts, like other labile mRNAs, harbor AREs in their 3′ UTRs that dictate the turnover of mRNA. This review is a survey of the literature related to IFN regulation by miRNA, ARE-BPs, and how these complexes interact dynamically on the 3′ UTR. Additionally, downstream effects of these post-transcriptional regulators on the immune response will be discussed. Review topics include past studies, current understanding, and future challenges in the study of post-transcriptional regulation affecting IFN responses. PMID:24702117

  16. The Retinoblastoma pathway regulates stem cell proliferation in freshwater planarians.

    PubMed

    Zhu, Shu Jun; Pearson, Bret J

    2013-01-15

    Freshwater planarians are flatworms of the Lophotrochozoan superphylum and are well known for their regenerative abilities, which rely on a large population of pluripotent adult stem cells. However, the mechanisms by which planarians maintain a precise population of adult stem cells while balancing proliferation and cell death, remain to be elucidated. Here we have identified, characterized, and functionally tested the core Retinoblastoma (Rb) pathway components in planarian adult stem cell biology. The Rb pathway is an ancient and conserved mechanism of proliferation control from plants to animals and is composed of three core components: an Rb protein, and a transcription factor heterodimer of E2F and DP proteins. Although the planarian genome contains all components of the Rb pathway, we found that they have undergone gene loss from the ancestral state, similar to other species in their phylum. The single Rb homolog (Smed-Rb) was highly expressed in planarian stem cells and was required for stem cell maintenance, similar to the Rb-homologs p107 and p130 in vertebrates. We show that planarians and their phylum have undergone the most severe reduction in E2F genes observed thus far, and the single remaining E2F was predicted to be a repressive-type E2F (Smed-E2F4-1). Knockdown of either Smed-E2F4-1 or its dimerization partner Dp (Smed-Dp) by RNAi resulted in temporary hyper-proliferation. Finally, we showed that known Rb-interacting genes in other systems, histone deacetylase 1 and cyclinD (Smed-HDAC1; Smed-cycD), were similar to Rb in expression and phenotypes when knocked down by RNAi, suggesting that these established interactions with Rb may also be conserved in planarians. Together, these results showed that planarians use the conserved components of the Rb tumor suppressor pathway to control proliferation and cell survival.

  17. Circadian acetylome reveals regulation of mitochondrial metabolic pathways.

    PubMed

    Masri, Selma; Patel, Vishal R; Eckel-Mahan, Kristin L; Peleg, Shahaf; Forne, Ignasi; Ladurner, Andreas G; Baldi, Pierre; Imhof, Axel; Sassone-Corsi, Paolo

    2013-02-26

    The circadian clock is constituted by a complex molecular network that integrates a number of regulatory cues needed to maintain organismal homeostasis. To this effect, posttranslational modifications of clock proteins modulate circadian rhythms and are thought to convert physiological signals into changes in protein regulatory function. To explore reversible lysine acetylation that is dependent on the clock, we have characterized the circadian acetylome in WT and Clock-deficient (Clock(-/-)) mouse liver by quantitative mass spectrometry. Our analysis revealed that a number of mitochondrial proteins involved in metabolic pathways are heavily influenced by clock-driven acetylation. Pathways such as glycolysis/gluconeogenesis, citric acid cycle, amino acid metabolism, and fatty acid metabolism were found to be highly enriched hits. The significant number of metabolic pathways whose protein acetylation profile is altered in Clock(-/-) mice prompted us to link the acetylome to the circadian metabolome previously characterized in our laboratory. Changes in enzyme acetylation over the circadian cycle and the link to metabolite levels are discussed, revealing biological implications connecting the circadian clock to cellular metabolic state.

  18. Prefrontal–striatal pathway underlies cognitive regulation of craving

    PubMed Central

    Kober, Hedy; Mende-Siedlecki, Peter; Kross, Ethan F.; Weber, Jochen; Mischel, Walter; Hart, Carl L.; Ochsner, Kevin N.

    2010-01-01

    The ability to control craving for substances that offer immediate rewards but whose long-term consumption may pose serious risks lies at the root of substance use disorders and is critical for mental and physical health. Despite its importance, the neural systems supporting this ability remain unclear. Here, we investigated this issue using functional imaging to examine neural activity in cigarette smokers, the most prevalent substance-dependent population in the United States, as they used cognitive strategies to regulate craving for cigarettes and food. We found that the cognitive down-regulation of craving was associated with (i) activity in regions previously associated with regulating emotion in particular and cognitive control in general, including dorsomedial, dorsolateral, and ventrolateral prefrontal cortices, and (ii) decreased activity in regions previously associated with craving, including the ventral striatum, subgenual cingulate, amygdala, and ventral tegmental area. Decreases in craving correlated with decreases in ventral striatum activity and increases in dorsolateral prefrontal cortex activity, with ventral striatal activity fully mediating the relationship between lateral prefrontal cortex and reported craving. These results provide insight into the mechanisms that enable cognitive strategies to effectively regulate craving, suggesting that it involves neural dynamics parallel to those involved in regulating other emotions. In so doing, this study provides a methodological tool and conceptual foundation for studying this ability across substance using populations and developing more effective treatments for substance use disorders. PMID:20679212

  19. Prefrontal-striatal pathway underlies cognitive regulation of craving.

    PubMed

    Kober, Hedy; Mende-Siedlecki, Peter; Kross, Ethan F; Weber, Jochen; Mischel, Walter; Hart, Carl L; Ochsner, Kevin N

    2010-08-17

    The ability to control craving for substances that offer immediate rewards but whose long-term consumption may pose serious risks lies at the root of substance use disorders and is critical for mental and physical health. Despite its importance, the neural systems supporting this ability remain unclear. Here, we investigated this issue using functional imaging to examine neural activity in cigarette smokers, the most prevalent substance-dependent population in the United States, as they used cognitive strategies to regulate craving for cigarettes and food. We found that the cognitive down-regulation of craving was associated with (i) activity in regions previously associated with regulating emotion in particular and cognitive control in general, including dorsomedial, dorsolateral, and ventrolateral prefrontal cortices, and (ii) decreased activity in regions previously associated with craving, including the ventral striatum, subgenual cingulate, amygdala, and ventral tegmental area. Decreases in craving correlated with decreases in ventral striatum activity and increases in dorsolateral prefrontal cortex activity, with ventral striatal activity fully mediating the relationship between lateral prefrontal cortex and reported craving. These results provide insight into the mechanisms that enable cognitive strategies to effectively regulate craving, suggesting that it involves neural dynamics parallel to those involved in regulating other emotions. In so doing, this study provides a methodological tool and conceptual foundation for studying this ability across substance using populations and developing more effective treatments for substance use disorders.

  20. MiRSEA: Discovering the pathways regulated by dysfunctional MicroRNAs

    PubMed Central

    Xu, Yanjun; Jiang, Ying; Zhang, Chunlong; Li, Chunquan; Li, Xia

    2016-01-01

    Recent studies have shown that dysfunctional microRNAs (miRNAs) are involved in the progression of various cancers. Dysfunctional miRNAs may jointly regulate their target genes and further alter the activities of canonical biological pathways. Identification of the pathways regulated by a group of dysfunctional miRNAs could help uncover the pathogenic mechanisms of cancer and facilitate development of new drug targets. Current miRNA-pathway analyses mainly use differentially-expressed miRNAs to predict the shared pathways on which they act. However, these methods fail to consider the level of differential expression level, which could improve our understanding of miRNA function. We propose a novel computational method, MicroRNA Set Enrichment Analysis (MiRSEA), to identify the pathways regulated by dysfunctional miRNAs. MiRSEA integrates the differential expression levels of miRNAs with the strength of miRNA pathway associations to perform direct enrichment analysis using miRNA expression data. We describe the MiRSEA methodology and illustrate its effectiveness through analysis of data from hepatocellular cancer, gastric cancer and lung cancer. With these analyses, we show that MiRSEA can successfully detect latent biological pathways regulated by dysfunctional miRNAs. We have implemented MiRSEA as a freely available R-based package on CRAN (https://cran.r-project.org/web/packages/MiRSEA/). PMID:27474169

  1. Method to determine transcriptional regulation pathways in organisms

    DOEpatents

    Gardner, Timothy S.; Collins, James J.; Hayete, Boris; Faith, Jeremiah

    2012-11-06

    The invention relates to computer-implemented methods and systems for identifying regulatory relationships between expressed regulating polypeptides and targets of the regulatory activities of such regulating polypeptides. More specifically, the invention provides a new method for identifying regulatory dependencies between biochemical species in a cell. In particular embodiments, provided are computer-implemented methods for identifying a regulatory interaction between a transcription factor and a gene target of the transcription factor, or between a transcription factor and a set of gene targets of the transcription factor. Further provided are genome-scale methods for predicting regulatory interactions between a set of transcription factors and a corresponding set of transcriptional target substrates thereof.

  2. Regulation of Hh/Gli signaling by dual ubiquitin pathways.

    PubMed

    Jiang, Jin

    2006-11-01

    The Hedgehog (Hh) signaling pathway governs cell growth and patterning in animal development. Malfunction of several pathway components, including the key transcriptional effector Ci/Gli proteins, leads to a variety of human disorders including several malignancies. Ci/Gli activity is controlled by multi-layered regulatory mechanisms, the most prominent of which is the ubiquitin-mediated proteolysis. In the absence of Hh, Ci/Gli is proteolytically processed into a truncated form that functions as a transcriptional repressor of the Hh pathway. Ci processing is mediated by an SCF (Skip1/Cul1/F-box protein) ubiquitin ligase in which the F-box protein Slimb/beta-TRCP bridges Ci to the ubiquitin ligase. Recent studies in Drosophila and mammalian cultured cells have demonstrated that sequential phosphorylation of Ci/Gli by PKA, GSK3, and CKI creates multiple docking sites that can recruit SCF(Slimb/beta-TRCP), which then promotes Ci/Gli ubiquitination followed by proteasome-mediated processing. Recently, an E3 ubiquitin ligase consisting of the BTB (Broad Complex, Tramtrack, and Bric a Brac) protein HIB (Hh induced MATH and BTB protein) and Cullin 3 (Cul3) has been identified that acts in a negative feedback loop to fine-tune Hh signaling responses by degrading full length Ci. In eye imaginal discs where Hh signals coordinate cell proliferation and differentiation, HIB is highly expressed in the differentiating cells to prevent aberrant Hh signaling activity and ensure normal eye development. Tissue- and developmental stage-specific expression of HIB and its homologs in vertebrates may provide a conserved mechanism for ensuring precision in spatial and temporal control of Hh signaling.

  3. The Lysosome Rupture-activated TAK1-JNK Pathway Regulates NLRP3 Inflammasome Activation*

    PubMed Central

    Okada, Masahiro; Matsuzawa, Atsushi; Yoshimura, Akihiko; Ichijo, Hidenori

    2014-01-01

    Lysosome rupture triggers NLRP3 inflammasome activation in macrophages. However, the underlying mechanism is not fully understood. Here we showed that the TAK1-JNK pathway, a MAPK signaling pathway, is activated through lysosome rupture and that this activation is necessary for the complete activation of the NLRP3 inflammasome through the oligomerization of an adapter protein, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC). We also revealed that the activation of the TAK1-JNK pathway is sustained through Ca2+ ions and that calcium/calmodulin-dependent protein kinase type II functions upstream of the TAK1-JNK pathway and specifically regulates lysosome rupture-induced NLRP3 inflammasome activation. These data suggest a novel role for the TAK1-JNK pathway as a critical regulator of NLRP3 inflammasome activation. PMID:25288801

  4. Transcriptomic analysis in the developing zebrafish embryo after compound exposure: Individual gene expression and pathway regulation

    SciTech Connect

    Hermsen, Sanne A.B.; Pronk, Tessa E.; Brandhof, Evert-Jan van den; Ven, Leo T.M. van der; Piersma, Aldert H.

    2013-10-01

    The zebrafish embryotoxicity test is a promising alternative assay for developmental toxicity. Classically, morphological assessment of the embryos is applied to evaluate the effects of compound exposure. However, by applying differential gene expression analysis the sensitivity and predictability of the test may be increased. For defining gene expression signatures of developmental toxicity, we explored the possibility of using gene expression signatures of compound exposures based on commonly expressed individual genes as well as based on regulated gene pathways. Four developmental toxic compounds were tested in concentration-response design, caffeine, carbamazepine, retinoic acid and valproic acid, and two non-embryotoxic compounds, D-mannitol and saccharin, were included. With transcriptomic analyses we were able to identify commonly expressed genes, which were mostly development related, after exposure to the embryotoxicants. We also identified gene pathways regulated by the embryotoxicants, suggestive of their modes of action. Furthermore, whereas pathways may be regulated by all compounds, individual gene expression within these pathways can differ for each compound. Overall, the present study suggests that the use of individual gene expression signatures as well as pathway regulation may be useful starting points for defining gene biomarkers for predicting embryotoxicity. - Highlights: • The zebrafish embryotoxicity test in combination with transcriptomics was used. • We explored two approaches of defining gene biomarkers for developmental toxicity. • Four compounds in concentration-response design were tested. • We identified commonly expressed individual genes as well as regulated gene pathways. • Both approaches seem suitable starting points for defining gene biomarkers.

  5. microRNA regulation of Wnt signaling pathways in development and disease

    PubMed Central

    Song, Jia L.; Nigam, Priya; Tektas, Senel S.; Selva, Erica

    2015-01-01

    Wnt signaling pathways and microRNAs (miRNAs) are critical regulators of development. Aberrant Wnt signaling pathways and miRNA levels lead to developmental defects and diverse human pathologies including but not limited to cancer. Wnt signaling pathways regulate a plethora of cellular processes during embryonic development and maintain homeostasis of adult tissues. A majority of Wnt signaling components are regulated by miRNAs which are small noncoding RNAs that are expressed in both animals and plants. In animal cells, miRNAs fine tune gene expression by pairing primarily to the 3′untranslated region of protein coding mRNAs to repress target mRNA translation and/or induce target degradation. miRNA-mediated regulation of signaling transduction pathways is important in modulating dose-sensitive response of cells to signaling molecules. This review discusses components of the Wnt signaling pathways that are regulated by miRNAs in the context of development and diseases. A fundamental understanding of miRNA functions in Wnt signaling transduction pathways may yield new insight into crosstalks of regulatory mechanisms essential for development and disease pathophysiology leading to novel therapeutics. PMID:25843779

  6. Regulation of mat responses by a differentiation MAPK pathway in Saccharomyces cerevisiae.

    PubMed

    Karunanithi, Sheelarani; Joshi, Jyoti; Chavel, Colin; Birkaya, Barbara; Grell, Laura; Cullen, Paul J

    2012-01-01

    Fungal species exhibit diverse behaviors when presented with extracellular challenges. Pathogenic fungi can undergo cell differentiation and biofilm formation in response to fluctuating nutrient levels, and these responses are required for virulence. In the model fungal eukaryote Saccharomyces cerevisiae, nutrient limitation induces filamentous growth and biofilm/mat formation. Both responses require the same signal transduction (MAPK) pathway and the same cell adhesion molecule (Flo11) but have been studied under different conditions. We found that filamentous growth and mat formation are aspects of a related response that is regulated by the MAPK pathway. Cells in yeast-form mats differentiated into pseudohyphae in response to nutrient limitation. The MAPK pathway regulated mat expansion (in the plane of the XY-axis) and substrate invasion (downward in the plane of the Z-axis), which optimized the mat's response to extracellular nutrient levels. The MAPK pathway also regulated an upward growth pattern (in the plane of the Z-axis) in response to nutrient limitation and changes in surface rigidity. Upward growth allowed for another level of mat responsiveness and resembled a type of colonial chemorepulsion. Together our results show that signaling pathways play critical roles in regulating social behaviors in which fungal cells participate. Signaling pathways may regulate similar processes in pathogens, whose highly nuanced responses are required for virulence.

  7. Chemical combinations elucidate pathway interactions and regulation relevant to Hepatitis C replication

    PubMed Central

    Owens, Christopher M; Mawhinney, Christina; Grenier, Jill M; Altmeyer, Ralf; Lee, Margaret S; Borisy, Alexis A; Lehár, Joseph; Johansen, Lisa M

    2010-01-01

    The search for effective Hepatitis C antiviral therapies has recently focused on host sterol metabolism and protein prenylation pathways that indirectly affect viral replication. However, inhibition of the sterol pathway with statin drugs has not yielded consistent results in patients. Here, we present a combination chemical genetic study to explore how the sterol and protein prenylation pathways work together to affect hepatitis C viral replication in a replicon assay. In addition to finding novel targets affecting viral replication, our data suggest that the viral replication is strongly affected by sterol pathway regulation. There is a marked transition from antagonistic to synergistic antiviral effects as the combination targets shift downstream along the sterol pathway. We also show how pathway regulation frustrates potential hepatitis C therapies based on the sterol pathway, and reveal novel synergies that selectively inhibit hepatitis C replication over host toxicity. In particular, combinations targeting the downstream sterol pathway enzymes produced robust and selective synergistic inhibition of hepatitis C replication. Our findings show how combination chemical genetics can reveal critical pathway connections relevant to viral replication, and can identify potential treatments with an increased therapeutic window. PMID:20531405

  8. Mechanisms and pathways of innate immune activation and regulation in health and cancer

    PubMed Central

    Cui, Jun; Chen, Yongjun; Wang, Helen Y; Wang, Rong-Fu

    2015-01-01

    Research on innate immune signaling and regulation has recently focused on pathogen recognition receptors (PRRs) and their signaling pathways. Members of PRRs sense diverse microbial invasions or danger signals, and initiate innate immune signaling pathways, leading to proinflammatory cytokines production, which, in turn, instructs adaptive immune response development. Despite the diverse functions employed by innate immune signaling to respond to a variety of different pathogens, the innate immune response must be tightly regulated. Otherwise, aberrant, uncontrolled immune responses will lead to harmful, or even fatal, consequences. Therefore, it is essential to better discern innate immune signaling and many regulators, controlling various signaling pathways, have been identified. In this review, we focus on the recent advances in our understanding of the activation and regulation of innate immune signaling in the host response to pathogens and cancer. PMID:25625930

  9. MicroRNAs regulate mitochondrial apoptotic pathway in myocardial ischemia-reperfusion-injury.

    PubMed

    Makhdoumi, Pouran; Roohbakhsh, Ali; Karimi, Gholamreza

    2016-12-01

    MicroRNAs (miRNAs) are small non-coding RNAs that act as post-transcriptional gene regulators. They are involved in the pathogenesis of different disorders including heart diseases. MiRNAs contribute to ischemia/reperfusion injury (I/RI) by altering numerous key signaling elements. Together with alterations in the various potential signaling pathways, modification in miRNA expression has been suggested as a part of the response network following ischemia/reperfusion (I/R). In addition, cardiac mitochondrial homeostasis is closely associated with cardiac function and impairment of mitochondrial activity occurred after ischemia/reperfusion injury. MiRNAs play a key role in the regulation of mitochondrial apoptotic pathway and signaling proteins. In this review, we summarize the knowledge currently available regarding the molecular mechanisms of miRNA-regulated mitochondrial functions during ischemia/reperfusion injury. This regulation occurs in different stages of mitochondrial apoptosis pathway.

  10. Protein Kinase D Regulates Cell Death Pathways in Experimental Pancreatitis

    PubMed Central

    Yuan, Jingzhen; Liu, Yannan; Tan, Tanya; Guha, Sushovan; Gukovsky, Ilya; Gukovskaya, Anna; Pandol, Stephen J.

    2012-01-01

    Inflammation and acinar cell necrosis are two major pathological responses of acute pancreatitis, a serious disorder with no current therapies directed to its molecular pathogenesis. Serine/threonine protein kinase D family, which includes PKD/PKD1, PKD2, and PKD3, has been increasingly implicated in the regulation of multiple physiological and pathophysiological effects. We recently reported that PKD/PKD1, the predominant PKD isoform expressed in rat pancreatic acinar cells, mediates early events of pancreatitis including NF-κB activation and inappropriate intracellular digestive enzyme activation. In current studies, we investigated the role and mechanisms of PKD/PKD1 in the regulation of necrosis in pancreatic acinar cells by using two novel small molecule PKD inhibitors CID755673 and CRT0066101 and molecular approaches in in vitro and in vivo experimental models of acute pancreatitis. Our results demonstrated that both CID755673 and CRT0066101 are PKD-specific inhibitors and that PKD/PKD1 inhibition by either the chemical inhibitors or specific PKD/PKD1 siRNAs attenuated necrosis while promoting apoptosis induced by pathological doses of cholecystokinin-octapeptide (CCK) in pancreatic acinar cells. Conversely, up-regulation of PKD expression in pancreatic acinar cells increased necrosis and decreased apoptosis. We further showed that PKD/PKD1 regulated several key cell death signals including inhibitors of apoptotic proteins, caspases, receptor-interacting protein kinase 1 to promote necrosis. PKD/PKD1 inhibition by CID755673 significantly ameliorated necrosis and severity of pancreatitis in an in vivo experimental model of acute pancreatitis. Thus, our studies indicate that PKD/PKD1 is a key mediator of necrosis in acute pancreatitis and that PKD/PKD1 may represent a potential therapeutic target in acute pancreatitis. PMID:22470346

  11. Central genes, pathways and modules that regulate bone mass.

    PubMed

    Quiros-Gonzalez, Isabel; Yadav, Vijay K

    2014-11-01

    Bones are structures that give the shape and defined features to vertebrates, protect several soft organs and perform multiple endocrine influences on other organs. To achieve these functions bones are first modeled early during life and then constantly remodeled throughout life. The process of bone (re)modeling happens simultaneously at multitude of locations in the skeleton and ensures that vertebrates have a mechanically strong yet a flexible skeleton to the most part of their life. Given the extent of its occurrence in the body, bone remodeling is a highly energy demanding process and is co-ordinated with other physiological processes as diverse as energy metabolism, sleep-wake cycle and reproduction. Neuronal circuits in the brain play a very important role in the coordination of bone remodeling with other organ system functions, and perform this function in sync with environmental and peripheral hormonal cues. In this review, we will focus on the roles of hormonal signals and neural circuits that originate in, or impinge on, the brain in the regulation of bone mass. We will provide herein an updated view of how advances in molecular genetics have refined the neural circuits involved in the regulation of bone mass, from the whole brain level to the specific neuronal populations and their neurotransmitters. This will help to understand the mechanisms whereby vertebrate brain regulates bone mass by fine-tuning metabolic signals that originate in the brain or elsewhere in the body.

  12. Brain areas and pathways in the regulation of glucose metabolism.

    PubMed

    Diepenbroek, Charlene; Serlie, Mireille J; Fliers, Eric; Kalsbeek, Andries; la Fleur, Susanne E

    2013-01-01

    Glucose is the most important source of fuel for the brain and its concentration must be kept within strict boundaries to ensure the organism's optimal fitness. To maintain glucose homeostasis, an optimal balance between glucose uptake and glucose output is required. Besides managing acute changes in plasma glucose concentrations, the brain controls a daily rhythm in glucose concentrations. The various nuclei within the hypothalamus that are involved in the control of both these processes are well known. However, novel studies indicate an additional role for brain areas that are originally appreciated in other processes than glucose metabolism. Therefore, besides the classic hypothalamic pathways, we will review cortico-limbic brain areas and their role in glucose metabolism.

  13. Complex regulation of HSC emergence by the Notch signaling pathway

    PubMed Central

    Butko, Emerald; Pouget, Claire; Traver, David

    2016-01-01

    Hematopoietic stem cells are formed during embryonic development, and serve as the foundation of the definitive blood program for life. Notch signaling has been well established as an essential direct contributor to HSC specification. However, several recent studies have indicated that the contribution of Notch signaling is complex. HSC specification requires multiple Notch signaling inputs, some received directly by hematopoietic precursors, and others that occur indirectly within neighboring somites. Of note, proinflammatory signals provided by primitive myeloid cells are needed for HSC specification via upregulation of the Notch pathway in hemogenic endothelium. In addition to multiple requirements for Notch activation, recent studies indicate that Notch signaling must subsequently be repressed to permit HSC emergence. Finally, Notch must then be reactivated to maintain HSC fate. In this review, we discuss the growing understanding of the dynamic contributions of Notch signaling to the establishment of hematopoiesis during development. PMID:26586199

  14. Interacting appetite-regulating pathways in the hypothalamic regulation of body weight.

    PubMed

    Kalra, S P; Dube, M G; Pu, S; Xu, B; Horvath, T L; Kalra, P S

    1999-02-01

    elucidating the precise role of leptin in daily patterning of feeding in the rat. 4) A neural timing mechanism also operates upstream from the ARN in the daily management of energy homeostasis. Although the precise anatomical boundaries are not clearly defined, this device is likely to be composed of a group of neurons that integrate incoming internal and external information for the timely onset of the drive to eat. Evidently, this network operates independently in primates, but it is entrained to the circadian time keeper in the SCN of rodents. Apart from its role in the onset of drive to eat, the circadian patterns of gene expression of NPY, GAL, and POMC denote independent control of the timing device on the synthesis and availability for release of orexigenic signals. The VMN-DMN-PVN complex is apparently an integrated constituent of the timing mechanism in this context, because lesions in each of these sites result in loss of regulated feeding. The accumulated evidence points to the PVN and surrounding neural sites within this framework as the primary sites of release and action of various orexigenic and anorexigenic signals. A novel finding is the identification of the interconnected wiring of the DMN-mPVN axis that may mediate leptin restraint on NPY-induced feeding. The chemical phenotypes of leptin and NPY target neurons in this axis remain to be identified. These multiple orexigenic and anorexigenic pathways in the hypothalamic ARN appear to represent redundancy, a characteristic of regulated biological systems to provide a "fail-safe" neural mechanism to meet an organism's constant energy needs for growth and maintenance. Within this formulation, the coexisting orexigenic signals (NPY, NE, GAL, GABA, and AgrP) represent either another level of redundancy or it is possible that these signals operate within the ARN as reinforcing agents to varying degrees under different circumstances. (ABSTRACT TRUNCATED)

  15. A sugar phosphatase regulates the methylerythritol phosphate (MEP) pathway in malaria parasites.

    PubMed

    Guggisberg, Ann M; Park, Jooyoung; Edwards, Rachel L; Kelly, Megan L; Hodge, Dana M; Tolia, Niraj H; Odom, Audrey R

    2014-07-24

    Isoprenoid biosynthesis through the methylerythritol phosphate (MEP) pathway generates commercially important products and is a target for antimicrobial drug development. MEP pathway regulation is poorly understood in microorganisms. Here we employ a forward genetics approach to understand MEP pathway regulation in the malaria parasite, Plasmodium falciparum. The antimalarial fosmidomycin inhibits the MEP pathway enzyme deoxyxylulose 5-phosphate reductoisomerase (DXR). Fosmidomycin-resistant P. falciparum are enriched for changes in the PF3D7_1033400 locus (hereafter referred to as PfHAD1), encoding a homologue of haloacid dehalogenase (HAD)-like sugar phosphatases. We describe the structural basis for loss-of-function PfHAD1 alleles and find that PfHAD1 dephosphorylates a variety of sugar phosphates, including glycolytic intermediates. Loss of PfHAD1 is required for fosmidomycin resistance. Parasites lacking PfHAD1 have increased MEP pathway metabolites, particularly the DXR substrate, deoxyxylulose 5-phosphate. PfHAD1 therefore controls substrate availability to the MEP pathway. Because PfHAD1 has homologues in plants and bacteria, other HAD proteins may be MEP pathway regulators.

  16. Cooperation of multiple signaling pathways in CD40-regulated gene expression in B lymphocytes

    PubMed Central

    Dadgostar, Hajir; Zarnegar, Brian; Hoffmann, Alexander; Qin, Xiao-Feng; Truong, Uyen; Rao, Govinda; Baltimore, David; Cheng, Genhong

    2002-01-01

    CD40/CD40L interaction is essential for multiple biological events in T dependent humoral immune responses, including B cell survival and proliferation, germinal center and memory B cell formation, and antibody isotype switching and affinity maturation. By using high-density microarrays, we examined gene expression in primary mouse B lymphocytes after multiple time points of CD40L stimulation. In addition to genes involved in cell survival and growth, which are also induced by other mitogens such as lipopolysaccharide, CD40L specifically activated genes involved in germinal center formation and T cell costimulatory molecules that facilitate T dependent humoral immunity. Next, by examining the roles of individual CD40-activated signal transduction pathways, we dissected the overall CD40-mediated response into genes independently regulated by the individual pathways or collectively by all pathways. We also found that gene down-regulation is a significant part of the overall response and that the p38 pathway plays an important role in this process, whereas the NF-κB pathway is important for the up-regulation of primary response genes. Our finding of overlapping independent control of gene expression modules by different pathways suggests, in principle, that distinct biological behaviors that depend on distinct gene expression subsets can be manipulated by targeting specific signaling pathways. PMID:11830667

  17. Carbohydrate Metabolism in Archaea: Current Insights into Unusual Enzymes and Pathways and Their Regulation

    PubMed Central

    Esser, Dominik; Rauch, Bernadette

    2014-01-01

    SUMMARY The metabolism of Archaea, the third domain of life, resembles in its complexity those of Bacteria and lower Eukarya. However, this metabolic complexity in Archaea is accompanied by the absence of many “classical” pathways, particularly in central carbohydrate metabolism. Instead, Archaea are characterized by the presence of unique, modified variants of classical pathways such as the Embden-Meyerhof-Parnas (EMP) pathway and the Entner-Doudoroff (ED) pathway. The pentose phosphate pathway is only partly present (if at all), and pentose degradation also significantly differs from that known for bacterial model organisms. These modifications are accompanied by the invention of “new,” unusual enzymes which cause fundamental consequences for the underlying regulatory principles, and classical allosteric regulation sites well established in Bacteria and Eukarya are lost. The aim of this review is to present the current understanding of central carbohydrate metabolic pathways and their regulation in Archaea. In order to give an overview of their complexity, pathway modifications are discussed with respect to unusual archaeal biocatalysts, their structural and mechanistic characteristics, and their regulatory properties in comparison to their classic counterparts from Bacteria and Eukarya. Furthermore, an overview focusing on hexose metabolic, i.e., glycolytic as well as gluconeogenic, pathways identified in archaeal model organisms is given. Their energy gain is discussed, and new insights into different levels of regulation that have been observed so far, including the transcript and protein levels (e.g., gene regulation, known transcription regulators, and posttranslational modification via reversible protein phosphorylation), are presented. PMID:24600042

  18. Regulation of PCP by the Fat signaling pathway

    PubMed Central

    Matis, Maja; Axelrod, Jeffrey D.

    2013-01-01

    Planar cell polarity (PCP) in epithelia, orthogonal to the apical–basal axis, is essential for numerous developmental events and physiological functions. Drosophila model systems have been at the forefront of studies revealing insights into mechanisms regulating PCP and have revealed distinct signaling modules. One of these, involving the atypical cadherins Fat and Dachsous and the ectokinase Four-jointed, appears to link the direction of cell polarization to the tissue axes. We discuss models for the function of this signaling module as well as several unanswered questions that may guide future investigations. PMID:24142873

  19. Overlapping and interactive pathways regulating appetite and craving.

    PubMed

    Kalra, Satya P; Kalra, Pushpa S

    2004-01-01

    Multidisciplinary research in recent years has delineated the hypothalamic hardcore wiring that encodes appetitive drive. The appetite regulating network (ARN) consisting of distinct orexigenic and anorexigenic circuitries operates in the arcuate nucleus-paraventricular nucleus axis of the hypothalamus to propagate and relay the appetitive drive, and is subject to modulation by excitatory and inhibitory messages from the lateral hypothalamus and ventromedial nucleus, respectively. Reciprocal afferent humoral signals, comprised of anorexigenic leptin from white adipose tissue and orexigenic ghrelin from stomach, to the ARN integrate the moment-to-moment regulation of energy homeostasis. Various loci in the ARN and afferent hormonal feedback circuitry in the rodent brain are important for food craving elicited by drugs of abuse. This convergence of neurochemical and hormonal signaling has now paved the way to address the fundamental question of whether cellular and molecular events that underlie the appetitive drive in response to diminished energy stores in the body are akin to drug craving during withdrawal in humans.

  20. Pathway and regulation of erythritol formation in Leuconostoc oenos.

    PubMed Central

    Veiga-da-Cunha, M; Santos, H; Van Schaftingen, E

    1993-01-01

    It was recently observed that Leuconostoc oenos GM, a wine lactic acid bacterium, produced erythritol anaerobically from glucose but not from fructose or ribose and that this production was almost absent in the presence of O2. In this study, the pathway of formation of erythritol from glucose in L. oenos was shown to involve the isomerization of glucose 6-phosphate to fructose 6-phosphate by a phosphoglucose isomerase, the cleavage of fructose 6-phosphate by a phosphoketolase, the reduction of erythrose 4-phosphate by an erythritol 4-phosphate dehydrogenase and, finally, the hydrolysis of erythritol 4-phosphate to erythritol by a phosphatase. Fructose 6-phosphate phosphoketolase was copurified with xylulose 5-phosphate phosphoketolase, and the activity of the latter was competitively inhibited by fructose 6-phosphate, with a Ki of 26 mM, corresponding to the Km of fructose 6-phosphate phosphoketolase (22 mM). These results suggest that the two phosphoketolase activities are borne by a single enzyme. Extracts of L. oenos were also found to contain NAD(P)H oxidase, which must be largely responsible for the reoxidation of NADPH and NADH in cells incubated in the presence of O2. In cells incubated with glucose, the concentrations of glucose 6-phosphate and of fructose 6-phosphate were higher in the absence of O2 than in its presence, explaining the stimulation by anaerobiosis of erythritol production. The increase in the hexose 6-phosphate concentration is presumably the result of a functional inhibition of glucose 6-phosphate dehydrogenase because of a reduction in the availability of NADP. PMID:8391532

  1. Regulation of Intrinsic and Extrinsic Apoptotic Pathways in Osteosarcoma Cells Following Oleandrin Treatment

    PubMed Central

    Ma, Yunlong; Zhu, Bin; Yong, Lei; Song, Chunyu; Liu, Xiao; Yu, Huilei; Wang, Peng; Liu, Zhongjun; Liu, Xiaoguang

    2016-01-01

    Our previous study has reported the anti-tumor effect of oleandrin on osteosarcoma (OS) cells. In the current study, we mainly explored its potential regulation on intrinsic and extrinsic apoptotic pathway in OS cells. Cells apoptosis, reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were detected using fluorescence staining and flow cytometry. Caspase-3 activity was detected using a commercial kit. The levels of cytoplasmic cytochrome c, mitochondrial cytochrome c, bcl-2, bax, caspase-9, Fas, FasL, caspase-8 and caspase-3 were detected by Western blotting. z-VAD-fmk was applied to block both intrinsic and extrinsic apoptosis pathways, and cells apoptosis was also tested. Furthermore, we used z-LEHD-fmk and Fas blocking antibody to inhibit intrinsic and extrinsic pathways, separately, and the selectivity of oleandrin on these pathways was explored. Results showed that oleandrin induced the apoptosis of OS cells, which was accompanied by an increase in ROS and a decrease in MMP. Furthermore, cytochrome c level was reduced in mitochondria but elevated in the cytoplasm. Caspase-3 activity was enhanced by oleandrin in a concentration- and time-dependent manner. Oleandrin also down-regulated the expression of bcl-2, but up-regulated bax, caspase-9, Fas, FasL, caspase-8 and caspase-3. In addition, the suppression of both apoptotic pathways by z-VAD-fmk greatly reverted the oleandrin-induced apoptosis. Moreover, the suppression of one pathway by a corresponding inhibitor did not affect the regulation of oleandrin on another pathway. Taken together, we concluded that oleandrin induced apoptosis of OS cells via activating both intrinsic and extrinsic apoptotic pathways. PMID:27886059

  2. Integrated pathway-based transcription regulation network mining and visualization based on gene expression profiles.

    PubMed

    Kibinge, Nelson; Ono, Naoaki; Horie, Masafumi; Sato, Tetsuo; Sugiura, Tadao; Altaf-Ul-Amin, Md; Saito, Akira; Kanaya, Shigehiko

    2016-06-01

    Conventionally, workflows examining transcription regulation networks from gene expression data involve distinct analytical steps. There is a need for pipelines that unify data mining and inference deduction into a singular framework to enhance interpretation and hypotheses generation. We propose a workflow that merges network construction with gene expression data mining focusing on regulation processes in the context of transcription factor driven gene regulation. The pipeline implements pathway-based modularization of expression profiles into functional units to improve biological interpretation. The integrated workflow was implemented as a web application software (TransReguloNet) with functions that enable pathway visualization and comparison of transcription factor activity between sample conditions defined in the experimental design. The pipeline merges differential expression, network construction, pathway-based abstraction, clustering and visualization. The framework was applied in analysis of actual expression datasets related to lung, breast and prostrate cancer.

  3. Adaptive Control Model Reveals Systematic Feedback and Key Molecules in Metabolic Pathway Regulation

    PubMed Central

    Moffitt, Richard A.; Merrill, Alfred H.; Wang, May D.

    2011-01-01

    Abstract Robust behavior in metabolic pathways resembles stabilized performance in systems under autonomous control. This suggests we can apply control theory to study existing regulation in these cellular networks. Here, we use model-reference adaptive control (MRAC) to investigate the dynamics of de novo sphingolipid synthesis regulation in a combined theoretical and experimental case study. The effects of serine palmitoyltransferase over-expression on this pathway are studied in vitro using human embryonic kidney cells. We report two key results from comparing numerical simulations with observed data. First, MRAC simulations of pathway dynamics are comparable to simulations from a standard model using mass action kinetics. The root-sum-square (RSS) between data and simulations in both cases differ by less than 5%. Second, MRAC simulations suggest systematic pathway regulation in terms of adaptive feedback from individual molecules. In response to increased metabolite levels available for de novo sphingolipid synthesis, feedback from molecules along the main artery of the pathway is regulated more frequently and with greater amplitude than from other molecules along the branches. These biological insights are consistent with current knowledge while being new that they may guide future research in sphingolipid biology. In summary, we report a novel approach to study regulation in cellular networks by applying control theory in the context of robust metabolic pathways. We do this to uncover potential insight into the dynamics of regulation and the reverse engineering of cellular networks for systems biology. This new modeling approach and the implementation routines designed for this case study may be extended to other systems. Supplementary Material is available at www.liebertonline.com/cmb. PMID:21314456

  4. Quercitrin attenuates osteoporosis in ovariectomized rats by regulating mitogen-activated protein kinase (MAPK) signaling pathways.

    PubMed

    Xing, Li-Zhi; Ni, Huai-Jun; Wang, Yu-Ling

    2017-03-13

    MAPK signaling pathways are crucial in regulating osteogenesis, a genetic disorder affecting the bones. Quercitrin, a type of flavonoid, is widely distributed in nature and involved in many pharmacological activities. But its osteoprotective functions and mechanism in osteoporosis are far from being understood clearly. In this paper, the MAPK upregulation was observed in the ovariectomy-induced bone loss. Quercitrin was found to downregulate MAPK signaling pathways and prevent the ovariectomy-induced deterioration of bone mineral density (BMD), trabecular microstructure, and bone mechanical characteristics. In this study, quercitrin was seen to prevent the progression of the postmenopausal osteoporosis among the rats, which may be mediated by the downregulated MAPK signaling pathways.

  5. Regulation of respiration in plants: a role for alternative metabolic pathways.

    PubMed

    van Dongen, Joost T; Gupta, Kapuganti J; Ramírez-Aguilar, Santiago J; Araújo, Wagner L; Nunes-Nesi, Adriano; Fernie, Alisdair R

    2011-08-15

    Respiratory metabolism includes the reactions of glycolysis, the tricarboxylic acid cycle and the mitochondrial electron transport chain, but is also directly linked with many other metabolic pathways such as protein and lipid biosynthesis and photosynthesis via photorespiration. Furthermore, any change in respiratory activity can impact the redox status of the cell and the production of reactive oxygen species. In this review, it is discussed how respiration is regulated and what alternative pathways are known that increase the metabolic flexibility of this vital metabolic process. By looking at the adaptive responses of respiration to hypoxia or changes in the oxygen availability of a cell, the integration of regulatory responses of various pathways is illustrated.

  6. Role of bile acids in the regulation of the metabolic pathways

    PubMed Central

    Taoka, Hiroki; Yokoyama, Yoko; Morimoto, Kohkichi; Kitamura, Naho; Tanigaki, Tatsuya; Takashina, Yoko; Tsubota, Kazuo; Watanabe, Mitsuhiro

    2016-01-01

    Recent studies have revealed that bile acids (BAs) are not only facilitators of dietary lipid absorption but also important signaling molecules exerting multiple physiological functions. Some major signaling pathways involving the nuclear BAs receptor farnesoid X receptor and the G protein-coupled BAs receptor TGR5/M-BAR have been identified to be the targets of BAs. BAs regulate their own homeostasis via signaling pathways. BAs also affect diverse metabolic pathways including glucose metabolism, lipid metabolism and energy expenditure. This paper suggests the mechanism of controlling metabolism via BA signaling and demonstrates that BA signaling is an attractive therapeutic target of the metabolic syndrome. PMID:27433295

  7. Transcriptomic analysis in the developing zebrafish embryo after compound exposure: individual gene expression and pathway regulation.

    PubMed

    Hermsen, Sanne A B; Pronk, Tessa E; van den Brandhof, Evert-Jan; van der Ven, Leo T M; Piersma, Aldert H

    2013-10-01

    The zebrafish embryotoxicity test is a promising alternative assay for developmental toxicity. Classically, morphological assessment of the embryos is applied to evaluate the effects of compound exposure. However, by applying differential gene expression analysis the sensitivity and predictability of the test may be increased. For defining gene expression signatures of developmental toxicity, we explored the possibility of using gene expression signatures of compound exposures based on commonly expressed individual genes as well as based on regulated gene pathways. Four developmental toxic compounds were tested in concentration-response design, caffeine, carbamazepine, retinoic acid and valproic acid, and two non-embryotoxic compounds, d-mannitol and saccharin, were included. With transcriptomic analyses we were able to identify commonly expressed genes, which were mostly development related, after exposure to the embryotoxicants. We also identified gene pathways regulated by the embryotoxicants, suggestive of their modes of action. Furthermore, whereas pathways may be regulated by all compounds, individual gene expression within these pathways can differ for each compound. Overall, the present study suggests that the use of individual gene expression signatures as well as pathway regulation may be useful starting points for defining gene biomarkers for predicting embryotoxicity.

  8. Role of calcium in regulation of phosphoinositide signaling pathway.

    PubMed

    Patel, J; Keith, R A; Salama, A I; Moore, W C

    1991-01-01

    Using primary neuronal cultures we have examined the role of extracellular Ca2+ in a receptor-regulated phosphoinositide turnover. We report that receptor (glutamic acid and acetylcholine)-activated phosphoinositide turnover requires the presence of extracellular Ca2+ (EC50 = 21.1 microM). The requirement for Ca2+ appears to be at an intracellular level and is highly selective for Ca2+. We also found that several inorganic and organic Ca2+ channel blockers, including La3+ and verapamil, inhibit phosphoinositide turnover. However, the pharmacological profile of these agents in this regard was distinct from their actions at the voltage-sensitive Ca2+ channels. To explain the above requirement for extracellular Ca2+ in agonist-stimulated phosphoinositide turnover and its sensitivity to Ca(2+)-channel blockers, we propose a hypothetical model suggesting that Ca2+, following IP-3-mediated mobilization, exerts a facilitatory action on the activity of receptor-phospholipase C complex. We further propose that in the absence of extracellular Ca2+ or in the presence of certain Ca(2+)-channel blockers, refilling of calciosomes is ineffectual or inhibited, causing its depletion and subsequent inactivation of agonist-stimulated phosphoinositide turnover.

  9. Dissecting dysfunctional crosstalk pathways regulated by miRNAs during glioma progression

    PubMed Central

    Li, Feng; Li, Xiang; Feng, Li; Shi, Xinrui; Wang, Lihua; Li, Xia

    2016-01-01

    Glioma is a malignant nervous system tumor with a high fatality rate and poor prognosis. MicroRNAs (miRNAs) are important post-transcriptional modulators of glioma initiation and progression. Tumor progression often results from dysfunctional co-operation between pathways regulated by miRNAs. We therefore constructed a glioma progression-related miRNA-pathway crosstalk network that not only revealed some key miRNA-pathway patterns, but also helped characterize the functional roles of miRNAs during glioma progression. Our data indicate that crosstalk between cell cycle and p53 pathways is associated with grade II to grade III progression, while cell communications-related pathways involving regulation of actin cytoskeleton and adherens junctions are associated with grade IV glioblastoma progression. Furthermore, miRNAs and their crosstalk pathways may be useful for stratifying glioma and glioblastoma patients into groups with short or long survival times. Our data indicate that a combination of miRNA and pathway crosstalk information can be used for survival prediction. PMID:27013589

  10. The flavonoid pathway regulates the petal colors of cotton flower.

    PubMed

    Tan, Jiafu; Wang, Maojun; Tu, Lili; Nie, Yichun; Lin, Yongjun; Zhang, Xianlong

    2013-01-01

    Although biochemists and geneticists have studied the cotton flower for more than one century, little is known about the molecular mechanisms underlying the dramatic color change that occurs during its short developmental life following blooming. Through the analysis of world cotton germplasms, we found that all of the flowers underwent color changes post-anthesis, but there is a diverse array of petal colors among cotton species, with cream, yellow and red colors dominating the color scheme. Genetic and biochemical analyses indicated that both the original cream and red colors and the color changes post-anthesis were related to flavonoid content. The anthocyanin content and the expression of biosynthesis genes were both increased from blooming to one day post-anthesis (DPA) when the flower was withering and undergoing abscission. Our results indicated that the color changes and flavonoid biosynthesis of cotton flowers were precisely controlled and genetically regulated. In addition, flavonol synthase (FLS) genes involved in flavonol biosynthesis showed specific expression at 11 am when the flowers were fully opened. The anthocyanidin reductase (ANR) genes, which are responsible for proanthocyanidins biosynthesis, showed the highest expression at 6 pm on 0 DPA, when the flowers were withered. Light showed primary, moderate and little effects on flavonol, anthocyanin and proanthocyanidin biosynthesis, respectively. Flavonol biosynthesis was in response to light exposure, while anthocyanin biosynthesis was involved in flower color changes. Further expression analysis of flavonoid genes in flowers of wild type and a flavanone 3-hydroxylase (F3H) silenced line showed that the development of cotton flower color was controlled by a complex interaction between genes and light. These results present novel information regarding flavonoids metabolism and flower development.

  11. ERK signaling pathway regulates sleep duration through activity-induced gene expression during wakefulness.

    PubMed

    Mikhail, Cyril; Vaucher, Angélique; Jimenez, Sonia; Tafti, Mehdi

    2017-01-24

    Wakefulness is accompanied by experience-dependent synaptic plasticity and an increase in activity-regulated gene transcription. Wake-induced genes are certainly markers of neuronal activity and may also directly regulate the duration of and need for sleep. We stimulated murine cortical cultures with the neuromodulatory signals that are known to control wakefulness in the brain and found that norepinephrine alone or a mixture of these neuromodulators induced activity-regulated gene transcription. Pharmacological inhibition of the various signaling pathways involved in the regulation of gene expression indicated that the extracellular signal-regulated kinase (ERK) pathway is the principal one mediating the effects of waking neuromodulators on gene expression. In mice, ERK phosphorylation in the cortex increased and decreased with wakefulness and sleep. Whole-body or cortical neuron-specific deletion of Erk1 or Erk2 significantly increased the duration of wakefulness in mice, and pharmacological inhibition of ERK phosphorylation decreased sleep duration and increased the duration of wakefulness bouts. Thus, this signaling pathway, which is highly conserved from Drosophila to mammals, is a key pathway that links waking experience-induced neuronal gene expression to sleep duration and quality.

  12. Tbx2 regulates anterior neural specification by repressing FGF signaling pathway.

    PubMed

    Cho, Gun-Sik; Park, Dong-Seok; Choi, Sun-Cheol; Han, Jin-Kwan

    2017-01-15

    During early embryogenesis, FGF signals regulate the antero-posterior (AP) patterning of the neural plate by promoting posterior cell fates. In particular, BMP signal-mediated attenuation of FGF pathway plays a critical role in the determination of the anterior neural region. Here we show that Tbx2, a T-box transcriptional repressor regulates anterior neural specification by suppressing FGF8 signaling pathway in Xenopus embryo. Tbx2 is expressed in the anterior edge of the neural plate in early neurulae. Overexpression and knockdown of Tbx2 induce expansion and reduction in the expression of anterior neural markers, respectively. It also suppresses FGF8-induced ERK phosphorylation and neural caudalization. Tbx2, which is a target gene of BMP signal, down-regulates FGF8 signaling by inhibiting the expression of Flrt3, a positive regulator of this pathway. We found that Tbx2 binds directly to the T-box element located in the promoter region of Flrt3 gene, thereby interfering with the activity of the promoter. Consistently, Tbx2 augmentation of anterior neural formation is inhibited by co-expression of Flrt3. Furthermore, disruption of the anterior-most structures such as eyes in Tbx2-depleted embryos can be rescued by inhibition of Flrt3 function or FGF signaling. Taken together, our results suggest that Tbx2 mediates BMP signal to down-regulate FGF signaling pathway by repressing Flrt3 expression for anterior tissue formation.

  13. Interspecies Complementation of the LuxR Family Pathway-Specific Regulator Involved in Macrolide Biosynthesis.

    PubMed

    Mo, SangJoon; Yoon, Yeo Joon

    2016-01-01

    PikD is a widely known pathway-specific regulator for controlling pikromycin production in Streptomyces venezuelae ATCC 15439, which is a representative of the large ATP-binding regulator of the LuxR family (LAL) in Streptomyces sp. RapH and FkbN also belong to the LAL family of transcriptional regulators, which show greatest homology with the ATP-binding motif and helix-turn-helix DNA-binding motif of PikD. Overexpression of pikD and heterologous expression of rapH and fkbN led to enhanced production of pikromycin by approximately 1.8-, 1.6-, and 1.6-fold in S. venezuelae, respectively. Cross-complementation of rapH and fkbN in the pikD deletion mutant (ΔpikD) restored pikromycin and derived macrolactone production. Overall, these results show that heterologous expression of rapH and fkbN leads to the overproduction of pikromycin and its congeners from the pikromycin biosynthetic pathway in S. venezuelae, and they have the same functionality as the pathwayspecific transcriptional activator for the pikromycin biosynthetic pathway in the ΔpikD strain. These results also show extensive "cross-communication" between pathway-specific regulators of streptomycetes and suggest revision of the current paradigm for pathwayspecific versus global regulation of secondary metabolism in Streptomyces species.

  14. Homeostatic response to hypoxia is regulated by the N-end rule pathway in plants.

    PubMed

    Gibbs, Daniel J; Lee, Seung Cho; Isa, Nurulhikma Md; Gramuglia, Silvia; Fukao, Takeshi; Bassel, George W; Correia, Cristina Sousa; Corbineau, Françoise; Theodoulou, Frederica L; Bailey-Serres, Julia; Holdsworth, Michael J

    2011-10-23

    Plants and animals are obligate aerobes, requiring oxygen for mitochondrial respiration and energy production. In plants, an unanticipated decline in oxygen availability (hypoxia), as caused by roots becoming waterlogged or foliage submergence, triggers changes in gene transcription and messenger RNA translation that promote anaerobic metabolism and thus sustain substrate-level ATP production. In contrast to animals, oxygen sensing has not been ascribed to a mechanism of gene regulation in response to oxygen deprivation in plants. Here we show that the N-end rule pathway of targeted proteolysis acts as a homeostatic sensor of severe low oxygen levels in Arabidopsis, through its regulation of key hypoxia-response transcription factors. We found that plants lacking components of the N-end rule pathway constitutively express core hypoxia-response genes and are more tolerant of hypoxic stress. We identify the hypoxia-associated ethylene response factor group VII transcription factors of Arabidopsis as substrates of this pathway. Regulation of these proteins by the N-end rule pathway occurs through a characteristic conserved motif at the amino terminus initiating with Met-Cys. Enhanced stability of one of these proteins, HRE2, under low oxygen conditions improves hypoxia survival and reveals a molecular mechanism for oxygen sensing in plants via the evolutionarily conserved N-end rule pathway. SUB1A-1, a major determinant of submergence tolerance in rice, was shown not to be a substrate for the N-end rule pathway despite containing the N-terminal motif, indicating that it is uncoupled from N-end rule pathway regulation, and that enhanced stability may relate to the superior tolerance of Sub1 rice varieties to multiple abiotic stresses.

  15. Skin sensitizers differentially regulate signaling pathways in MUTZ-3 cells in relation to their individual potency

    PubMed Central

    2014-01-01

    Background Due to the recent European legislations posing a ban of animal tests for safety assessment within the cosmetic industry, development of in vitro alternatives for assessment of skin sensitization is highly prioritized. To date, proposed in vitro assays are mainly based on single biomarkers, which so far have not been able to classify and stratify chemicals into subgroups, related to risk or potency. Methods Recently, we presented the Genomic Allergen Rapid Detection (GARD) assay for assessment of chemical sensitizers. In this paper, we show how the genome wide readout of GARD can be expanded and used to identify differentially regulated pathways relating to individual chemical sensitizers. In this study, we investigated the mechanisms of action of a range of skin sensitizers through pathway identification, pathway classification and transcription factor analysis and related this to the reactive mechanisms and potency of the sensitizing agents. Results By transcriptional profiling of chemically stimulated MUTZ-3 cells, 33 canonical pathways intimately involved in sensitization to chemical substances were identified. The results showed that metabolic processes, cell cycling and oxidative stress responses are the key events activated during skin sensitization, and that these functions are engaged differently depending on the reactivity mechanisms of the sensitizing agent. Furthermore, the results indicate that the chemical reactivity groups seem to gradually engage more pathways and more molecules in each pathway with increasing sensitizing potency of the chemical used for stimulation. Also, a switch in gene regulation from up to down regulation, with increasing potency, was seen both in genes involved in metabolic functions and cell cycling. These observed pathway patterns were clearly reflected in the regulatory elements identified to drive these processes, where 33 regulatory elements have been proposed for further analysis. Conclusions This study

  16. Protective effects of pogostone against LPS-induced acute lung injury in mice via regulation of Keap1-Nrf2/NF-κB signaling pathways.

    PubMed

    Sun, Chao-Yue; Xu, Lie-Qiang; Zhang, Zhen-Biao; Chen, Chao-Hui; Huang, Yong-Zhong; Su, Zu-Qing; Guo, Hui-Zhen; Chen, Xiao-Ying; Zhang, Xie; Liu, Yu-Hong; Chen, Jian-Nan; Lai, Xiao-Ping; Li, Yu-Cui; Su, Zi-Ren

    2016-03-01

    Pogostone, a major component of Pogostemon cablin, has been demonstrated to possess antibacterial, anti-fungal, immunosuppressive and anti-inflammatory properties. To investigate the potential therapeutic effect of pogostone on lipopolysaccharide (LPS)-induced acute lung injury (ALI), mice were pretreated with pogostone prior to LPS exposure. After LPS challenge, the lungs were excised and the histological changes, wet to dry weight ratios, MPO activity reflecting neutrophil infiltration, and MDA activity reflecting oxidative stress were examined. The inflammatory cytokines in the BALF were determined by ELISA assay. Moreover, the expressions of p65 and phosphorylated p65 subunit of NF-κB, and Nrf2 in the nucleus in lung tissues were measured by Western blot analysis, and meanwhile the dependent genes of NF-κB and Nrf2 were assessed by RT-qPCR. The results showed that pretreatment with pogostone markedly improved survival rate, attenuated the histological alterations in the lung, reduced the MPO and MDA levels, decreased the wet/dry weight ratio of lungs, down-regulated the level of pro-inflammatory mediators including TNF-a, IL-1β and IL-6. Furthermore, pretreatment with pogostone enhanced the Nrf2 dependent genes including NQO-1, GCLC and HO-1 but suppressed NF-κB regulated genes including TNF-α, IL-1β and IL-6. The mechanism behind the protective effect was correlated with its regulation on the balance between Keap1-Nrf2 and NF-κB signaling pathways. Therefore, pogostone may be considered as a potential therapeutic agent for preventing and treating ALI.

  17. Claudin-7 indirectly regulates the integrin/FAK signaling pathway in human colon cancer tissue.

    PubMed

    Ding, Lei; Wang, Liyong; Sui, Leiming; Zhao, Huanying; Xu, Xiaoxue; Li, Tengyan; Wang, Xiaonan; Li, Wenjing; Zhou, Ping; Kong, Lu

    2016-08-01

    The claudin family of proteins is integral to the structure and function of tight junctions. The role of claudin-7 (Cldn-7, CLDN7) in regulating the integrin/focal adhesion kinase (FAK)/ERK signaling pathway remains poorly understood. Therefore, we investigated differences in gene expression, primarily focusing on CLDN7 and integrin/FAK/ERK signaling pathway genes, between colon cancer and adjacent normal tissues. Quantitative real-time reverse transcription-PCR and immunohistochemistry were utilized to verify the results of mRNA and protein expression, respectively. In silico analysis was used to predict co-regulation between Cldn-7 and integrin/FAK/ERK signaling pathway components, and the STRING database was used to analyze protein-protein interaction pairs among these proteins. Meta-analysis of expression microarrays in The Cancer Genome Atlas (TCGA) database was used to identify significant correlations between Cldn-7 and components of predicted genes in the integrin/FAK/ERK signaling pathway. Our results showed marked cancer stage-specific decreases in the protein expression of Cldn-7, Gelsolin, MAPK1 and MAPK3 in colon cancer samples, and the observed changes for all proteins except Cldn-7 were in agreement with changes in the corresponding mRNA levels. Cldn-7 might indirectly regulate MAPK3 via KRT8 due to KRT8 co-expression with MAPK3 or CLDN7. Our bioinformatics methods supported the hypothesis that Cldn-7 does not directly regulate any genes in the integrin/FAK/ERK signaling pathway. These factors may participate in a common network that regulates cancer progression in which the MAPK pathway serves as the central node.

  18. A constitutive active MAPK/ERK pathway due to BRAFV600E positively regulates AHR pathway in PTC

    PubMed Central

    Regazzo, Daniela; Bertazza, Loris; Galuppini, Francesca; Guzzardo, Vincenza; Jaffrain-Rea, Marie Lise; Vianello, Federica; Ciato, Denis; Ceccato, Filippo; Watutantrige-Fernando, Sara; Bisognin, Andrea; Bortoluzzi, Stefania; Pennelli, Gianmaria; Boscaro, Marco; Scaroni, Carla; Mian, Caterina

    2015-01-01

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor mediating the toxicity and tumor-promoting properties of dioxin. AHR has been reported to be overexpressed and constitutively active in a variety of solid tumors, but few data are currently available concerning its role in thyroid cancer. In this study we quantitatively explored a series of 51 paired-normal and papillary thyroid carcinoma (PTC) tissues for AHR-related genes. We identified an increased AHR expression/activity in PTC, independently from its nuclear dimerization partner and repressor but strictly related to a constitutive active MAPK/ERK pathway. The AHR up-regulation followed by an increased expression of AHR target genes was confirmed by a meta-analysis of published microarray data, suggesting a ligand-independent active AHR pathway in PTC. In-vitro studies using a PTC-derived cell line (BCPAP) and HEK293 cells showed that BRAFV600E may directly modulate AHR localization, induce AHR expression and activity in an exogenous ligand-independent manner. The AHR pathway might represent a potential novel therapeutic target for PTC in the clinical practice. PMID:26392334

  19. A constitutive active MAPK/ERK pathway due to BRAFV600E positively regulates AHR pathway in PTC.

    PubMed

    Occhi, Gianluca; Barollo, Susi; Regazzo, Daniela; Bertazza, Loris; Galuppini, Francesca; Guzzardo, Vincenza; Jaffrain-Rea, Marie Lise; Vianello, Federica; Ciato, Denis; Ceccato, Filippo; Watutantrige-Fernando, Sara; Bisognin, Andrea; Bortoluzzi, Stefania; Pennelli, Gianmaria; Boscaro, Marco; Scaroni, Carla; Mian, Caterina

    2015-10-13

    The aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor mediating the toxicity and tumor-promoting properties of dioxin. AHR has been reported to be overexpressed and constitutively active in a variety of solid tumors, but few data are currently available concerning its role in thyroid cancer. In this study we quantitatively explored a series of 51 paired-normal and papillary thyroid carcinoma (PTC) tissues for AHR-related genes. We identified an increased AHR expression/activity in PTC, independently from its nuclear dimerization partner and repressor but strictly related to a constitutive active MAPK/ERK pathway. The AHR up-regulation followed by an increased expression of AHR target genes was confirmed by a meta-analysis of published microarray data, suggesting a ligand-independent active AHR pathway in PTC. In-vitro studies using a PTC-derived cell line (BCPAP) and HEK293 cells showed that BRAFV600E may directly modulate AHR localization, induce AHR expression and activity in an exogenous ligand-independent manner. The AHR pathway might represent a potential novel therapeutic target for PTC in the clinical practice.

  20. Molecular chaperone Hsp27 regulates the Hippo tumor suppressor pathway in cancer

    PubMed Central

    Vahid, Sepideh; Thaper, Daksh; Gibson, Kate F.; Bishop, Jennifer L.; Zoubeidi, Amina

    2016-01-01

    Heat shock protein 27 (Hsp27) is a molecular chaperone highly expressed in aggressive cancers, where it is involved in numerous pro-tumorigenic signaling pathways. Using functional genomics we identified for the first time that Hsp27 regulates the gene signature of transcriptional co-activators YAP and TAZ, which are negatively regulated by the Hippo Tumor Suppressor pathway. The Hippo pathway inactivates YAP by phosphorylating and increasing its cytoplasmic retention with the 14.3.3 proteins. Gain and loss of function experiments in prostate, breast and lung cancer cells showed that Hsp27 knockdown induced YAP phosphorylation and cytoplasmic localization while overexpression of Hsp27 displayed opposite results. Mechanistically, Hsp27 regulates the Hippo pathway by accelerating the proteasomal degradation of ubiquitinated MST1, the core Hippo kinase, resulting in reduced phosphorylation/activity of LATS1 and MOB1, its downstream effectors. Importantly, our in vitro results were supported by data from human tumors; clinically, high expression of Hsp27 in prostate tumors is correlated with increased expression of YAP gene signature and reduced phosphorylation of YAP in lung and invasive breast cancer clinical samples. This study reveals for the first time a link between Hsp27 and the Hippo cascade, providing a novel mechanism of deregulation of this tumor suppressor pathway across multiple cancers. PMID:27555231

  1. Regulating the ubiquitin/proteasome pathway via cAMP-signaling: neuroprotective potential

    PubMed Central

    Huang, He; Wang, Hu; Figueiredo-Pereira, Maria E.

    2013-01-01

    The cAMP-signaling pathway has been under intensive investigation for decades. It is a wonder that such a small simple molecule like cAMP can modulate a vast number of diverse processes in different types of cells. The ubiquitous involvement of cAMP-signaling in a variety of cellular events requires tight spatial and temporal control of its generation, propagation, compartmentalization, and elimination. Among the various steps of the cAMP-signaling pathway, G-protein coupled receptors, adenylate cyclases, phosphodiesterases, the two major cAMP targets, i.e. protein kinase A and exchange protein activated by cAMP, as well as the A-kinase anchoring proteins, are potential targets for drug development. Herein we review the recent progress on the regulation and manipulation of different steps of the cAMP-signaling pathway. We end by focusing on the emerging role of cAMP-signaling in modulating protein degradation via the ubiquitin/proteasome pathway. New discoveries on the regulation of the ubiquitin/proteasome pathway by cAMP-signaling support the development of new therapeutic approaches to prevent proteotoxicity in chronic neurodegenerative disorders and other human disease conditions associated with impaired protein turnover by the ubiquitin/proteasome pathway and the accumulation of ubiquitin-protein aggregates. PMID:23686612

  2. Putting the pieces together: How is the mitochondrial pathway of apoptosis regulated in cancer and chemotherapy?

    PubMed Central

    2014-01-01

    In order to solve a jigsaw puzzle, one must first have the complete picture to logically connect the pieces. However, in cancer biology, we are still gaining an understanding of all the signaling pathways that promote tumorigenesis and how these pathways can be pharmacologically manipulated by conventional and targeted therapies. Despite not having complete knowledge of the mechanisms that cause cancer, the signaling networks responsible for cancer are becoming clearer, and this information is serving as a solid foundation for the development of rationally designed therapies. One goal of chemotherapy is to induce cancer cell death through the mitochondrial pathway of apoptosis. Within this review, we present the pathways that govern the cellular decision to undergo apoptosis as three distinct, yet connected puzzle pieces: (1) How do oncogene and tumor suppressor pathways regulate apoptosis upstream of mitochondria? (2) How does the B-cell lymphoma 2 (BCL-2) family influence tumorigenesis and chemotherapeutic responses? (3) How is post-mitochondrial outer membrane permeabilization (MOMP) regulation of cell death relevant in cancer? When these pieces are united, it is possible to appreciate how cancer signaling directly impacts upon the fundamental cellular mechanisms of apoptosis and potentially reveals novel pharmacological targets within these pathways that may enhance chemotherapeutic success. PMID:25621172

  3. Trim65: A cofactor for regulation of the microRNA pathway

    PubMed Central

    Li, Shitao; Wang, Lingyan; Fu, Bishi; Dorf, Martin E

    2014-01-01

    MicroRNA (miRNA) comprise a large family of non-protein coding transcripts which regulate gene expression in diverse biological pathways of both plants and animals. We recently used a systematic proteomic approach to generate a protein interactome map of the human miRNA pathway involved in miRNA biogenesis and processing. The interactome expands the number of candidate proteins in the miRNA pathway and connects the network to other cellular processes. Functional analyses identified TRIM65 and at least 3 other proteins as novel regulators of the miRNA pathway. Biochemical studies established that TRIM65 forms stable complexes with TNRC6 proteins and these molecules co-localize in P-body-like structures. Gain of function and RNAi analyses reveal that TRIM65 negatively regulates miRNA-driven suppression of mRNA translation by targeting TNRC6 proteins for ubiquitination and degradation. The potential molecular mechanisms which regulate TRIM65 catalytic activity are discussed. PMID:25483047

  4. 78 FR 41703 - Regulation of Fuels and Fuel Additives: Additional Qualifying Renewable Fuel Pathways Under the...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-11

    ... unfarmed land in other countries into cropland for energy grass-based renewable fuel production... AGENCY 40 CFR Part 80 RIN 2060-AR85 Regulation of Fuels and Fuel Additives: Additional Qualifying Renewable Fuel Pathways Under the Renewable Fuel Standard Program; Final Rule Approving Renewable...

  5. Regulation of Smoothened Trafficking and Hedgehog Signaling by the SUMO Pathway.

    PubMed

    Ma, Guoqiang; Li, Shuang; Han, Yuhong; Li, Shuangxi; Yue, Tao; Wang, Bing; Jiang, Jin

    2016-11-21

    Hedgehog (Hh) signaling plays a central role in development and diseases. Hh activates its signal transducer and GPCR-family protein Smoothened (Smo) by inducing Smo phosphorylation, but whether Smo is activated through other post-translational modifications remains unexplored. Here we show that sumoylation acts in parallel with phosphorylation to promote Smo cell-surface expression and Hh signaling. We find that Hh stimulates Smo sumoylation by dissociating it from a desumoylation enzyme Ulp1. Sumoylation of Smo in turn recruits a deubiquitinase UBPY/USP8 to antagonize Smo ubiquitination and degradation, leading to its cell-surface accumulation and elevated Hh pathway activity. We also provide evidence that Shh stimulates sumoylation of mammalian Smo (mSmo) and that sumoylation promotes ciliary localization of mSmo and Shh pathway activity. Our findings reveal a conserved mechanism whereby the SUMO pathway promotes Hh signaling by regulating Smo subcellular localization and shed light on how sumoylation regulates membrane protein trafficking.

  6. Regulation of TRPM7 Function by IL-6 through the JAK2-STAT3 Signaling Pathway

    PubMed Central

    Wang, Jing; Zhao, Yin; Luo, Zhenzhao; Gao, Yan; Shi, Jing

    2016-01-01

    Aims Previous studies have demonstrated that expression of the TRPM7 channel, which may induce delayed cell death by mediating calcium influx, is precisely regulated. However, functional regulation of TRPM7 channels by endogenous molecules has not been elucidated. The proinflammatory cytokine IL-6 contributes to regulation of Ca2+ influx in cerebral ischemia, but the role of IL-6 in regulating TRPM7 functioning is unknown. Thus, we here investigated the interaction between IL-6 and TRPM7 channels and the relevant mechanisms. Materials and Methods Using whole-cell patch-clamping, we first investigated the effect of IL-6 on TRPM7-like currents in primary cultured cortical neurons. Next, TRPM7-overexpressing HEK293 cells were used to confirm the effect of IL-6/sIL-6R on TRPM7. Finally, we used specific signaling pathway inhibitors to investigate the signaling pathways involved. Results IL-6 or IL-6/sIL-6R dose-dependently inhibited inward TRPM7 currents, in both primary cultured neurons and HEK293 cells overexpressing TRPM7. In intracellular Mg2+-free conditions, extracellular Ca2+ or the α-kinase domain of TRPM7 did not participate in this regulation. The inhibitory effect of IL-6 on TRPM7 could be blocked by specific inhibitors of the JAK2−STAT3 pathway, but not of the PI3K, ERK1/2, or PLC pathways. Conclusions IL-6 inhibits the inward TRPM7 current via the JAK2−STAT3 signaling pathway. PMID:27010689

  7. PDK1-Akt pathway regulates radial neuronal migration and microtubules in the developing mouse neocortex.

    PubMed

    Itoh, Yasuhiro; Higuchi, Maiko; Oishi, Koji; Kishi, Yusuke; Okazaki, Tomohiko; Sakai, Hiroshi; Miyata, Takaki; Nakajima, Kazunori; Gotoh, Yukiko

    2016-05-24

    Neurons migrate a long radial distance by a process known as locomotion in the developing mammalian neocortex. During locomotion, immature neurons undergo saltatory movement along radial glia fibers. The molecular mechanisms that regulate the speed of locomotion are largely unknown. We now show that the serine/threonine kinase Akt and its activator phosphoinositide-dependent protein kinase 1 (PDK1) regulate the speed of locomotion of mouse neocortical neurons through the cortical plate. Inactivation of the PDK1-Akt pathway impaired the coordinated movement of the nucleus and centrosome, a microtubule-dependent process, during neuronal migration. Moreover, the PDK1-Akt pathway was found to control microtubules, likely by regulating the binding of accessory proteins including the dynactin subunit p150(glued) Consistent with this notion, we found that PDK1 regulates the expression of cytoplasmic dynein intermediate chain and light intermediate chain at a posttranscriptional level in the developing neocortex. Our results thus reveal an essential role for the PDK1-Akt pathway in the regulation of a key step of neuronal migration.

  8. Regulation of the transforming growth factor β pathway by reversible ubiquitylation

    PubMed Central

    Al-Salihi, Mazin A.; Herhaus, Lina; Sapkota, Gopal P.

    2012-01-01

    The transforming growth factor β (TGFβ) signalling pathway plays a central role during embryonic development and in adult tissue homeostasis. It regulates gene transcription through a signalling cascade from cell surface receptors to intracellular SMAD transcription factors and their nuclear cofactors. The extent, duration and potency of signalling in response to TGFβ cytokines are intricately regulated by complex biochemical processes. The corruption of these regulatory processes results in aberrant TGFβ signalling and leads to numerous human diseases, including cancer. Reversible ubiquitylation of pathway components is a key regulatory process that plays a critical role in ensuring a balanced response to TGFβ signals. Many studies have investigated the mechanisms by which various E3 ubiquitin ligases regulate the turnover and activity of TGFβ pathway components by ubiquitylation. Moreover, recent studies have shed new light into their regulation by deubiquitylating enzymes. In this report, we provide an overview of current understanding of the regulation of TGFβ signalling by E3 ubiquitin ligases and deubiquitylases. PMID:22724073

  9. The Atg1-Tor pathway regulates yolk catabolism in Drosophila embryos.

    PubMed

    Kuhn, Hallie; Sopko, Richelle; Coughlin, Margaret; Perrimon, Norbert; Mitchison, Tim

    2015-11-15

    Yolk provides an important source of nutrients during the early development of oviparous organisms. It is composed mainly of vitellogenin proteins packed into membrane-bound compartments called yolk platelets. Catabolism of yolk is initiated by acidification of the yolk platelet, leading to the activation of Cathepsin-like proteinases, but it is unknown how this process is triggered. Yolk catabolism initiates at cellularization in Drosophila melanogaster embryos. Using maternal shRNA technology we found that yolk catabolism depends on the Tor pathway and on the autophagy-initiating kinase Atg1. Whereas Atg1 was required for a burst of spatially regulated autophagy during late cellularization, autophagy was not required for initiating yolk catabolism. We propose that the conserved Tor metabolic sensing pathway regulates yolk catabolism, similar to Tor-dependent metabolic regulation on the lysosome.

  10. The STARS signaling pathway: a key regulator of skeletal muscle function.

    PubMed

    Lamon, Séverine; Wallace, Marita A; Russell, Aaron P

    2014-09-01

    During the last decade, the striated muscle activator of Rho signaling (STARS), a muscle-specific protein, has been proposed to play an increasingly important role in skeletal muscle growth, metabolism, regeneration and stress adaptation. STARS influences actin dynamics and, as a consequence, regulates the myocardin-related transcription factor A/serum response factor (MRTF-A/SRF) transcriptional program, a well-known pathway controlling skeletal muscle development and function. Muscle-specific stress conditions, such as exercise, positively regulates, while disuse and degenerative muscle diseases are associated with a downregulation of STARS and its downstream partners, suggesting a pivotal role for STARS in skeletal muscle health. This review provides a comprehensive overview of the known role and regulation of STARS and the members of its signaling pathway, RhoA, MRTF-A and SRF, in skeletal muscle.

  11. Intermediates in the constitutive and regulated secretory pathways released in vitro from semi-intact cells

    PubMed Central

    1992-01-01

    Regulated secretory cells have two pathways that transport secreted proteins from the Golgi complex to the cell surface. To identify carrier vesicles involved in regulated and constitutive secretion, PC12 pheochromocytoma cells were labeled with [35S]sulfate to identify markers for the two secretory pathways, then mechanically permeabilized and incubated in vitro. Small constitutive secretory vesicles, containing mostly sulfated proteoglycans, accumulated during an in vitro incubation with ATP. In the presence of GTP gamma S, the constitutive vesicles became significantly more dense, suggesting that a coated intermediate was stabilized. Larger immature regulated secretory granules, enriched in sulfated secretogranin II, also escaped from the permeabilized cells in vitro. During granule maturation, their density increased and the amount of cofractionating proteoglycans diminished. The data suggest that sorting continues during secretory granule maturation. PMID:1572894

  12. Time-course regulation of quercetin on cell survival/proliferation pathways in human hepatoma cells.

    PubMed

    Granado-Serrano, Ana Belén; Angeles Martín, María; Bravo, Laura; Goya, Luis; Ramos, Sonia

    2008-04-01

    Quercetin, a dietary flavonoid, has been shown to possess anticarcinogenic properties, but the precise molecular mechanisms of action are not thoroughly elucidated. This study was aimed at investigating the time-course regulation effect of quercetin on survival/proliferation pathways in a human hepatoma cell line (HepG2). Quercetin induced a significant time-dependent inactivation of the major survival signaling proteins, i. e., phosphatidylinositol 3-kinase (PI 3-kinase)/protein kinase B (AKT), extracellular regulated kinase (ERK), protein kinase C-alpha (PKC-alpha), in concert with a time-dependent activation of key death-related signals: c-jun amino-terminal kinase (JNK) and PKC-delta. These data suggest that quercetin exerts a tight regulation of survival/proliferation pathways that requires the integration of different signals and persists over time, being the balance of these regulatory signals what determines the fate of HepG2 cells.

  13. Cortactin phosphorylation regulates cell invasion through a pH-dependent pathway

    PubMed Central

    Magalhaes, Marco A. O.; Larson, Daniel R.; Mader, Christopher C.; Bravo-Cordero, Jose Javier; Gil-Henn, Hava; Oser, Matthew; Chen, Xiaoming; Koleske, Anthony J.

    2011-01-01

    Invadopodia are invasive protrusions with proteolytic activity uniquely found in tumor cells. Cortactin phosphorylation is a key step during invadopodia maturation, regulating Nck1 binding and cofilin activity. The precise mechanism of cortactin-dependent cofilin regulation and the roles of this pathway in invadopodia maturation and cell invasion are not fully understood. We provide evidence that cortactin–cofilin binding is regulated by local pH changes at invadopodia that are mediated by the sodium–hydrogen exchanger NHE1. Furthermore, cortactin tyrosine phosphorylation mediates the recruitment of NHE1 to the invadopodium compartment, where it locally increases the pH to cause the release of cofilin from cortactin. We show that this mechanism involving cortactin phosphorylation, local pH increase, and cofilin activation regulates the dynamic cycles of invadopodium protrusion and retraction and is essential for cell invasion in 3D. Together, these findings identify a novel pH-dependent regulation of cell invasion. PMID:22105349

  14. Inhibition of Fatty Acid Synthase Reduces Blastocyst Hatching through Regulation of the AKT Pathway in Pigs

    PubMed Central

    Guo, Jing; Kim, Nam-Hyung; Cui, Xiang-Shun

    2017-01-01

    Fatty acid synthase (FASN) is an enzyme responsible for the de novo synthesis of long-chain fatty acids. During oncogenesis, FASN plays a role in growth and survival rather than acting within the energy storage pathways. Here, the function of FASN during early embryonic development was studied using its specific inhibitor, C75. We found that the presence of the inhibitor reduced blastocyst hatching. FASN inhibition decreased Cpt1 expression, leading to a reduction in mitochondria numbers and ATP content. This inhibition of FASN resulted in the down-regulation of the AKT pathway, thereby triggering apoptosis through the activation of the p53 pathway. Activation of the apoptotic pathway also leads to increased accumulation of reactive oxygen species and autophagy. In addition, the FASN inhibitor impaired cell proliferation, a parameter of blastocyst quality for outgrowth. The level of OCT4, an important factor in embryonic development, decreased after treatment with the FASN inhibitor. These results show that FASN exerts an effect on early embryonic development by regulating both fatty acid oxidation and the AKT pathway in pigs. PMID:28107461

  15. The molecular choreography of protein synthesis: translational control, regulation, and pathways.

    PubMed

    Chen, Jin; Choi, Junhong; O'Leary, Seán E; Prabhakar, Arjun; Petrov, Alexey; Grosely, Rosslyn; Puglisi, Elisabetta Viani; Puglisi, Joseph D

    2016-01-01

    Translation of proteins by the ribosome regulates gene expression, with recent results underscoring the importance of translational control. Misregulation of translation underlies many diseases, including cancer and many genetic diseases. Decades of biochemical and structural studies have delineated many of the mechanistic details in prokaryotic translation, and sketched the outlines of eukaryotic translation. However, translation may not proceed linearly through a single mechanistic pathway, but likely involves multiple pathways and branchpoints. The stochastic nature of biological processes would allow different pathways to occur during translation that are biased by the interaction of the ribosome with other translation factors, with many of the steps kinetically controlled. These multiple pathways and branchpoints are potential regulatory nexus, allowing gene expression to be tuned at the translational level. As research focus shifts toward eukaryotic translation, certain themes will be echoed from studies on prokaryotic translation. This review provides a general overview of the dynamic data related to prokaryotic and eukaryotic translation, in particular recent findings with single-molecule methods, complemented by biochemical, kinetic, and structural findings. We will underscore the importance of viewing the process through the viewpoints of regulation, translational control, and heterogeneous pathways.

  16. Activation of the TOR Signalling Pathway by Glutamine Regulates Insect Fecundity.

    PubMed

    Zhai, Yifan; Sun, Zhongxiang; Zhang, Jianqing; Kang, Kui; Chen, Jie; Zhang, Wenqing

    2015-05-29

    The target of rapamycin (TOR) positively controls cell growth in response to nutrients such as amino acids. However, research on the specific nutrients sensed by TOR is limited. Glutamine (Gln), a particularly important amino acid involved in metabolism in organisms, is synthesised and catalysed exclusively by glutamine synthetase (GS), and our previous studies have shown that Gln may regulate fecundity in vivo levels of the brown planthopper (BPH) Nilaparvata lugens. Until now, it has remained unclear whether Gln activates or inhibits the TOR signalling pathway. Here, we performed the combined analyses of iTRAQ (isobaric tags for relative and absolute quantification) and DGE (tag-based digital gene expression) data in N. lugens at the protein and transcript levels after GS RNAi, and we found that 52 pathways overlap, including the TOR pathway. We further experimentally demonstrate that Gln activates the TOR pathway by promoting the serine/threonine protein kinase AKT and inhibiting the 5'AMP-activated protein kinase AMPK phosphorylation activity in the pest. Furthermore, TOR regulates the fecundity of N. lugens probably by mediating vitellogenin (Vg) expression. This work is the first report that Gln activates the TOR pathway in vivo.

  17. The SLIT-ROBO pathway: a regulator of cell function with implications for the reproductive system.

    PubMed

    Dickinson, Rachel E; Duncan, W Colin

    2010-04-01

    The secreted SLIT glycoproteins and their Roundabout (ROBO) receptors were originally identified as important axon guidance molecules. They function as a repulsive cue with an evolutionarily conserved role in preventing axons from migrating to inappropriate locations during the assembly of the nervous system. In addition the SLIT-ROBO interaction is involved in the regulation of cell migration, cell death and angiogenesis and, as such, has a pivotal role during the development of other tissues such as the lung, kidney, liver and breast. The cellular functions that the SLIT/ROBO pathway controls during tissue morphogenesis are processes that are dysregulated during cancer development. Therefore inactivation of certain SLITs and ROBOs is associated with advanced tumour formation and progression in disparate tissues. Recent research has indicated that the SLIT/ROBO pathway could also have important functions in the reproductive system. The fetal ovary expresses most members of the SLIT and ROBO families. The SLITs and ROBOs also appear to be regulated by steroid hormones and regulate physiological cell functions in adult reproductive tissues such as the ovary and endometrium. Furthermore several SLITs and ROBOs are aberrantly expressed during the development of ovarian, endometrial, cervical and prostate cancer. This review will examine the roles this pathway could have in the development, physiology and pathology of the reproductive system and highlight areas for future research that could further dissect the influence of the SLIT/ROBO pathway in reproduction.

  18. New insights into the regulation of plant immunity by amino acid metabolic pathways.

    PubMed

    Zeier, Jürgen

    2013-12-01

    Besides defence pathways regulated by classical stress hormones, distinct amino acid metabolic pathways constitute integral parts of the plant immune system. Mutations in several genes involved in Asp-derived amino acid biosynthetic pathways can have profound impact on plant resistance to specific pathogen types. For instance, amino acid imbalances associated with homoserine or threonine accumulation elevate plant immunity to oomycete pathogens but not to pathogenic fungi or bacteria. The catabolism of Lys produces the immune signal pipecolic acid (Pip), a cyclic, non-protein amino acid. Pip amplifies plant defence responses and acts as a critical regulator of plant systemic acquired resistance, defence priming and local resistance to bacterial pathogens. Asp-derived pyridine nucleotides influence both pre- and post-invasion immunity, and the catabolism of branched chain amino acids appears to affect plant resistance to distinct pathogen classes by modulating crosstalk of salicylic acid- and jasmonic acid-regulated defence pathways. It also emerges that, besides polyamine oxidation and NADPH oxidase, Pro metabolism is involved in the oxidative burst and the hypersensitive response associated with avirulent pathogen recognition. Moreover, the acylation of amino acids can control plant resistance to pathogens and pests by the formation of protective plant metabolites or by the modulation of plant hormone activity.

  19. The neutral sphingomyelinase pathway regulates packaging of the prion protein into exosomes.

    PubMed

    Guo, Belinda B; Bellingham, Shayne A; Hill, Andrew F

    2015-02-06

    Prion diseases are a group of transmissible, fatal neurodegenerative disorders associated with the misfolding of the host-encoded prion protein, PrP(C), into a disease-associated form, PrP(Sc). The transmissible prion agent is principally formed of PrP(Sc) itself and is associated with extracellular vesicles known as exosomes. Exosomes are released from cells both in vitro and in vivo, and have been proposed as a mechanism by which prions spread intercellularly. The biogenesis of exosomes occurs within the endosomal system, through formation of intraluminal vesicles (ILVs), which are subsequently released from cells as exosomes. ILV formation is known to be regulated by the endosomal sorting complexes required for transport (ESCRT) machinery, although an alternative neutral sphingomyelinase (nSMase) pathway has been suggested to also regulate this process. Here, we investigate a role for the nSMase pathway in exosome biogenesis and packaging of PrP into these vesicles. Inhibition of the nSMase pathway using GW4869 revealed a role for the nSMase pathway in both exosome formation and PrP packaging. In agreement, targeted knockdown of nSMase1 and nSMase2 in mouse neurons using lentivirus-mediated RNAi also decreases exosome release, demonstrating the nSMase pathway regulates the biogenesis and release of exosomes. We also demonstrate that PrP(C) packaging is dependent on nSMase2, whereas the packaging of disease-associated PrP(Sc) into exosomes occurs independently of nSMase2. These findings provide further insight into prion transmission and identify a pathway which directly assists exosome-mediated transmission of prions.

  20. The TOR signaling pathway regulates vegetative development and virulence in Fusarium graminearum.

    PubMed

    Yu, Fangwei; Gu, Qin; Yun, Yingzi; Yin, Yanni; Xu, Jin-Rong; Shim, Won-Bo; Ma, Zhonghua

    2014-07-01

    The target of rapamycin (TOR) signaling pathway plays critical roles in controlling cell growth in a variety of eukaryotes. However, the contribution of this pathway in regulating virulence of plant pathogenic fungi is unknown. We identified and characterized nine genes encoding components of the TOR pathway in Fusarium graminearum. Biological, genetic and biochemical functions of each component were investigated. The FgFkbp12-rapamycin complex binds to the FgTor kinase. The type 2A phosphatases FgPp2A, FgSit4 and FgPpg1 were found to interact with FgTap42, a downstream component of FgTor. Among these, we determined that FgPp2A is likely to be essential for F. graminearum survival, and FgSit4 and FgPpg1 play important roles in cell wall integrity by positively regulating the phosphorylation of FgMgv1, a key MAP kinase in the cell wall integrity pathway. In addition, the FgPpg1 interacting protein, FgTip41, is involved in regulating mycelial growth and virulence. Notably, FgTip41 does not interact with FgTap42 but with FgPpg1, suggesting the existence of FgTap42:FgPpg1:FgTip41 heterotrimer in F. graminearum, a complex not observed in the yeast model. Collectively, we defined a genetic regulatory framework that elucidates how the TOR pathway regulates virulence and vegetative development in F. graminearum.

  1. Regulation of necrotic cell death: p53, PARP1 and cyclophilin D-overlapping pathways of regulated necrosis?

    PubMed

    Ying, Yuan; Padanilam, Babu J

    2016-06-01

    In contrast to apoptosis and autophagy, necrotic cell death was considered to be a random, passive cell death without definable mediators. However, this dogma has been challenged by recent developments suggesting that necrotic cell death can also be a regulated process. Regulated necrosis includes multiple cell death modalities such as necroptosis, parthanatos, ferroptosis, pyroptosis, and mitochondrial permeability transition pore (MPTP)-mediated necrosis. Several distinctive executive molecules, particularly residing on the mitochondrial inner and outer membrane, amalgamating to form the MPTP have been defined. The c-subunit of the F1F0ATP synthase on the inner membrane and Bax/Bak on the outer membrane are considered to be the long sought components that form the MPTP. Opening of the MPTP results in loss of mitochondrial inner membrane potential, disruption of ATP production, increased ROS production, organelle swelling, mitochondrial dysfunction and consequent necrosis. Cyclophilin D, along with adenine nucleotide translocator and the phosphate carrier are considered to be important regulators involved in the opening of MPTP. Increased production of ROS can further trigger other necrotic pathways mediated through molecules such as PARP1, leading to irreversible cell damage. This review examines the roles of PARP1 and cyclophilin D in necrotic cell death. The hierarchical role of p53 in regulation and integration of key components of signaling pathway to elicit MPTP-mediated necrosis and ferroptosis is explored. In the context of recent insights, the indistinct role of necroptosis signaling in tubular necrosis after ischemic kidney injury is scrutinized. We conclude by discussing the participation of p53, PARP1 and cyclophilin D and their overlapping pathways to elicit MPTP-mediated necrosis and ferroptosis in acute kidney injury.

  2. Early Diagnosis of Clear Cell Kidney Cancer via VHL/HIF Pathway-regulated Circulating microRNA

    DTIC Science & Technology

    2013-09-01

    Cancer via VHL /HIF Pathway-regulated Circulating microRNA PRINCIPAL INVESTIGATOR: Allan Pantuck, MD...SUBTITLE Early Diagnosis of Clear Cell Kidney Cancer via VHL /HIF Pathway-regulated Circulating microRNA 5a. CONTRACT NUMBER 5b. GRANT NUMBER

  3. Hedgehog pathway regulators influence cervical cancer cell proliferation, survival and migration

    SciTech Connect

    Samarzija, Ivana; Beard, Peter

    2012-08-17

    Highlights: Black-Right-Pointing-Pointer Unknown cellular mutations complement papillomavirus-induced carcinogenesis. Black-Right-Pointing-Pointer Hedgehog pathway components are expressed by cervical cancer cells. Black-Right-Pointing-Pointer Hedgehog pathway activators and inhibitors regulate cervical cancer cell biology. Black-Right-Pointing-Pointer Cell immortalization by papillomavirus and activation of Hedgehog are independent. -- Abstract: Human papillomavirus (HPV) infection is considered to be a primary hit that causes cervical cancer. However, infection with this agent, although needed, is not sufficient for a cancer to develop. Additional cellular changes are required to complement the action of HPV, but the precise nature of these changes is not clear. Here, we studied the function of the Hedgehog (Hh) signaling pathway in cervical cancer. The Hh pathway can have a role in a number of cancers, including those of liver, lung and digestive tract. We found that components of the Hh pathway are expressed in several cervical cancer cell lines, indicating that there could exists an autocrine Hh signaling loop in these cells. Inhibition of Hh signaling reduces proliferation and survival of the cervical cancer cells and induces their apoptosis as seen by the up-regulation of the pro-apoptotic protein cleaved caspase 3. Our results indicate that Hh signaling is not induced directly by HPV-encoded proteins but rather that Hh-activating mutations are selected in cells initially immortalized by HPV. Sonic Hedgehog (Shh) ligand induces proliferation and promotes migration of the cervical cancer cells studied. Together, these results indicate pro-survival and protective roles of an activated Hh signaling pathway in cervical cancer-derived cells, and suggest that inhibition of this pathway may be a therapeutic option in fighting cervical cancer.

  4. Melatonin regulates aging and neurodegeneration through energy metabolism, epigenetics, autophagy and circadian rhythm pathways.

    PubMed

    Jenwitheesuk, Anorut; Nopparat, Chutikorn; Mukda, Sujira; Wongchitrat, Prapimpun; Govitrapong, Piyarat

    2014-09-22

    Brain aging is linked to certain types of neurodegenerative diseases and identifying new therapeutic targets has become critical. Melatonin, a pineal hormone, associates with molecules and signaling pathways that sense and influence energy metabolism, autophagy, and circadian rhythms, including insulin-like growth factor 1 (IGF-1), Forkhead box O (FoxOs), sirtuins and mammalian target of rapamycin (mTOR) signaling pathways. This review summarizes the current understanding of how melatonin, together with molecular, cellular and systemic energy metabolisms, regulates epigenetic processes in the neurons. This information will lead to a greater understanding of molecular epigenetic aging of the brain and anti-aging mechanisms to increase lifespan under healthy conditions.

  5. Utilization of economical substrate-derived carbohydrates by solventogenic clostridia: pathway dissection, regulation and engineering.

    PubMed

    Gu, Yang; Jiang, Yu; Yang, Sheng; Jiang, Weihong

    2014-10-01

    Solventogenic clostridia can produce acetone, butanol and ethanol (ABE) by using different carbohydrates. For economical reasons, the utilization of cheap and renewable biomass in clostridia-based ABE fermentation has recently attracted increasing interests. With the study of molecular microbiology and development of genetic tools, the understanding of carbohydrate metabolism in clostridia has increased in recent years. Here, we review the pioneering work in this field, with a focus on dissecting the pathways and describing the regulation of the metabolism of economical substrate-derived carbohydrates by clostridia. Recent progress in the metabolic engineering of carbohydrate utilization pathways is also described.

  6. Dicer-dependent pathway contribute to the osteogenesis mediated by regulation of Runx2

    PubMed Central

    Zhou, Jie; Hu, Yun; Chen, Yang; Yang, Lan; Song, Jinlin; Tang, Yuying; Deng, Feng; Zheng, Leilei

    2016-01-01

    Osteogenesis is mediated by sophisticated interactions of various molecular functions and biological processes, including post-transcriptional regulation. A range of miRNAs have been reported to regulate bone homeostasis and osteoblasts differentiation either positively or negatively through multiple signaling pathways. RNase III endonuclease Dicer is the key enzyme required for the biogenesis of miRNAs and small interfering RNAs. To determine the global influence of miRNAs on regulation of osteogenesis of pre-osteoblast cells, the transcriptional regulation of Dicer and the function of Dicer during osteoblast differentiation and mineralization were investigated. Runx2 binding directly to the Dicer promoter region was characterized in MC3T3-E1 cells by chromatin immunoprecipitation (ChIP) and luciferase promoter reporter assays. Overexpression or knockdown of Runx2 resulted in increase or decrease of Dicer expression, respectively. Furthermore, abatement of Dicer in MC3T3-E1 cells down-regulated the expression of osteogenic marker genes and mineralization ability, at least partly involving Dicer-dependent processing of the miR-21a-5p targeting PTEN via pAKT/pGSK3β/β-catenin signaling pathways. Taken together, the study demonstrates the role of Dicer in osteogenesis and suggests that Dicer is required, in part, for Runx2 regulation of osteoblast differentiation. PMID:28078008

  7. Procontractile G protein–mediated signaling pathways antagonistically regulate smooth muscle differentiation in vascular remodeling

    PubMed Central

    Althoff, Till F.; Juárez, Julián Albarrán; Troidl, Kerstin; Tang, Cong; Wang, Shengpeng; Wirth, Angela; Takefuji, Mikito; Wettschureck, Nina

    2012-01-01

    Vascular smooth muscle (Sm) cells (VSMCs) are highly plastic. Their differentiation state can be regulated by serum response factor (SRF), which activates genes involved in Sm differentiation and proliferation by recruiting cofactors, such as members of the myocardin family and ternary complex factors (TCFs), respectively. However, the extracellular cues and upstream signaling mechanisms regulating SRF-dependent VSMC differentiation under in vivo conditions are poorly understood. In this study, we show that the procontractile signaling pathways mediated by the G proteins G12/G13 and Gq/G11 antagonistically regulate VSMC plasticity in different models of vascular remodeling. In mice lacking Gα12/Gα13 or their effector, the RhoGEF protein LARG, RhoA-dependent SRF-regulation was blocked and down-regulation of VSMC differentiation marker genes was enhanced. This was accompanied by an excessive vascular remodeling and exacerbation of atherosclerosis. In contrast, Sm-specific Gαq/Gα11 deficiency blocked activation of extracellular signal-regulated kinase 1/2 and the TCF Elk-1, resulting in a reduced VSMC dedifferentiation in response to flow cessation or vascular injury. These data show that the balanced activity of both G protein–mediated pathways in VSMCs is required for an appropriate vessel remodeling response in vascular diseases and suggest new approaches to modulate Sm differentiation in vascular pathologies. PMID:23129751

  8. OTUB1 modulates c-IAP1 stability to regulate signalling pathways

    PubMed Central

    Goncharov, Tatiana; Niessen, Kyle; de Almagro, Maria Cristina; Izrael-Tomasevic, Anita; Fedorova, Anna V; Varfolomeev, Eugene; Arnott, David; Deshayes, Kurt; Kirkpatrick, Donald S; Vucic, Domagoj

    2013-01-01

    The cellular inhibitor of apoptosis (c-IAP) proteins are E3 ubiquitin ligases that are critical regulators of tumour necrosis factor (TNF) receptor (TNFR)-mediated signalling. Through their E3 ligase activity c-IAP proteins promote ubiquitination of receptor-interaction protein 1 (RIP1), NF-κB-inducing kinase (NIK) and themselves, and regulate the assembly of TNFR signalling complexes. Consequently, in the absence of c-IAP proteins, TNFR-mediated activation of NF-κB and MAPK pathways and the induction of gene expression are severely reduced. Here, we describe the identification of OTUB1 as a c-IAP-associated deubiquitinating enzyme that regulates c-IAP1 stability. OTUB1 disassembles K48-linked polyubiquitin chains from c-IAP1 in vitro and in vivo within the TWEAK receptor-signalling complex. Downregulation of OTUB1 promotes TWEAK- and IAP antagonist-stimulated caspase activation and cell death, and enhances c-IAP1 degradation. Furthermore, knockdown of OTUB1 reduces TWEAK-induced activation of canonical NF-κB and MAPK signalling pathways and modulates TWEAK-induced gene expression. Finally, suppression of OTUB1 expression in zebrafish destabilizes c-IAP (Birc2) protein levels and disrupts fish vasculature. These results suggest that OTUB1 regulates NF-κB and MAPK signalling pathways and TNF-dependent cell death by modulating c-IAP1 stability. PMID:23524849

  9. Regulation of the salvage pathway of deoxynucleotides synthesis in apoptosis induced by growth factor deprivation.

    PubMed Central

    Oliver, F J; Collins, M K; López-Rivas, A

    1996-01-01

    Here we describe changes in dNTP metabolism that precede DNA fragmentation in a model of apoptosis driven by deprivation of the cytokine interleukin 3 (IL-3). In haemopoietic BAF3 cells, IL-3 withdrawal leads to a rapid decrease in the size of dATP, dTTP and dGTP pools without affecting dCTP levels. This imbalance in dNTP pools precedes DNA fragmentation and is accompanied by down-regulation of enzymes controlling the de novo and salvage pathways of dNTP synthesis, ribonucleotide reductase and thymidine kinase (TK) respectively. Readdition of IL-3 results in a rapid, protein synthesis-independent restoration of normal dNTP pools, enhanced TK activity and increased precursor incorporation through the salvage pathway. Up-regulation of TK activity after IL-3 readdition is prevented by the protein kinase C (PKC) inhibitor staurosporin, but not by tyrosine kinase inhibitors. Furthermore activation of PKC by phorbol esters mimics the stimulatory effect of IL-3 on TK activity, suggesting that PKC might be involved in regulating this effect. These results indicate that regulation by IL-3 of the salvage pathway of dNTP synthesis plays a role in the maintenance of cellular dNTP pool balance and suggests that alterations in dNTP metabolism after IL-3 deprivation could be a relevant event in the commitment of haemopoietic cells to apoptosis. PMID:8687383

  10. The DAF-7 TGF-β signaling pathway regulates chemosensory receptor gene expression in C. elegans

    PubMed Central

    Nolan, Katherine M.; Sarafi-Reinach, Trina R.; Horne, Jennifer G.; Saffer, Adam M.; Sengupta, Piali

    2002-01-01

    Regulation of chemoreceptor gene expression in response to environmental or developmental cues provides a mechanism by which animals can alter their sensory responses. Here we demonstrate a role for the daf-7 TGF-β pathway in the regulation of expression of a subset of chemoreceptor genes in Caenorhabditis elegans. We describe a novel role of this pathway in maintaining receptor gene expression in the adult and show that the DAF-4 type II TGF-β receptor functions cell-autonomously to modulate chemoreceptor expression. We also find that the alteration of receptor gene expression in the ASI chemosensory neurons by environmental signals, such as levels of a constitutively produced pheromone, may be mediated via a DAF-7-independent pathway. Receptor gene expression in the ASI and ASH sensory neurons appears to be regulated via distinct mechanisms. Our results suggest that the expression of individual chemoreceptor genes in C. elegans is subject to multiple modes of regulation, thereby ensuring that animals exhibit the responses most appropriate for their developmental stage and environmental conditions. PMID:12464635

  11. Regulation of the retinoblastoma-E2F pathway by the ubiquitin-proteasome system.

    PubMed

    Sengupta, Satyaki; Henry, R William

    2015-10-01

    The retinoblastoma tumor suppressor (RB) and its related family members p107 and p130 regulate cell proliferation through the transcriptional repression of genes involved in cellular G1 to S phase transition. However, RB proteins are functionally versatile, and numerous genetic and biochemical studies point to expansive roles in cellular growth control, pluripotency, and apoptotic response. For the vast majority of genes, RB family members target the E2F family of transcriptional activators as an integral component of its gene regulatory mechanism. These interactions are regulated via reversible phosphorylation by Cyclin/Cyclin-dependent kinase (Cdk) complexes, a major molecular mechanism that regulates transcriptional output of RB/E2F target genes. Recent studies indicate an additional level of regulation involving the ubiquitin-proteasome system that renders pervasive control over each component of the RB pathway. Disruption of the genetic circuitry for proteasome-mediated targeting of the RB pathway has serious consequences on development and cellular transformation, and is associated with several forms of human cancer. In this review, we discuss the role of the ubiquitin-proteasome system in proteolytic control of RB-E2F pathway components, and recent data that points to surprising non-proteolytic roles for the ubiquitin-proteasome system in novel transcriptional regulatory mechanisms.

  12. mTOR regulate EMT through RhoA and Rac1 pathway in prostate cancer.

    PubMed

    Chen, XianGuo; Cheng, HaiYan; Pan, TengFei; Liu, Yi; Su, Yang; Ren, CuiPing; Huang, DaKe; Zha, XiaoJun; Liang, ChaoZhao

    2015-10-01

    Recently, an increasing number of studies have suggested that mTOR plays a critical role in the regulation of tumor cell motility, invasion and cancer metastasis. However, little is known about the signaling mechanisms in regulating epithelial-mesenchymal transition (EMT) of prostate cancer. In this study, we found that the expression levels of Raptor and Rictor in prostate cancer tissues were elevated, which may suggest that Raptor and Rictor signaling pathways are associated with prostate cancer progression and metastasis. Inhibition of mTORC1 or mTORC2 by knock down of Raptor or Rictor, respectively, migration and invasion of prostate cancer were attenuated. Furthermore, EMT, a characterized by the changed expression levels of various markers like E-cadherin, β-catenin, N-cadherin, and vimentin emergend following inhibition of Raptor or Rictor. Finally, the small GTPases (RhoA and Rac1) which were crucial regulatory proteins in cell migration and invasion were inactivited after downregulating Raptor and Rictor. These results suggest that mTOR regulate EMT at least in part by down regulation of RhoA and Rac1 signaling pathways. Our findings provide novel very attractive target strategies that the inhibition of mTOR signaling pathways may retard prostate cancer migration and invasion at early stages.

  13. Identification of pathways directly regulated by SHORT VEGETATIVE PHASE during vegetative and reproductive development in Arabidopsis

    PubMed Central

    2013-01-01

    Background MADS-domain transcription factors play important roles during plant development. The Arabidopsis MADS-box gene SHORT VEGETATIVE PHASE (SVP) is a key regulator of two developmental phases. It functions as a repressor of the floral transition during the vegetative phase and later it contributes to the specification of floral meristems. How these distinct activities are conferred by a single transcription factor is unclear, but interactions with other MADS domain proteins which specify binding to different genomic regions is likely one mechanism. Results To compare the genome-wide DNA binding profile of SVP during vegetative and reproductive development we performed ChIP-seq analyses. These ChIP-seq data were combined with tiling array expression analysis, induction experiments and qRT-PCR to identify biologically relevant binding sites. In addition, we compared genome-wide target genes of SVP with those published for the MADS domain transcription factors FLC and AP1, which interact with SVP during the vegetative and reproductive phases, respectively. Conclusions Our analyses resulted in the identification of pathways that are regulated by SVP including those controlling meristem development during vegetative growth and flower development whereas floral transition pathways and hormonal signaling were regulated predominantly during the vegetative phase. Thus, SVP regulates many developmental pathways, some of which are common to both of its developmental roles whereas others are specific to only one of them. PMID:23759218

  14. Soldier morphogenesis in the damp-wood termite is regulated by the insulin signaling pathway.

    PubMed

    Hattori, Akiko; Sugime, Yasuhiro; Sasa, Chifune; Miyakawa, Hitoshi; Ishikawa, Yuki; Miyazaki, Satoshi; Okada, Yasukazu; Cornette, Richard; Lavine, Laura Corley; Emlen, Douglas J; Koshikawa, Shigeyuki; Miura, Toru

    2013-07-01

    Eusocial insects exhibit various morphological castes associated with the division of labor within a colony. Termite soldiers possess defensive traits including mandibles that are greatly exaggerated and enlarged, as compared to termite reproductives and workers. The enlarged mandibles of soldiers are known to result from dynamic morphogenesis during soldier differentiation that can be induced by juvenile hormone and its analogs. However, the detailed developmental mechanisms still remain unresolved. Because the insulin/insulin-like growth factor signaling (IIS) pathway has been shown to regulate the relative sizes of organs (i.e., allometry) in other insects, we examined the expression profiles of major IIS factors in the damp-wood termite Hodotermopsis sjostedti, during soldier differentiation. The relative expression patterns of orthologs for termite InR (HsjInR), PKB/Akt (HsjPKB/Akt), and FOXO (HsjFOXO) suggest that HsjInR and HsjPKB/Akt were up-regulated in the period of elongation of mandibles during soldier development. In situ hybridization showed that HsjInR was strongly expressed in the mandibular epithelial tissues, and RNA interference (RNAi) for HsjInR disrupted soldier-specific morphogenesis including mandibular elongation. These results suggest that signaling through the IIS pathway is required for soldier-specific morphogenesis. In addition, up-regulation of the IIS pathway in other body tissues occurred at earlier stages of development, indicating that there is tissue-specific IIS regulation. Because the IIS pathway is generally thought to act upstream of JH in insects, our results suggest the damp-wood termite may have evolved a novel feedback loop between JH and IIS that enables social interactions, rather than nutrition, to regulate caste determination.

  15. Bioinformatics Analysis Reveals MicroRNAs Regulating Biological Pathways in Exercise-Induced Cardiac Physiological Hypertrophy

    PubMed Central

    Xu, Jiahong; Liu, Yang; Xie, Yuan

    2017-01-01

    Exercise-induced physiological cardiac hypertrophy is generally considered to be a type of adaptive change after exercise training and is beneficial for cardiovascular diseases. This study aims at investigating exercise-regulated microRNAs (miRNAs) and their potential biological pathways. Here, we collected 23 miRNAs from 8 published studies. MirPath v.3 from the DIANA tools website was used to execute the analysis, and TargetScan was used to predict the target genes. Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses were performed to identify potential pathways and functional annotations associated with exercise-induced physiological cardiac hypertrophy. Various miRNA targets and molecular pathways, such as Fatty acid elongation, Arrhythmogenic right ventricular cardiomyopathy (ARVC), and ECM-receptor interaction, were identified. This study could prompt the understanding of the regulatory mechanisms underlying exercise-induced physiological cardiac hypertrophy. PMID:28286759

  16. Bioinformatics Analysis Reveals MicroRNAs Regulating Biological Pathways in Exercise-Induced Cardiac Physiological Hypertrophy.

    PubMed

    Xu, Jiahong; Liu, Yang; Xie, Yuan; Zhao, Cuimei; Wang, Hongbao

    2017-01-01

    Exercise-induced physiological cardiac hypertrophy is generally considered to be a type of adaptive change after exercise training and is beneficial for cardiovascular diseases. This study aims at investigating exercise-regulated microRNAs (miRNAs) and their potential biological pathways. Here, we collected 23 miRNAs from 8 published studies. MirPath v.3 from the DIANA tools website was used to execute the analysis, and TargetScan was used to predict the target genes. Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analyses were performed to identify potential pathways and functional annotations associated with exercise-induced physiological cardiac hypertrophy. Various miRNA targets and molecular pathways, such as Fatty acid elongation, Arrhythmogenic right ventricular cardiomyopathy (ARVC), and ECM-receptor interaction, were identified. This study could prompt the understanding of the regulatory mechanisms underlying exercise-induced physiological cardiac hypertrophy.

  17. Agrin as a Mechanotransduction Signal Regulating YAP through the Hippo Pathway.

    PubMed

    Chakraborty, Sayan; Njah, Kizito; Pobbati, Ajaybabu V; Lim, Ying Bena; Raju, Anandhkumar; Lakshmanan, Manikandan; Tergaonkar, Vinay; Lim, Chwee Teck; Hong, Wanjin

    2017-03-07

    The Hippo pathway effectors YAP and TAZ act as nuclear sensors of mechanical signals in response to extracellular matrix (ECM) cues. However, the identity and nature of regulators in the ECM and the precise pathways relaying mechanoresponsive signals into intracellular sensors remain unclear. Here, we uncover a functional link between the ECM proteoglycan Agrin and the transcriptional co-activator YAP. Importantly, Agrin transduces matrix and cellular rigidity signals that enhance stability and mechanoactivity of YAP through the integrin-focal adhesion- and Lrp4/MuSK receptor-mediated signaling pathways. Agrin antagonizes focal adhesion assembly of the core Hippo components by facilitating ILK-PAK1 signaling and negating the functions of Merlin and LATS1/2. We further show that Agrin promotes oncogenesis through YAP-dependent transcription and is clinically relevant in human liver cancer. We propose that Agrin acts as a mechanotransduction signal in the ECM.

  18. Integrative Approaches to Enhance Understanding of Plant Metabolic Pathway Structure and Regulation1

    PubMed Central

    Tohge, Takayuki; Scossa, Federico; Fernie, Alisdair R.

    2015-01-01

    Huge insight into molecular mechanisms and biological network coordination have been achieved following the application of various profiling technologies. Our knowledge of how the different molecular entities of the cell interact with one another suggests that, nevertheless, integration of data from different techniques could drive a more comprehensive understanding of the data emanating from different techniques. Here, we provide an overview of how such data integration is being used to aid the understanding of metabolic pathway structure and regulation. We choose to focus on the pairwise integration of large-scale metabolite data with that of the transcriptomic, proteomics, whole-genome sequence, growth- and yield-associated phenotypes, and archival functional genomic data sets. In doing so, we attempt to provide an update on approaches that integrate data obtained at different levels to reach a better understanding of either single gene function or metabolic pathway structure and regulation within the context of a broader biological process. PMID:26371234

  19. Ligand concentration regulates the pathways of coupled protein folding and binding.

    PubMed

    Daniels, Kyle G; Tonthat, Nam K; McClure, David R; Chang, Yu-Chu; Liu, Xin; Schumacher, Maria A; Fierke, Carol A; Schmidler, Scott C; Oas, Terrence G

    2014-01-22

    Coupled ligand binding and conformational change plays a central role in biological regulation. Ligands often regulate protein function by modulating conformational dynamics, yet the order in which binding and conformational change occurs are often hotly debated. Here we show that the "conformational selection versus induced fit" distinction on which this debate is based is a false dichotomy because the mechanism depends on ligand concentration. Using the binding of pyrophosphate (PPi) to Bacillus subtilis RNase P protein as a model, we show that coupled reactions are best understood as a change in flux between competing pathways with distinct orders of binding and conformational change. The degree of partitioning through each pathway depends strongly on PPi concentration, with ligand binding redistributing the conformational ensemble toward the folded state by both increasing folding rates and decreasing unfolding rates. These results indicate that ligand binding induces marked and varied changes in protein conformational dynamics, and that the order of binding and conformational change is ligand concentration dependent.

  20. Essential Role of TGF-β/Smad Pathway on Statin Dependent Vascular Smooth Muscle Cell Regulation

    PubMed Central

    Rodríguez-Vita, Juan; Sánchez-Galán, Eva; Santamaría, Beatriz; Sánchez-López, Elsa; Rodrigues-Díez, Raquel; Blanco-Colio, Luís Miguel; Egido, Jesús; Ortiz, Alberto; Ruiz-Ortega, Marta

    2008-01-01

    Background The 3-hydroxy-3-methylglutaryl CoA reductase inhibitors (also called statins) exert proven beneficial effects on cardiovascular diseases. Recent data suggest a protective role for Transforming Growth Factor-β (TGF-β) in atherosclerosis by regulating the balance between inflammation and extracellular matrix accumulation. However, there are no studies about the effect of statins on TGF-β/Smad pathway in atherosclerosis and vascular cells. Methodology In cultured vascular smooth muscle cells (VSMCs) statins enhanced Smad pathway activation caused by TGF-β. In addition, statins upregulated TGF-β receptor type II (TRII), and increased TGF-β synthesis and TGF-β/Smad-dependent actions. In this sense, statins, through Smad activation, render VSMCs more susceptible to TGF-β induced apoptosis and increased TGF-β-mediated ECM production. It is well documented that high doses of statins induce apoptosis in cultured VSMC in the presence of serum; however the precise mechanism of this effect remains to be elucidated. We have found that statins-induced apoptosis was mediated by TGF-β/Smad pathway. Finally, we have described that RhoA inhibition is a common intracellular mechanisms involved in statins effects. The in vivo relevance of these findings was assessed in an experimental model of atherosclerosis in apolipoprotein E deficient mice: Treatment with Atorvastatin increased Smad3 phosphorylation and TRII overexpression, associated to elevated ECM deposition in the VSMCs within atheroma plaques, while apoptosis was not detected. Conclusions Statins enhance TGF-β/Smad pathway, regulating ligand levels, receptor, main signaling pathway and cellular responses of VSMC, including apoptosis and ECM accumulation. Our findings show that TGF-β/Smad pathway is essential for statins-dependent actions in VSMCs. PMID:19088845

  1. Functional CRISPR screening identifies the ufmylation pathway as a regulator of SQSTM1/p62

    PubMed Central

    DeJesus, Rowena; Moretti, Francesca; McAllister, Gregory; Wang, Zuncai; Bergman, Phil; Liu, Shanming; Frias, Elizabeth; Alford, John; Reece-Hoyes, John S; Lindeman, Alicia; Kelliher, Jennifer; Russ, Carsten; Knehr, Judith; Carbone, Walter; Beibel, Martin; Roma, Guglielmo; Ng, Aylwin; Tallarico, John A; Porter, Jeffery A; Xavier, Ramnik J; Mickanin, Craig; Murphy, Leon O; Hoffman, Gregory R; Nyfeler, Beat

    2016-01-01

    SQSTM1 is an adaptor protein that integrates multiple cellular signaling pathways and whose expression is tightly regulated at the transcriptional and post-translational level. Here, we describe a forward genetic screening paradigm exploiting CRISPR-mediated genome editing coupled to a cell selection step by FACS to identify regulators of SQSTM1. Through systematic comparison of pooled libraries, we show that CRISPR is superior to RNAi in identifying known SQSTM1 modulators. A genome-wide CRISPR screen exposed MTOR signalling and the entire macroautophagy machinery as key regulators of SQSTM1 and identified several novel modulators including HNRNPM, SLC39A14, SRRD, PGK1 and the ufmylation cascade. We show that ufmylation regulates SQSTM1 by eliciting a cell type-specific ER stress response which induces SQSTM1 expression and results in its accumulation in the cytosol. This study validates pooled CRISPR screening as a powerful method to map the repertoire of cellular pathways that regulate the fate of an individual target protein. DOI: http://dx.doi.org/10.7554/eLife.17290.001 PMID:27351204

  2. Hedgehog signaling pathway regulated the target genes for adipogenesis in silkworm Bombyx mori.

    PubMed

    Liang, Shuang; Chen, Rui-Ting; Zhang, Deng-Pan; Xin, Hu-Hu; Lu, Yan; Wang, Mei-Xian; Miao, Yun-Gen

    2015-10-01

    Hedgehog (Hh) signals regulate invertebrate and vertebrate development, yet the role of the pathway in adipose development remains poorly understood. In this report, we found that Hh pathway components are expressed in the fat body of silkworm larvae. Functional analysis of these components in a BmN cell line model revealed that activation of the Hh gene stimulated transcription of Hh pathway components, but inhibited the expression of the adipose marker gene AP2. Conversely, specific RNA interference-mediated knockdown of Hh resulted in increased AP2 expression. This further showed the regulation of Hh signal on the adipose marker gene. In silkworm larval models, enhanced adipocyte differentiation and an increase in adipocyte cell size were observed in silkworms that had been treated with a specific Hh signaling pathway antagonist, cyclopamine. The fat-body-specific Hh blockade tests were consistent with Hh signaling inhibiting silkworm adipogenesis. Our results indicate that the role of Hh signaling in inhibiting fat formation is conserved in vertebrates and invertebrates.

  3. Genome-wide analysis of a Wnt1-regulated transcriptional network implicates neurodegenerative pathways.

    PubMed

    Wexler, Eric M; Rosen, Ezra; Lu, Daning; Osborn, Gregory E; Martin, Elizabeth; Raybould, Helen; Geschwind, Daniel H

    2011-10-04

    Wnt proteins are critical to mammalian brain development and function. The canonical Wnt signaling pathway involves the stabilization and nuclear translocation of β-catenin; however, Wnt also signals through alternative, noncanonical pathways. To gain a systems-level, genome-wide view of Wnt signaling, we analyzed Wnt1-stimulated changes in gene expression by transcriptional microarray analysis in cultured human neural progenitor (hNP) cells at multiple time points over a 72-hour time course. We observed a widespread oscillatory-like pattern of changes in gene expression, involving components of both the canonical and the noncanonical Wnt signaling pathways. A higher-order, systems-level analysis that combined independent component analysis, waveform analysis, and mutual information-based network construction revealed effects on pathways related to cell death and neurodegenerative disease. Wnt effectors were tightly clustered with presenilin1 (PSEN1) and granulin (GRN), which cause dominantly inherited forms of Alzheimer's disease and frontotemporal dementia (FTD), respectively. We further explored a potential link between Wnt1 and GRN and found that Wnt1 decreased GRN expression by hNPs. Conversely, GRN knockdown increased WNT1 expression, demonstrating that Wnt and GRN reciprocally regulate each other. Finally, we provided in vivo validation of the in vitro findings by analyzing gene expression data from individuals with FTD. These unbiased and genome-wide analyses provide evidence for a connection between Wnt signaling and the transcriptional regulation of neurodegenerative disease genes.

  4. Regulation of GIRK channel deactivation by Galpha(q) and Galpha(i/o) pathways.

    PubMed

    Mark, M D; Ruppersberg, J P; Herlitze, S

    2000-09-01

    G protein regulated inward rectifying potassium channels (GIRKs) are activated by G protein coupled receptors (GPCRs) via the G protein betagamma subunits. However, little is known about the effects of different GPCRs on the deactivation kinetics of transmitter-mediated GIRK currents. In the present study we investigated the influence of different GPCRs in the presence and absence of RGS proteins on the deactivation kinetics of GIRK channels by coexpressing the recombinant protein subunits in Xenopus oocytes. The stimulation of both G(i/o)- and G(q)-coupled pathways accelerated GIRK deactivation. GIRK currents deactivated faster upon stimulation of G(i/o)- and G(q)-coupled pathways by P(2)Y(2) receptors (P(2)Y(2)Rs) than upon activation of the G(i/o)-coupled pathway alone via muscarinic acetylcholine receptor M2 (M(2) mAChRs). This acceleration was found to be dependent on phospholipase C (PLC) and protein kinase C (PKC) activities and intracellular calcium. With the assumption that RGS2 has a higher affinity for Galpha(q) than Galpha(i/o), we demonstrated that the deactivation kinetics of GIRK channels can be differentially regulated by the relative amount of RGS proteins. These data indicate that transmitter-mediated deactivation of GIRK currents is modulated by crosstalk between G(i/o)- and G(q)-coupled pathways.

  5. 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.

  6. Cdk9 T-loop Phosphorylation is Regulated by the Calcium Signaling Pathway

    PubMed Central

    Ramakrishnan, Rajesh; Rice, Andrew P.

    2011-01-01

    Eukaryotic RNA polymerase II transcriptional elongation is a tightly regulated process and is dependent upon positive transcription elongation factor-b (P-TEFb). The core P-TEFb complex is composed of Cdk9 and Cyclin T and is essential for the expression of most protein coding genes. Cdk9 kinase function is dependent upon phosphorylation of Thr186 in its T-loop. In this study, we examined kinases and signaling pathways that influence Cdk9 T-loop phosphorylation. Using an RNAi screen in HeLa cells, we found that Cdk9 T-loop phosphorylation is regulated by Calcium/Calmodulin- dependent kinase 1D (CaMK1D). Using small molecules inhibitors in HeLa cells and primary CD4+ T lymphocytes, we found that the Ca2+ signaling pathway is required for Cdk9 T-loop phosphorylation. Inhibition of Ca2+ signaling led to dephosphorylation of Thr186 on Cdk9. In reporter plasmid assays, inhibition of the Ca2+ signaling pathway repressed the PCNA promoter and HIV-1 Tat transactivation of the HIV-1 LTR, but not HTLV-1 Tax transactivation of the HTLV-1 LTR, suggesting that perturbation of the Ca2+ pathway and reduction of Cdk9 T-loop phosphorylation inhibits transcription units that have a rigorous requirement for P-TEFb function. PMID:21448926

  7. ERK5 pathway regulates the phosphorylation of tumour suppressor hDlg during mitosis

    SciTech Connect

    Inesta-Vaquera, Francisco A.; Campbell, David G.; Arthur, J. Simon C.; Cuenda, Ana

    2010-08-13

    Research highlights: {yields} hDlg is phosphorylated during mitosis in multiple residues. {yields} Prospho-hDlg is excluded from the midbody during mitosis. {yields} hDlg is not phosphorylated by p38{gamma} or JNK1/2 during mitosis. {yields} ERK5 pathway mediates hDlg phosphorylation in mitosis. -- Abstract: Human disc-large (hDlg) is a scaffold protein critical for the maintenance of cell polarity and adhesion. hDlg is thought to be a tumour suppressor that regulates the cell cycle and proliferation. However, the mechanism and pathways involved in hDlg regulation during these processes is still unclear. Here we report that hDlg is phosphorylated during mitosis, and we establish the identity of at least three residues phosphorylated in hDlg; some are previously unreported. Phosphorylation affects hDlg localisation excluding it from the contact point between the two daughter cells. Our results reveal a previously unreported pathway for hDlg phosphorylation in mitosis and show that ERK5 pathway mediates hDlg cell cycle dependent phosphorylation. This is likely to have important implications in the correct timely mitotic entry and mitosis progression.

  8. Lovastatin inhibits the extracellular-signal-regulated kinase pathway in immortalized rat brain neuroblasts

    PubMed Central

    Cerezo-Guisado, Maria Isabel; GarcíA-Román, Natalia; García-MaríN, Luis Jesús; Álvarez-Barrientos, Alberto; Bragado, Maria Julia; Lorenzo, Maria Jesús

    2006-01-01

    We have shown previously that lovastatin, a 3-hydroxy-3-methyl- glutaryl coenzyme A reductase inhibitor, induces apoptosis in spontaneously immortalized rat brain neuroblasts. In the present study, we analysed the intracellular signal transduction pathways by which lovastatin induces neuroblast apoptosis. We showed that lovastatin efficiently inhibited Ras activation, which was associ-ated with a significant decrease in ERK1/2 (extracellular-signal-regulated kinase 1/2) phosphorylation. Lovastatin also decreased CREB phosphorylation and CREB-mediated gene expression. The effects of lovastatin on the Ras/ERK1/2/CREB pathway were time- and concentration-dependent and fully prevented by meva-lonate. In addition, we showed that two MEK [MAPK (mitogen-activated protein kinase)/ERK kinase] inhibitors, PD98059 and PD184352, were poor inducers of apoptosis in serum-treated neuroblasts. However, these inhibitors significantly increased apop-tosis induced by lovastatin treatment. Furthermore, we showed that pharmacological inhibition of both MEK and phosphoinos-itide 3-kinase activities was able to induce neuroblast apoptosis with similar efficacy as lovastatin. Our results suggest that lovast-atin triggers neuroblast apoptosis by regulating several signalling pathways, including the Ras/ERK1/2 pathway. These findings might also contribute to elucidate the intracellular mechanisms involved in the central nervous system side effects associated with statin therapy. PMID:16952276

  9. Signaling molecules and pathways regulating the fate of spermatogonial stem cells

    PubMed Central

    He, Zuping; Kokkinaki, Maria; Dym, Martin

    2009-01-01

    Spermatogenesis is the process that involves the division and differentiation of spermatogonial stem cells (SSCs) into mature spermatozoa. SSCs are a subpopulation of type A spermatogonia resting on the basement membrane in the mammalian testis. Self-renewal and differentiation of SSCs are the foundation of normal spermatogenesis, and thus a better understanding of molecular mechanisms and signaling pathways in the SSCs is of paramount importance for the regulation of spermatogenesis and may eventually lead to novel targets for male contraception as well as for gene therapy of male infertility and testicular cancer. Uncovering the molecular mechanisms is also of great interest to a better understanding of SSC aging and for developing novel therapeutic strategies for degenerative diseases in view of the recent work demonstrating the pluripotent potential of the SSC. Progress has recently been made in elucidating the signaling molecules and pathways that determine cell fate decisions of SSCs. In this review, we first address the morphological features, phenotypic characteristics, and the potential of SSCs. And then we focus on the recent advances in defining the key signaling molecules and crucial signaling pathways regulating self-renewal and differentiation of SSCs. The association of aberrant expression of signaling molecules and cascades with abnormal spermatogenesis and testicular cancer are also discussed. Finally we point out potential future directions to pursue in research on signaling pathways of SSCs. PMID:19263492

  10. Hypertensive stretch regulates endothelial exocytosis of Weibel-Palade bodies through VEGF receptor 2 signaling pathways.

    PubMed

    Xiong, Yan; Hu, Zhenqian; Han, Xiaofan; Jiang, Beibei; Zhang, Rongli; Zhang, Xiaoyu; Lu, Yao; Geng, Chenyang; Li, Wei; He, Yulong; Huo, Yingqing; Shibuya, Masabumi; Luo, Jincai

    2013-06-01

    Regulated endothelial exocytosis of Weibel-Palade bodies (WPBs), the first stage in leukocyte trafficking, plays a pivotal role in inflammation and injury. Acute mechanical stretch has been closely associated with vascular inflammation, although the precise mechanism is unknown. Here, we show that hypertensive stretch regulates the exocytosis of WPBs of endothelial cells (ECs) through VEGF receptor 2 (VEGFR2) signaling pathways. Stretch triggers a rapid release (within minutes) of von Willebrand factor and interleukin-8 from WPBs in cultured human ECs, promoting the interaction between leukocytes and ECs through the translocation of P-selectin to the cell membrane. We further show that hypertensive stretch significantly induces P-selectin translocation of intact ECs and enhances leukocyte adhesion both ex vivo and in vivo. Stretch-induced endothelial exocytosis is mediated via a VEGFR2/PLCγ1/calcium pathway. Interestingly, stretch also induces a negative feedback via a VEGFR2/Akt/nitric oxide pathway. Such dual effects are confirmed using pharmacological and genetic approaches in carotid artery segments, as well as in acute hypertensive mouse models. These studies reveal mechanical stretch as a potent agonist for endothelial exocytosis, which is modulated by VEGFR2 signaling. Thus, VEGFR2 signaling pathways may represent novel therapeutic targets in limiting hypertensive stretch-related inflammation.

  11. Cdk9 T-loop phosphorylation is regulated by the calcium signaling pathway.

    PubMed

    Ramakrishnan, Rajesh; Rice, Andrew P

    2012-02-01

    Eukaryotic RNA polymerase II transcriptional elongation is a tightly regulated process and is dependent upon positive transcription elongation factor-b (P-TEFb). The core P-TEFb complex is composed of Cdk9 and Cyclin T and is essential for the expression of most protein coding genes. Cdk9 kinase function is dependent upon phosphorylation of Thr186 in its T-loop. In this study, we examined kinases and signaling pathways that influence Cdk9 T-loop phosphorylation. Using an RNAi screen in HeLa cells, we found that Cdk9 T-loop phosphorylation is regulated by Ca(2+)/calmodulin-dependent kinase 1D (CaMK1D). Using small molecules inhibitors in HeLa cells and primary CD4(+) T lymphocytes, we found that the Ca(2+) signaling pathway is required for Cdk9 T-loop phosphorylation. Inhibition of Ca(2+) signaling led to dephosphorylation of Thr186 on Cdk9. In reporter plasmid assays, inhibition of the Ca(2+) signaling pathway repressed the PCNA promoter and HIV-1 Tat transactivation of the HIV-1 LTR, but not HTLV-1 Tax transactivation of the HTLV-1 LTR, suggesting that perturbation of the Ca(2+) pathway and reduction of Cdk9 T-loop phosphorylation inhibits transcription units that have a rigorous requirement for P-TEFb function.

  12. Cell cycle regulators interact with pathways that modulate microtubule stability in Saccharomyces cerevisiae.

    PubMed

    Shohat-Tal, Aya; Eshel, Dan

    2011-12-01

    The integrity of mitosis is dependent upon strict regulation of microtubule stability and dynamics. Although much information has been accumulated on regulators of the microtubule cytoskeleton, our knowledge of the specific pathways involved is still limited. Here we designed genetic screens to identify regulators of microtubule stability that are dispensable in the wild type yet become essential under microtubule-disrupting conditions. We found that the transcriptional cofactor Swi6p and activator Swi4p, as well as the G(2)/M-specific cyclin Clb2p, are required in a microtubule-destabilizing environment. Swi6p and Swi4p can combine as a transcriptional complex, called the SBF complex (SBF for Swi4/6 cell cycle box [SCB]-binding factor) that is functionally homologous to the metazoan DP1/2-E2F complex and that controls the G(1)/S transition through the genes it regulates. We show that Swi6p's contribution to microtubule stability can be either dependent or independent of the SBF complex. The SBF-dependent pathway requires downregulation of SBF complex levels and may thereby reroute the transcriptional program in favor of greater microtubule stability. This pathway can be triggered by overexpression of Fcp1p, a phosphatase in the general transcription machinery, or by expression of an allele of SWI6 that is associated with reduced transcription from SBF-controlled promoters. The SBF-independent pathway is activated by a constitutively nuclear allele of Swi6p. Our results introduce novel roles in microtubule stability for genes whose participation in the process may be masked under normal conditions yet nonetheless acquire a dominant role when microtubule stability is compromised.

  13. Mechanisms of JAK/STAT pathway negative regulation by the short coreceptor Eye Transformer/Latran.

    PubMed

    Fisher, Katherine H; Stec, Wojciech; Brown, Stephen; Zeidler, Martin P

    2016-02-01

    Transmembrane receptors interact with extracellular ligands to transduce intracellular signaling cascades, modulate target gene expression, and regulate processes such as proliferation, apoptosis, differentiation, and homeostasis. As a consequence, aberrant signaling events often underlie human disease. Whereas the vertebrate JAK/STAT signaling cascade is transduced via multiple receptor combinations, the Drosophila pathway has only one full-length signaling receptor, Domeless (Dome), and a single negatively acting receptor, Eye Transformer/Latran (Et/Lat). Here we investigate the molecular mechanisms underlying Et/Lat activity. We demonstrate that Et/Lat negatively regulates the JAK/STAT pathway activity and can bind to Dome, thus reducing Dome:Dome homodimerization by creating signaling-incompetent Dome:Et/Lat heterodimers. Surprisingly, we find that Et/Lat is able to bind to both JAK and STAT92E but, despite the presence of putative cytokine-binding motifs, does not detectably interact with pathway ligands. We find that Et/Lat is trafficked through the endocytic machinery for lysosomal degradation but at a much slower rate than Dome, a difference that may enhance its ability to sequester Dome into signaling-incompetent complexes. Our data offer new insights into the molecular mechanism and regulation of Et/Lat in Drosophila that may inform our understanding of how short receptors function in other organisms.

  14. [PPARγ up-regulates TGFβ/smad signal pathway repressor c-Ski].

    PubMed

    Li, Gong-bo; Li, Jun; Zeng, Yi-jun; Zhong, Dan; Wu, Geng-ze; Fu, Xiao-hong; He, Feng-tian; Dai, Shuang-shuang

    2011-02-25

    TGFβ/smad pathway is recognized as an important signal pathway to promote the pathogenesis of atherosclerosis (AS). Peroxisome proliferator-activated receptor γ (PPARγ) activation is considered to be important in modulating AS. Herein, we investigated the regulation of PPARγ on c-Ski, the repressor of TGFβ/smad pathway, in rat AS model and cultured vascular smooth muscle cells (VSMCs). c-Ski mRNA and protein expression were detected by real-time PCR and Western blot, respectively, in vivo and in vitro with treatment of PPARγ agonist rosiglitazone and antagonist GW9662. The proliferation and collagen secretion of VSMCs after c-Ski transfection were investigated. The underlying mechanism was further investigated by online program NUBIScan and luciferase reporter gene analysis. Results showed that both mRNA and protein expressions of c-Ski in the AS lesions was down-regulated in vivo, while in cultured VSMCs, c-Ski transfection significantly suppressed the proliferation and collagen secretion of rat VSMCs. Rosiglitazone significantly up-regulated mRNA and protein levels of c-Ski in VSMCs, which could be blocked by GW9662. Online NUBIScan analysis suggested possible PPARγ binding sites in the promoter region of c-Ski. In addition, luciferase activity of c-Ski reporter gene was also increased obviously in the presence of rosiglitazone. These results indicate that c-Ski is one of the newly found target genes of PPARγ and thus involved in the anti-AS effect of PPARγ.

  15. Divergent androgen regulation of unfolded protein response pathways drives prostate cancer

    PubMed Central

    Sheng, Xia; Arnoldussen, Yke Jildouw; Storm, Margrethe; Tesikova, Martina; Nenseth, Hatice Zeynep; Zhao, Sen; Fazli, Ladan; Rennie, Paul; Risberg, Bjørn; Wæhre, Håkon; Danielsen, Håvard; Mills, Ian G; Jin, Yang; Hotamisligil, Gökhan; Saatcioglu, Fahri

    2015-01-01

    The unfolded protein response (UPR) is a homeostatic mechanism to maintain endoplasmic reticulum (ER) function. The UPR is activated by various physiological conditions as well as in disease states, such as cancer. As androgens regulate secretion and development of the normal prostate and drive prostate cancer (PCa) growth, they may affect UPR pathways. Here, we show that the canonical UPR pathways are directly and divergently regulated by androgens in PCa cells, through the androgen receptor (AR), which is critical for PCa survival. AR bound to gene regulatory sites and activated the IRE1α branch, but simultaneously inhibited PERK signaling. Inhibition of the IRE1α arm profoundly reduced PCa cell growth in vitro as well as tumor formation in preclinical models of PCa in vivo. Consistently, AR and UPR gene expression were correlated in human PCa, and spliced XBP-1 expression was significantly upregulated in cancer compared with normal prostate. These data establish a genetic switch orchestrated by AR that divergently regulates the UPR pathways and suggest that targeting IRE1α signaling may have therapeutic utility in PCa. PMID:25864123

  16. A GRHL3-regulated repair pathway suppresses immune-mediated epidermal hyperplasia.

    PubMed

    Gordon, William M; Zeller, Michael D; Klein, Rachel H; Swindell, William R; Ho, Hsiang; Espetia, Francisco; Gudjonsson, Johann E; Baldi, Pierre F; Andersen, Bogi

    2014-12-01

    Dermal infiltration of T cells is an important step in the onset and progression of immune-mediated skin diseases such as psoriasis; however, it is not known whether epidermal factors play a primary role in the development of these diseases. Here, we determined that the prodifferentiation transcription factor grainyhead-like 3 (GRHL3), which is essential during epidermal development, is dispensable for adult skin homeostasis, but required for barrier repair after adult epidermal injury. Consistent with activation of a GRHL3-regulated repair pathway in psoriasis, we found that GRHL3 is upregulated in lesional skin and binds known epidermal differentiation gene targets. Using an imiquimod-induced model of immune-mediated epidermal hyperplasia, we found that mice lacking GRHL3 have an exacerbated epidermal damage response, greater sensitivity to disease induction, delayed resolution of epidermal lesions, and resistance to anti-IL-22 therapy compared with WT animals. ChIP-Seq and gene expression profiling of murine skin revealed that while GRHL3 regulates differentiation pathways both during development and during repair from immune-mediated damage, it targets distinct sets of genes in the 2 processes. In particular, GRHL3 suppressed a number of alarmin and other proinflammatory genes after immune injury. This study identifies a GRHL3-regulated epidermal barrier repair pathway that suppresses disease initiation and helps resolve existing lesions in immune-mediated epidermal hyperplasia.

  17. Spn1 Regulates the GNBP3-Dependent Toll Signaling Pathway in Drosophila melanogaster▿

    PubMed Central

    Fullaondo, Ane; García-Sánchez, Susana; Sanz-Parra, Arantza; Recio, Emma; Lee, So Young; Gubb, David

    2011-01-01

    The Drosophila genome encodes 29 serpins, most of unknown function. We show here that Spn1 is an active protease inhibitor of the serpin superfamily. Spn1 inhibits trypsin in vitro and regulates the Toll-mediated immune response in vivo. Expression of the Toll-dependent transcripts Drosomycin and IM1 is increased in Spn1 null mutants. Overexpression of Spn1 reduces the induction of Drosomycin upon immune challenge with fungi but not Gram-positive bacteria. Similar reductions in Drosomycin levels are observed in the psh, spz, and grass mutants of the Toll signaling pathway. These results support a role of Spn1 as a repressor of Toll activation upon fungal infection. Epistatic analysis places Spn1 upstream of Spätzle processing enzyme and Grass, in the fungal cell wall-activated side branch of the pathway. Overexpression of the pattern recognition receptor GNBP3 activates the β-1,3-glucan-sensitive side branch of the Toll pathway. The resultant increased Drosomycin level is reduced by concomitant overexpression of Spn1, confirming that Spn1 regulates the fungal cell wall side branch. Spn1 null mutants show altered susceptibility to fungal infection compared to the wild type, demonstrating a requirement for Spn1 in the fine regulation of the immune response. PMID:21576362

  18. A GRHL3-regulated repair pathway suppresses immune-mediated epidermal hyperplasia

    PubMed Central

    Gordon, William M.; Zeller, Michael D.; Klein, Rachel H.; Swindell, William R.; Ho, Hsiang; Espetia, Francisco; Gudjonsson, Johann E.; Baldi, Pierre F.; Andersen, Bogi

    2014-01-01

    Dermal infiltration of T cells is an important step in the onset and progression of immune-mediated skin diseases such as psoriasis; however, it is not known whether epidermal factors play a primary role in the development of these diseases. Here, we determined that the prodifferentiation transcription factor grainyhead-like 3 (GRHL3), which is essential during epidermal development, is dispensable for adult skin homeostasis, but required for barrier repair after adult epidermal injury. Consistent with activation of a GRHL3-regulated repair pathway in psoriasis, we found that GRHL3 is upregulated in lesional skin and binds known epidermal differentiation gene targets. Using an imiquimod-induced model of immune-mediated epidermal hyperplasia, we found that mice lacking GRHL3 have an exacerbated epidermal damage response, greater sensitivity to disease induction, delayed resolution of epidermal lesions, and resistance to anti–IL-22 therapy compared with WT animals. ChIP-Seq and gene expression profiling of murine skin revealed that while GRHL3 regulates differentiation pathways both during development and during repair from immune-mediated damage, it targets distinct sets of genes in the 2 processes. In particular, GRHL3 suppressed a number of alarmin and other proinflammatory genes after immune injury. This study identifies a GRHL3-regulated epidermal barrier repair pathway that suppresses disease initiation and helps resolve existing lesions in immune-mediated epidermal hyperplasia. PMID:25347468

  19. Mutations modulating the Argos-regulated signaling pathway in Drosophila eye development.

    PubMed

    Taguchi, A; Sawamoto, K; Okano, H

    2000-04-01

    Argos is a secreted protein that contains an EGF-like domain and acts as an inhibitor of Drosophila EGF receptor activation. To identify genes that function in the Argos-regulated signaling pathway, we performed a genetic screen for enhancers and suppressors of the eye phenotype caused by the overexpression of argos. As a result, new alleles of known genes encoding components of the EGF receptor pathway, such as Star, sprouty, bulge, and clown, were isolated. To study the role of clown in development, we examined the eye and wing phenotypes of the clown mutants in detail. In the eye discs of clown mutants, the pattern of neuronal differentiation was impaired, showing a phenotype similar to those caused by a gain-of-function EGF receptor mutation and overexpression of secreted Spitz, an activating ligand for the EGF receptor. There was also an increased number of pigment cells in the clown eyes. Epistatic analysis placed clown between argos and Ras1. In addition, we found that clown negatively regulated the development of wing veins. These results suggest that the clown gene product is important for the Argos-mediated inhibition of EGF receptor activation during the development of various tissues. In addition to the known genes, we identified six mutations of novel genes. Genetic characterization of these mutants suggested that they have distinct roles in cell differentiation and/or survival regulated by the EGF receptor pathway.

  20. Emerging Molecular Pathways Governing Dietary Regulation of Neural Stem Cells during Aging

    PubMed Central

    de Lucia, Chiara; Murphy, Tytus; Thuret, Sandrine

    2017-01-01

    Aging alters cellular and molecular processes, including those of stem cells biology. In particular, changes in neural stem cells (NSCs) are linked to cognitive decline associated with aging. Recently, the systemic environment has been shown to alter both NSCs regulation and age-related cognitive decline. Interestingly, a well-documented and naturally occurring way of altering the composition of the systemic environment is through diet and nutrition. Furthermore, it is well established that the presence of specific nutrients as well as the overall increase or reduction of calorie intake can modulate conserved molecular pathways and respectively reduce or increase lifespan. In this review, we examine these pathways in relation to their function on NSCs and cognitive aging. We highlight the importance of the Sirtuin, mTOR and Insulin/Insulin like growth factor-1 pathways as well as the significant role played by epigenetics in the dietary regulation of NSCs and the need for further research to exploit nutrition as a mode of intervention to regulate NSCs aging. PMID:28194114

  1. TGF-beta and TNF-a affect cell surface proteoglycan and sialic acid expression on vascular endothelial cells.

    PubMed

    Doiron, Amber L; Kirkpatrick, Allison P; Rinker, Kristina D

    2004-01-01

    Atherosclerosis is the formation of plaques in the arterial wall brought about by numerous events including the accumulation of oxidized low density lipoprotein (LDL), stimulation of inflammatory responses, the release of cytokines, and the attachment of monocytes to the arterial wall. Proteoglycans are implicated in many aspects of atherosclerosis including the metabolism of lipoproteins, regulation of cytokine activity, cell adhesion, and modification of the extracellular matrix. Due to their complex role in molecular recognition and cellular adhesion, the glycosaminoglycan (GAG) chains attached to the proteoglycan core and sialic acids on the terminal ends of the glycan chains are of interest. This study investigated the effects of exposure to transforming growth factor-beta 1 (TGF-beta 1) and tumor necrosis factor-a (TNF-a) on the expression of cell surface GAGs and sialic acids on human umbilical vein endothelial cells (HUVECs). Initial results show that TGF-beta 1 affected GAG expression compared to a control condition. Results also show that the combination of TGF-beta 1 and TNF-a affected GAG expression differently than does TGF-beta 1 alone. Additionally, TNF-a decreased the number of sialic acid residues per cell and TGF-beta 1 slightly upregulated sialic acid expression as compared to the control. The combination of the two cytokines showed a larger upward trend in this value. These data indicate that TNF-a and TGF-beta 1 play a role in the expression of GAG chains and sialic acids on the cell surface. Further study may clarify the implications of these findings for atherosclerosis.

  2. HIF-1α pathway: role, regulation and intervention for cancer therapy

    PubMed Central

    Masoud, Georgina N.; Li, Wei

    2015-01-01

    Hypoxia-inducible factor-1 (HIF-1) has been recognized as an important cancer drug target. Many recent studies have provided convincing evidences of strong correlation between elevated levels of HIF-1 and tumor metastasis, angiogenesis, poor patient prognosis as well as tumor resistance therapy. It was found that hypoxia (low O2 levels) is a common character in many types of solid tumors. As an adaptive response to hypoxic stress, hypoxic tumor cells activate several survival pathways to carry out their essential biological processes in different ways compared with normal cells. Recent advances in cancer biology at the cellular and molecular levels highlighted the HIF-1α pathway as a crucial survival pathway for which novel strategies of cancer therapy could be developed. However, targeting the HIF-1α pathway has been a challenging but promising progresses have been made in the past twenty years. This review summarizes the role and regulation of the HIF-1α in cancer, and recent therapeutic approaches targeting this important pathway. PMID:26579469

  3. Targeting the TLR9-MyD88 pathway in the regulation of adaptive immune responses

    PubMed Central

    Huang, Xiaopei; Yang, Yiping

    2010-01-01

    IMPORTANCE OF THE FIELD Toll-like receptors (TLRs) are innate immune receptors critical in the innate immune defense against invading pathogens. Recent advances also reveal a crucial role for TLRs in shaping adaptive immune responses, conferring a potential therapeutic value to their modulation in the treatment of diseases. AREAS COVERED IN THIS REVIEW The aim of this review is to discuss TLR9, the TLR9-MyD88 signaling pathway and its role in regulation of adaptive immune responses, as well as potential therapeutic implications by targeting this pathway. WHAT THE READER WILL GAIN This review shows that the TLR9-MyD88 signaling pathway plays a critical role in promoting adaptive immune responses and that modulation of this pathway may have enormous therapeutic potential in enhancing vaccine potency, controlling autoimmunity, as well as improving the outcome of viral vector-mediated gene therapy. TAKE HOME MESSAGE Although TLR9 agonists have been used as adjuvants for enhancing vaccine potency, further exploitation of the TLR9-MyD88 pathway and its dynamic interaction with the immune system in vivo is needed to provide more effective therapeutic inventions in the design of vaccines for infectious diseases, allergies and cancer, in the control of autoimmunity, as well as in the improvement of viral vector-mediated gene therapy. PMID:20560798

  4. Transcriptional regulators in the Hippo signaling pathway control organ growth in Xenopus tadpole tail regeneration.

    PubMed

    Hayashi, Shinichi; Ochi, Haruki; Ogino, Hajime; Kawasumi, Aiko; Kamei, Yasuhiro; Tamura, Koji; Yokoyama, Hitoshi

    2014-12-01

    The size and shape of tissues are tightly controlled by synchronized processes among cells and tissues to produce an integrated organ. The Hippo signaling pathway controls both cell proliferation and apoptosis by dual signal-transduction states regulated through a repressive kinase cascade. Yap1 and Tead, transcriptional regulators that act downstream of the Hippo signaling kinase cascade, have essential roles in regulating cell proliferation. In amphibian limb or tail regeneration, the local tissue outgrowth terminates when the correct size is reached, suggesting that organ size is strictly controlled during epimorphic organ-level regeneration. We recently demonstrated that Yap1 is required for the regeneration of Xenopus tadpole limb buds (Hayashi et al., 2014, Dev. Biol. 388, 57-67), but the molecular link between the Hippo pathway and organ size control in vertebrate epimorphic regeneration is not fully understood. To examine the requirement of Hippo pathway transcriptional regulators in epimorphic regeneration, including organ size control, we inhibited these regulators during Xenopus tadpole tail regeneration by overexpressing a dominant-negative form of Yap (dnYap) or Tead4 (dnTead4) under a heat-shock promoter in transgenic animal lines. Each inhibition resulted in regeneration defects accompanied by reduced cell mitosis and increased apoptosis. Single-cell gene manipulation experiments indicated that Tead4 cell-autonomously regulates the survival of neural progenitor cells in the regenerating tail. In amphibians, amputation at the proximal level of the tail (deep amputation) results in faster regeneration than that at the distal level (shallow amputation), to restore the original-sized tail with similar timing. However, dnTead4 overexpression abolished the position-dependent differential growth rate of tail regeneration. These results suggest that the transcriptional regulators in the Hippo pathway, Tead4 and Yap1, are required for general vertebrate

  5. Sugar Allocation to Metabolic Pathways is Tightly Regulated and Affects the Virulence of Streptococcus mutans

    PubMed Central

    Kawada-Matsuo, Miki; Oogai, Yuichi; Komatsuzawa, Hitoshi

    2016-01-01

    Bacteria take up and metabolize sugar as a carbohydrate source for survival. Most bacteria can utilize many sugars, including glucose, sucrose, and galactose, as well as amino sugars, such as glucosamine and N-acetylglucosamine. After entering the cytoplasm, the sugars are mainly allocated to the glycolysis pathway (energy production) and to various bacterial component biosynthesis pathways, including the cell wall, nucleic acids and amino acids. Sugars are also utilized to produce several virulence factors, such as capsule and lipoteichoic acid. Glutamine-fructose-6-phosphate aminotransferase (GlmS) and glucosamine-6-phosphate deaminase (NagB) have crucial roles in sugar distribution to the glycolysis pathway and to cell wall biosynthesis. In Streptococcus mutans, a cariogenic pathogen, the expression levels of glmS and nagB are coordinately regulated in response to the presence or absence of amino sugars. In addition, the disruption of this regulation affects the virulence of S. mutans. The expression of nagB and glmS is regulated by NagR in S. mutans, but the precise mechanism underlying glmS regulation is not clear. In Staphylococcus aureus and Bacillus subtilis, the mRNA of glmS has ribozyme activity and undergoes self-degradation at the mRNA level. However, there is no ribozyme activity region on glmS mRNA in S. mutans. In this review article, we summarize the sugar distribution, particularly the coordinated regulation of GlmS and NagB expression, and its relationship with the virulence of S. mutans. PMID:28036052

  6. Circadian regulation of the hepatic endobiotic and xenobitoic detoxification pathways: the time matters.

    PubMed

    Zmrzljak, Ursula Prosenc; Rozman, Damjana

    2012-04-16

    Metabolic processes have to be regulated tightly to prevent waste of energy and to ensure sufficient detoxification. Most anabolic processes operate in a timely manner when energy intake is the highest, while catabolism takes place in energy spending periods. Endobiotic and xenobiotic metabolism are therefore under circadian control. Circadian regulation is mediated through the suprachiasmatic nucleus (SCN), a master autonomous oscillator of the brain. Although many peripheral organs have their own oscillators, the SCN is important in orchestrating and entraining organs according to the environmental light cues. However, light is not the only signal for entrainment of internal clocks. For endobiotic and xenobitoic detoxification pathways, the food composition and intake regime are equally important. The rhythm of the liver as an organ where the major metabolic pathways intersect depends on SCN signals, signals from endocrine tissues, and, importantly, the type and time of feeding or xenobiotics ingestion. Several enzymes are involved in detoxification processes. Phase I is composed mainly of cytochromes P450, which are regulated by nuclear receptors. Phase II enzymes modify the phase I metabolites, while phase III includes membrane transporters responsible for the elimination of modified xenobiotics. Phases I-III of drug metabolism are under strong circadian regulation, starting with the drug-sensing nuclear receptors and ending with drug transporters. Disturbed circadian regualtion (jet-lag, shift work, and dysfunction of core clock genes) leads to changed periods of activity, sleep disorders, disturbed glucose homeostasis, breast or colon cancer, and metabolic syndrome. As many xenobiotics influence the circadian rhythm of the liver, bad drug administration timing can worsen the above listed effects. This review will cover the major hepatic circadian regulation of endogenous and xenobiotic metabolic pathways and will provide examples of how good timing of drug

  7. WDR26 in Advanced Breast Cancer: A Novel Regulator of the P13K/AKT Pathway

    DTIC Science & Technology

    2015-10-01

    AWARD NUMBER: W81XWH-14-1-0539 TITLE: WDR26 in Advanced Breast Cancer : A Novel Regulator of the P13K/AKT Pathway PRINCIPAL INVESTIGATOR...5a. CONTRACT NUMBER WDR26 in Advanced Breast Cancer : A Novel Regulator of the P13K/AKT Pathway 5b. GRANT NUMBER W81XWH-14-1-0539 5c. PROGRAM...SUPPLEMENTARY NOTES 14. ABSTRACT The PI3K/AKT pathway is one of the most deregulated pathways in breast cancers (>70%), and a major contributor to tumor

  8. The silent information regulator 1 (Sirt1) is a positive regulator of the Notch pathway in Drosophila.

    PubMed

    Horvath, Matej; Mihajlovic, Zorana; Slaninova, Vera; Perez-Gomez, Raquel; Moshkin, Yuri; Krejci, Alena

    2016-11-15

    The silent information regulator 1 (Sirt1) has been shown to have negative effects on the Notch pathway in several contexts. We bring evidence that Sirt1 has a positive effect on Notch activation in Drosophila, in the context of sensory organ precursor specification and during wing development. The phenotype of Sirt1 mutant resembles weak Notch loss-of-function phenotypes, and genetic interactions of Sirt1 with the components of the Notch pathway also suggest a positive role for Sirt1 in Notch signalling. Sirt1 is necessary for the efficient activation of enhancer of split [E(spl)] genes by Notch in S2N cells. Additionally, the Notch-dependent response of several E(spl) genes is sensitive to metabolic stress caused by 2-deoxy-d-glucose treatment, in a Sirt1-dependent manner. We found Sirt1 associated with several proteins involved in Notch repression as well as activation, including the cofactor exchange factor Ebi (TBL1), the RLAF/LAF histone chaperone complex and the Tip60 acetylation complex. Moreover, Sirt1 participates in the deacetylation of the CSL transcription factor Suppressor of Hairless. The role of Sirt1 in Notch signalling is, therefore, more complex than previously recognized, and its diverse effects may be explained by a plethora of Sirt1 substrates involved in the regulation of Notch signalling.

  9. Deubiquitylating enzyme USP9x regulates hippo pathway activity by controlling angiomotin protein turnover

    PubMed Central

    Thanh Nguyen, Hung; Andrejeva, Diana; Gupta, Rajat; Choudhary, Chunaram; Hong, Xin; Eichhorn, Pieter J A; Loya, Anand C; Cohen, Stephen M

    2016-01-01

    The Hippo pathway has been identified as a key barrier for tumorigenesis, acting through downregulation of YAP/TAZ activity. Elevated YAP/TAZ activity has been documented in many human cancers. Ubiquitylation has been shown to play a key role in regulating YAP/TAZ activity through downregulation of a number of Hippo pathway components. Several ubiquitin ligase complexes have been implicated in this process, however, little is known about the deubiquitylating enzymes that counteract these activities to regulate YAP/TAZ. Here we identify the deubiquitylating enzyme USP9x as a regulator of YAP/TAZ activity. We demonstrate that USPx regulates ubiquitin-mediated turnover of the YAP inhibitor, Angiomotin. USP9x acts to deubiquitylate Angiomotin at lysine 496, resulting in stabilization of Angiomotin and lower YAP/TAZ activity. USP9x mRNA levels were reduced in several cancers. Clinically, USP9x mRNA levels were reduced in several cancers with low USPx expression correlating with poor prognosis in renal clear cell carcinoma. Our data indicate that USP9x may be a useful biomarker for renal clear cell carcinoma. PMID:27462448

  10. SLOB, a SLOWPOKE Channel Binding Protein, Regulates Insulin Pathway Signaling and Metabolism in Drosophila

    PubMed Central

    Sheldon, Amanda L.; Zhang, Jiaming; Fei, Hong; Levitan, Irwin B.

    2011-01-01

    There is ample evidence that ion channel modulation by accessory proteins within a macromolecular complex can regulate channel activity and thereby impact neuronal excitability. However, the downstream consequences of ion channel modulation remain largely undetermined. The Drosophila melanogaster large conductance calcium-activated potassium channel SLOWPOKE (SLO) undergoes modulation via its binding partner SLO-binding protein (SLOB). Regulation of SLO by SLOB influences the voltage dependence of SLO activation and modulates synaptic transmission. SLO and SLOB are expressed especially prominently in median neurosecretory cells (mNSCs) in the pars intercerebralis (PI) region of the brain; these cells also express and secrete Drosophila insulin like peptides (dILPs). Previously, we found that flies lacking SLOB exhibit increased resistance to starvation, and we reasoned that SLOB may regulate aspects of insulin signaling and metabolism. Here we investigate the role of SLOB in metabolism and find that slob null flies exhibit changes in energy storage and insulin pathway signaling. In addition, slob null flies have decreased levels of dilp3 and increased levels of takeout, a gene known to be involved in feeding and metabolism. Targeted expression of SLOB to mNSCs rescues these alterations in gene expression, as well as the metabolic phenotypes. Analysis of fly lines mutant for both slob and slo indicate that the effect of SLOB on metabolism and gene expression is via SLO. We propose that modulation of SLO by SLOB regulates neurotransmission in mNSCs, influencing downstream insulin pathway signaling and metabolism. PMID:21850269

  11. The TSC-mTOR signaling pathway regulates the innate inflammatory response.

    PubMed

    Weichhart, Thomas; Costantino, Giuseppina; Poglitsch, Marko; Rosner, Margit; Zeyda, Maximilian; Stuhlmeier, Karl M; Kolbe, Thomas; Stulnig, Thomas M; Hörl, Walter H; Hengstschläger, Markus; Müller, Mathias; Säemann, Marcus D

    2008-10-17

    The innate inflammatory immune response must be tightly controlled to avoid damage to the host. Here, we showed that the tuberous sclerosis complex-mammalian target of rapamycin (TSC-mTOR) pathway regulated inflammatory responses after bacterial stimulation in monocytes, macrophages, and primary dendritic cells. Inhibition of mTOR by rapamycin promoted production of proinflammatory cytokines via the transcription factor NF-kappaB but blocked the release of interleukin-10 via the transcription factor STAT3. Conversely, deletion of TSC2, the key negative regulator of mTOR, diminished NF-kappaB but enhanced STAT3 activity and reversed this proinflammatory cytokine shift. Rapamycin-hyperactivated monocytes displayed a strong T helper 1 (Th1) cell- and Th17 cell-polarizing potency. Inhibition of mTOR in vivo regulated the inflammatory response and protected genetically susceptible mice against lethal Listeria monocytogenes infection. These data identify the TSC2-mTOR pathway as a key regulator of innate immune homeostasis with broad clinical implications for infectious and autoimmune diseases, vaccination, cancer, and transplantation.

  12. Time-course regulation of survival pathways by epicatechin on HepG2 cells.

    PubMed

    Granado-Serrano, Ana Belén; Angeles Martín, María; Goya, Luis; Bravo, Laura; Ramos, Sonia

    2009-02-01

    Polyphenols, such as epicatechin, have been reported to exhibit a wide range of biological activities. The objective of the present study was to investigate the time-dependent regulation by epicatechin of survival/proliferation pathways in HepG2 cells. Treatment of HepG2 cells with 10 micromol/L epicatechin did not result in any cell damage up to 18 h, as evaluated by the lactate dehydrogenase assay. Moreover, the enhanced cell death evoked by an oxidative stress induced with tert-butyl hydroperoxide was prevented in the cells pretreated 4 or 18 h with epicatechin. Epicatechin-induced survival was a rapid event that was accompanied by early and sustained activation of major survival signaling proteins, such as AKT/phosphatidylinositol 3-kinase and extracellular-regulated kinase (activated from 5 min to 18 h), as well as protein kinase C (PKC)-alpha (30 min to 18 h), in concert with unaltered c-jun N-amino terminal kinase levels and early inactivation of key death-related signals like PKC-delta (5 min to 18 h). Additionally, reactive oxygen species generation was transiently reduced when cells were treated with 10 micromol/L epicatechin (15-240 min). These data suggest that epicatechin induces cellular survival through a tight regulation of survival/proliferation pathways that requires the integration of different signals and persists over time, the ultimate effect on HepG2 cells being regulated by the balance among these signals.

  13. Merlin inhibits growth hormone-regulated Raf-ERKs pathways by binding to Grb2 protein

    SciTech Connect

    Lim, Jung Yeon; Kim, Hongtae; Jeun, Sin-Soo . E-mail: ssjeun@catholic.ac.kr; Kang, Seok-Gu; Lee, Kyung-Jin

    2006-02-24

    Numerous studies have suggested that the NF2 protein merlin is involved in the regulation of abnormal cell growth and proliferation. In this study, to better understand the merlin's mechanisms that contribute to the inhibition of tumorigenesis, we examined the potential action of merlin on the cell proliferative signaling pathways in response to growth hormone (GH). Merlin effectively attenuated the GH-induced serum response element (SRE) and Elk-1-mediated transcriptional activation, as well as the endogenous SRE-regulated gene c-fos expression in NIH3T3 cells. In addition, merlin prevented the Raf-1 complex activation process, which resulted in the suppression of MAP kinase/ERK, extracellular signal-regulated kinase (ERKs), and Elk-1 phosphorylation, which are the downstream signals of Raf-1. Moreover, it was shown that merlin interacted with endogenous growth factor receptor bound 2 (Grb2) protein and inhibited its expression. These results suggest that merlin contributes, via its protein-to-protein interaction with Grb2 and consequent inhibition of the MAPK pathways, to the regulation of the abnormal cell proliferation, and this provides a further mechanism underlying the tumor suppressor function of merlin.

  14. Pebble/ECT2 RhoGEF negatively regulates the Wingless/Wnt signaling pathway.

    PubMed

    Greer, Elisabeth R; Chao, Anna T; Bejsovec, Amy

    2013-12-01

    Wingless (Wg)/Wnt signaling is essential for patterning invertebrate and vertebrate embryos, and inappropriate Wnt activity is associated with a variety of human cancers. Despite intensive study, Wnt pathway mechanisms are not fully understood. We have discovered a new mechanism for regulating the Wnt pathway: activity of a Rho guanine nucleotide exchange factor (GEF) encoded by pebble (pbl) in Drosophila and ECT2 in humans. This RhoGEF has an essential role in cytokinesis, but also plays an unexpected, conserved role in inhibiting Wg/Wnt activity. Loss and gain of pbl function in Drosophila embryos cause pattern defects that indicate altered Wg activity. Both Pbl and ECT2 repress Wg/Wnt target gene expression in cultured Drosophila and human cells. The GEF activity is required for Wnt regulation, whereas other protein domains important for cytokinesis are not. Unlike most negative regulators of Wnt activity, Pbl/ECT2 functions downstream of Armadillo (Arm)/beta-catenin stabilization. Our results indicate GTPase regulation at a novel point in Wg/Wnt signal transduction, and provide new insight into the categorization of ECT2 as a human proto-oncogene.

  15. Mechanistic relationship between the vagal afferent pathway, central nervous system and peripheral organs in appetite regulation.

    PubMed

    Ueno, Hiroaki; Nakazato, Masamitsu

    2016-11-01

    The hypothalamus is a center of food intake and energy metabolism regulation. Information signals from peripheral organs are mediated through the circulation or the vagal afferent pathway and input into the hypothalamus, where signals are integrated to determine various behaviors, such as eating. Numerous appetite-regulating peptides are expressed in the central nervous system and the peripheral organs, and interact in a complex manner. Of such peptides, gut peptides are known to bind to receptors at the vagal afferent pathway terminal that extend into the mucosal layer of the digestive tract, modulate the electrical activity of the vagus nerve, and subsequently send signals to the solitary nucleus and furthermore to the hypothalamus. All peripheral peptides other than ghrelin suppress appetite, and they synergistically suppress appetite through the vagus nerve. In contrast, the appetite-enhancing peptide, ghrelin, antagonizes the actions of appetite-suppressing peptides through the vagus nerve, and appetite-suppressing peptides have attenuated effects in obesity as a result of inflammation in the vagus nerve. With greater understanding of the mechanism for food intake and energy metabolism regulation, medications that apply the effects of appetite-regulating peptides or implantable devices that electrically stimulate the vagus nerve are being investigated as novel treatments for obesity in basic and clinical studies.

  16. Regulation of monocarboxylate transporter 1 in skeletal muscle cells by intracellular signaling pathways.

    PubMed

    Narumi, Katsuya; Furugen, Ayako; Kobayashi, Masaki; Otake, Sho; Itagaki, Shirou; Iseki, Ken

    2010-01-01

    Skeletal muscle is the major producer of lactic acid in the body, but its oxidative fibers also use lactic acid as a respiratory fuel. Monocarboxylate transporter (MCT) 1 has been suggested to play a major role in influx of L-lactic acid for oxidation. The regulation mechanism of MCT1 was characterized utilizing rhabdomyosarcoma cells as an in vitro skeletal muscle model. The uptake of L-lactic acid via MCT1 was studied in the presence of various intracellular regulatory pathways, including pathways mediated by protein kinases A, C and G (PKA, PKC and PKG), protein tyrosine kinase (PTK), and Ca2+/calmodulin modulators. The results showed that PKG-, PTK-, and Ca2+/calmodulin-mediated regulatory pathways play no role in the regulation of L-lactic acid uptake, but a role for PKC- and PKA-mediated pathways was apparent. Uptake of L-lactic acid appeared to be stimulated by phorbol 12-myristate 13-acetate (PMA, a PKC activator) via an increase in Vmax of transport processes with no alteration in Km. In parallel, PMA treatment also resulted in an increase in the level of MCT1 expression. On the other hand, exposure to 8-Br-cAMP, a cAMP analog, and to forskolin, an adenylyl cyclase activator, resulted in a significant decrease in L-lactic acid uptake. Additionally, 8-Br-cAMP reduced Vmax but not Km values. Parallel to the decrease in Vmax of L-lactic acid uptake, the level of MCT1 expression was decreased in response to incubation with 8-Br-cAMP. These results indicate the possible involvement of a PKC- and PKA-mediated pathway associated with expression of MCT1 and lactate transport.

  17. Phosphatidylinositol 3-kinase pathway regulates sperm viability but not capacitation on boar spermatozoa.

    PubMed

    Aparicio, I M; Bragado, M J; Gil, M C; Garcia-Herreros, M; Gonzalez-Fernandez, L; Tapia, J A; Garcia-Marin, L J

    2007-08-01

    Phosphatidylinositol 3-kinase (PI3-K) plays an important role in cell survival in somatic cells and recent data pointed out a role for this kinase in sperm capacitation and acrosome reaction (AR). This study was undertaken to evaluate the role of PI3-K pathway on porcine spermatozoa capacitation, AR, and viability using two unrelated PI3-K inhibitors, LY294002 and wortmannin. In boar spermatozoa, we have identified the presence of PDK1, PKB/Akt, and PTEN, three of the main key components of the PI3-K pathway. Incubation of boar sperm in a capacitating medium (TCM) caused a significant increase in the percentage of capacitated (25 +/- 2 to 34 +/- 1% P < 0.05, n = 6) and acrosome reacted (1 +/- 1 to 11 +/- 1% P < 0.01, n = 6) spermatozoa compared with sperm in basal medium (TBM). Inhibition of PI3-K did affect neither the capacitation status nor AR nor protein p32 tyrosine phosphorylation of boar spermatozoa incubated in TBM or TCM. Boar sperm viability in TBM was significantly decreased by 40 and 20% after pretreatment with LY294002 or wortmannin, respectively. Similar results were observed after incubation of boar spermatozoa in TCM. Treatment of boar spermatozoa with the analog of cAMP, 8Br-cAMP significantly prevented the reduction on sperm viability. Our results provide evidence for an important role of the PI3-K pathway in the regulation of boar sperm viability and suggests that other signaling pathways different from PI3-K must be activated downstream of cAMP to contribute to regulation of sperm viability. Finally, in our conditions the PI3-K pathway seems not related with boar sperm capacitation or AR.

  18. The mitochondria-regulated death pathway mediates asbestos-induced alveolar epithelial cell apoptosis.

    PubMed

    Panduri, Vijayalakshmi; Weitzman, Sigmund A; Chandel, Navdeep; Kamp, David W

    2003-02-01

    The mechanisms underlying asbestos-induced pulmonary toxicity are not fully understood. Alveolar epithelial cell (AEC) apoptosis by iron-derived reactive oxygen species (ROS) is one important mechanism implicated. The two major pathways regulating apoptosis include (i) the mitochondrial death (intrinsic) pathway caused by DNA damage, and (ii) the plasma-membrane death receptor (extrinsic) pathway. However, it is unknown whether asbestos activates either death pathway in AEC. We determined whether asbestos triggers AEC mitochondrial dysfunction by exposing cells (A549 and rat alveolar type II) to amosite asbestos and assessing mitochondrial membrane potential changes (deltapsi(m)) using a fluorometric technique involving tetremethylrhodamine ethyl ester (TMRE) and mitotracker green. Unlike inert particulates (titanium dioxide and glass beads), amosite asbestos caused dose- and time-dependent reductions in deltapsi(m). Asbestos-induced deltapsi(m) was associated with the release of cytochrome c from the mitochondria to the cytoplasm as well as activation of caspase 9, a mitochondrial-activated caspase. In contrast, a lower level of caspase 8, the death receptor-activated caspase, was detected in asbestos-exposed AEC. An iron chelator (phytic acid or deferoxamine) or a hydroxyl radical scavenger (sodium benzoate) each blocked asbestos-induced reductions in deltapsi(m) and caspase 9 activation, suggesting a role for iron-derived ROS. Finally, Bcl-X(L), a mitochondrial antiapoptotic protein that prevents cell death by preserving the outer mitochondrial membrane integrity, blocked asbestos-induced decreases in A549 cell deltapsi(m) and reduced apoptosis as assessed by DNA fragmentation. We conclude that asbestos-induced AEC apoptosis results from mitochondrial dysfunction, in part due to iron-derived ROS, which is followed by the release of cytochrome c and caspase 9 activation. Our findings suggest an important role for the mitochondria-regulated death pathway in the

  19. Dishevelled genes mediate a conserved mammalian PCP pathway to regulate convergent extension during neurulation

    PubMed Central

    Wang, Jianbo; Hamblet, Natasha S.; Mark, Sharayne; Dickinson, Mary E.; Brinkman, Brendan C.; Segil, Neil; Fraser, Scott E.; Chen, Ping; Wallingford, John B.; Wynshaw-Boris, Anthony

    2014-01-01

    The planar cell polarity (PCP) pathway is conserved throughout evolution, but it mediates distinct developmental processes. In Drosophila, members of the PCP pathway localize in a polarized fashion to specify the cellular polarity within the plane of the epithelium, perpendicular to the apicobasal axis of the cell. In Xenopus and zebrafish, several homologs of the components of the fly PCP pathway control convergent extension. We have shown previously that mammalian PCP homologs regulate both cell polarity and polarized extension in the cochlea in the mouse. Here we show, using mice with null mutations in two mammalian Dishevelled homologs, Dvl1 and Dvl2, that during neurulation a homologous mammalian PCP pathway regulates concomitant lengthening and narrowing of the neural plate, a morphogenetic process defined as convergent extension. Dvl2 genetically interacts with Loop-tail, a point mutation in the mammalian PCP gene Vangl2, during neurulation. By generating Dvl2 BAC (bacterial artificial chromosome) transgenes and introducing different domain deletions and a point mutation identical to the dsh1 allele in fly, we further demonstrated a high degree of conservation between Dvl function in mammalian convergent extension and the PCP pathway in fly. In the neuroepithelium of neurulating embryos, Dvl2 shows DEP domain-dependent membrane localization, a pre-requisite for its involvement in convergent extension. Intriguing, the Loop-tail mutation that disrupts both convergent extension in the neuroepithelium and PCP in the cochlea does not disrupt Dvl2 membrane distribution in the neuroepithelium, in contrast to its drastic effect on Dvl2 localization in the cochlea. These results are discussed in light of recent models on PCP and convergent extension. PMID:16571627

  20. Feedback inhibition of deoxy-D-xylulose-5-phosphate synthase regulates the methylerythritol 4-phosphate pathway.

    PubMed

    Banerjee, Aparajita; Wu, Yan; Banerjee, Rahul; Li, Yue; Yan, Honggao; Sharkey, Thomas D

    2013-06-07

    The 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway leads to the biosynthesis of isopentenyl diphosphate (IDP) and dimethylallyl diphosphate (DMADP), the precursors for isoprene and higher isoprenoids. Isoprene has significant effects on atmospheric chemistry, whereas other isoprenoids have diverse roles ranging from various biological processes to applications in commercial uses. Understanding the metabolic regulation of the MEP pathway is important considering the numerous applications of this pathway. The 1-deoxy-D-xylulose-5-phosphate synthase (DXS) enzyme was cloned from Populus trichocarpa, and the recombinant protein (PtDXS) was purified from Escherichia coli. The steady-state kinetic parameters were measured by a coupled enzyme assay. An LC-MS/MS-based assay involving the direct quantification of the end product of the enzymatic reaction, 1-deoxy-D-xylulose 5-phosphate (DXP), was developed. The effect of different metabolites of the MEP pathway on PtDXS activity was tested. PtDXS was inhibited by IDP and DMADP. Both of these metabolites compete with thiamine pyrophosphate for binding with the enzyme. An atomic structural model of PtDXS in complex with thiamine pyrophosphate and Mg(2+) was built by homology modeling and refined by molecular dynamics simulations. The refined structure was used to model the binding of IDP and DMADP and indicated that IDP and DMADP might bind with the enzyme in a manner very similar to the binding of thiamine pyrophosphate. The feedback inhibition of PtDXS by IDP and DMADP constitutes an important mechanism of metabolic regulation of the MEP pathway and indicates that thiamine pyrophosphate-dependent enzymes may often be affected by IDP and DMADP.

  1. Kaempferol regulates OPN-CD44 pathway to inhibit the atherogenesis of apolipoprotein E deficient mice

    SciTech Connect

    Xiao, Hong-Bo; Lu, Xiang-Yang; Sun, Zhi-Liang; Zhang, Heng-Bo

    2011-12-15

    Recent studies show that osteopontin (OPN) and its receptor cluster of differentiation 44 (CD44) are two pro-inflammatory cytokines contributing to the development of atherosclerosis. The objective of this study was to explore the inhibitory effect of kaempferol, a naturally occurring flavonoid compound, on atherogenesis and the mechanisms involved. The experiments were performed in aorta and plasma from C57BL/6J control and apolipoprotein E-deficient (ApoE{sup -/-}) mice treated or not with kaempferol (50 or 100 mg/kg, intragastrically) for 4 weeks. Kaempferol treatment decreased atherosclerotic lesion area, improved endothelium-dependent vasorelaxation, and increased the maximal relaxation value concomitantly with decrease in the half-maximum effective concentration, plasma OPN level, aortic OPN expression, and aortic CD44 expression in ApoE{sup -/-} mice. In addition, treatment with kaempferol also significantly decreased reactive oxygen species production in mice aorta. The present results suggest that kaempferol regulates OPN-CD44 pathway to inhibit the atherogenesis of ApoE{sup -/-} mice. -- Graphical abstract: Kaempferol regulates OPN-CD44 pathway to inhibit the atherogenesis of ApoE{sup -/-} mice. Highlights: Black-Right-Pointing-Pointer OPN-CD44 pathway plays a critical role in the development of atherosclerosis. Black-Right-Pointing-Pointer We examine lesion area, OPN and CD44 changes after kaempferol treatment. Black-Right-Pointing-Pointer Kaempferol treatment decreased atherosclerotic lesion area in ApoE{sup -/-} mice. Black-Right-Pointing-Pointer Kaempferol treatment decreased aortic OPN and CD44 expressions in ApoE{sup -/-} mice. Black-Right-Pointing-Pointer Kaempferol regulates OPN-CD44 pathway to inhibit the atherogenesis.

  2. Nrf2 pathway regulates multidrug-resistance-associated protein 1 in small cell lung cancer.

    PubMed

    Ji, Lili; Li, Hui; Gao, Pan; Shang, Guoguo; Zhang, Donna D; Zhang, Nong; Jiang, Tao

    2013-01-01

    Although multidrug-resistance-associated protein-1 (MRP1) is a major contributor to multi-drug resistance (MDR), the regulatory mechanism of Mrp1 still remains unclear. Nrf2 is a transcription factor that regulates cellular defense response through antioxidant response elements (AREs) in normal tissues. Recently, Nrf2 has emerged as an important contributor to chemo-resistance in tumor tissues. In the present study, the role of Nrf2-ARE pathway on regulation of Mrp1 was investigated. Compared with H69 lung cancer cells, H69AR cells with MDR showed significantly higher Nrf2-ARE pathway activity and expression of Mrp1 as well. When Nrf2 was knocked down in H69AR cells, MRP1's expression decreased accordingly. Moreover, those H69AR cells with reduced Nrf2 level restored sensitivity to chemo-drugs. To explore how Nrf2-ARE pathway regulates Mrp1, the promoter of Mrp1 gene was searched, and two putative AREs--ARE1 and ARE2--were found. Using reporter gene and ChIP assay, both ARE1 and ARE2 showed response to and interaction with Nrf2. In 40 cases of cancer tissues, the expression of Nrf2 and MRP1 was measured by immunohistochemistry (IHC). As the quantitive data of IHC indicated, both Nrf2 and MRP1 showed significantly higher expression in tumor tissue than adjacent non-tumor tissue. And more important, the correlation analysis of the two genes proved that their expression was correlative. Taken together, theses data suggested that Nrf2-ARE pathway is required for the regulatory expression of Mrp1 and implicated Nrf2 as a new therapeutic target for MDR.

  3. Pathway-specific regulation revisited: cross-regulation of multiple disparate gene clusters by PAS-LuxR transcriptional regulators.

    PubMed

    Vicente, Cláudia M; Payero, Tamara D; Santos-Aberturas, Javier; Barreales, Eva G; de Pedro, Antonio; Aparicio, Jesús F

    2015-06-01

    PAS-LuxR regulators are highly conserved proteins devoted to the control of antifungal production by binding to operators located in given promoters of polyene biosynthetic genes. The canonical operator of PimM, archetype of this class of regulators, has been used here to search for putative targets of orthologous protein PteF in the genome of Streptomyces avermitilis, finding 97 putative operators outside the pentaene filipin gene cluster (pte). The processes putatively affected included genetic information processing; energy, carbohydrate, and lipid metabolism; DNA replication and repair; morphological differentiation; secondary metabolite biosynthesis; and transcriptional regulation, among others. Seventeen of these operators were selected, and their binding to PimM DNA-binding domain was assessed by electrophoretic mobility shift assays. Strikingly, the protein bound all predicted operators suggesting a direct control over targeted processes. As a proof of concept, we studied the biosynthesis of the ATP-synthase inhibitor oligomycin whose gene cluster included two operators. Regulator mutants showed a severe loss of oligomycin production, whereas gene complementation of the mutant restored phenotype, and gene duplication in the wild-type strain boosted oligomycin production. Comparative gene expression analyses in parental and mutant strains by reverse transcription-quantitative polymerase chain reaction of selected olm genes corroborated production results. These results demonstrate that PteF is able to cross-regulate the biosynthesis of two related secondary metabolites, filipin and oligomycin, but might be extended to all the processes indicated above. This study highlights the complexity of the network of interactions in which PAS-LuxR regulators are involved and opens new possibilities for the manipulation of metabolite production in Streptomycetes.

  4. BMAL1-dependent regulation of the mTOR signaling pathway delays aging.

    PubMed

    Khapre, Rohini V; Kondratova, Anna A; Patel, Sonal; Dubrovsky, Yuliya; Wrobel, Michelle; Antoch, Marina P; Kondratov, Roman V

    2014-01-01

    The circadian clock, an internal time-keeping system, has been linked with control of aging, but molecular mechanisms of regulation are not known. BMAL1 is a transcriptional factor and core component of the circadian clock; BMAL1 deficiency is associated with premature aging and reduced lifespan. Here we report that activity of mammalian Target of Rapamycin Complex 1 (mTORC1) is increased upon BMAL1 deficiency both in vivo and in cell culture. Increased mTOR signaling is associated with accelerated aging; in accordance with that, treatment with the mTORC1 inhibitor rapamycin increased lifespan of Bmal1-/- mice by 50%. Our data suggest that BMAL1 is a negative regulator of mTORC1 signaling. We propose that the circadian clock controls the activity of the mTOR pathway through BMAL1-dependent mechanisms and this regulation is important for control of aging and metabolism.

  5. Universality and diversity in the signal transduction pathway that regulates seasonal reproduction in vertebrates

    PubMed Central

    Nakane, Yusuke; Yoshimura, Takashi

    2014-01-01

    Most vertebrates living outside the tropical zone show robust physiological responses in response to seasonal changes in photoperiod, such as seasonal reproduction, molt, and migration. The highly sophisticated photoperiodic mechanism in Japanese quail has been used to uncover the mechanism of seasonal reproduction. Molecular analysis of quail mediobasal hypothalamus (MBH) revealed that local thyroid hormone activation within the MBH plays a critical role in the photoperiodic response of gonads. This activation is accomplished by two gene switches: thyroid hormone-activating (DIO2) and thyroid hormone-inactivating enzymes (DIO3). Functional genomics studies have shown that long-day induced thyroid-stimulating hormone (TSH) in the pars tuberalis (PT) of the pituitary gland regulates DIO2/3 switching. In birds, light information received directly by deep brain photoreceptors regulates PT TSH. Recent studies demonstrated that Opsin 5-positive cerebrospinal fluid (CSF)-contacting neurons are deep brain photoreceptors that regulate avian seasonal reproduction. Although the involvement of TSH and DIO2/3 in seasonal reproduction has been confirmed in various mammals, the light input pathway that regulates PT TSH in mammals differs from that of birds. In mammals, the eye is the only photoreceptor organ and light information received by the eye is transmitted to the pineal gland through the circadian pacemaker, the suprachiasmatic nucleus. Nocturnal melatonin secretion from the pineal gland indicates the length of night and regulates the PT TSH. In fish, the regulatory machinery for seasonal reproduction, from light input to neuroendocrine output, has been recently demonstrated in the coronet cells of the saccus vasculosus (SV). The SV is unique to fish and coronet cells are CSF-contacting neurons. Here, we discuss the universality and diversity of signal transduction pathways that regulate vertebrate seasonal reproduction. PMID:24959116

  6. LINGO-1 Regulates Oligodendrocyte Differentiation through the Cytoplasmic Gelsolin Signaling Pathway.

    PubMed

    Shao, Zhaohui; Lee, Xinhua; Huang, Guanrong; Sheng, Guoqing; Henderson, Christopher E; Louvard, Daniel; Sohn, Jiho; Pepinsky, Blake; Mi, Sha

    2017-03-22

    Differentiation and maturation of oligodendrocyte progenitor cells (OPCs) involve the assembly and disassembly of actin microfilaments. However, how actin dynamics are regulated during this process remains poorly understood. Leucine-rich repeat and Ig-like domain-containing Nogo receptor interacting protein 1 (LINGO-1) is a negative regulator of OPC differentiation. We discovered that anti-LINGO-1 antibody-promoted OPC differentiation was accompanied by upregulation of cytoplasmic gelsolin (cGSN), an abundant actin-severing protein involved in the depolymerization of actin filaments. Treating rat OPCs with cGSN siRNA reduced OPC differentiation, whereas overexpression of cGSN promoted OPC differentiation in vitro and remyelination in vivo Furthermore, coexpression of cGSN and LINGO-1 blocked the inhibitory effect of LINGO-1. Our study demonstrates that cGSN works downstream of LINGO-1 signaling pathway, which enhances actin dynamics and is essential for OPC morphogenesis and differentiation. This finding may lead to novel therapeutic approaches for the treatment of demyelinating diseases such as multiple sclerosis (MS).SIGNIFICANCE STATEMENT Myelin loss and subsequent axon degeneration contributes to a variety of neurological diseases, such as multiple sclerosis (MS). Understanding the regulation of myelination by oligodendrocytes is therefore critical for developing therapies for the treatment of MS. We previously demonstrated that leucine-rich repeat and Ig-like domain-containing Nogo receptor interacting protein 1 (LINGO-1) is a negative regulator of oligodendrocyte differentiation and that anti-LINGO-1 promotes remyelination in preclinical animal models for MS and in a phase II acute optic neuritis clinical trial (RENEW). The mechanism by which LINGO-1 regulates oligodendrocyte differentiation is unknown. Here, we demonstrate that LINGO-1 regulates oligodendrocyte differentiation and maturation through the cytoplasmic gelsolin signaling pathway, providing new

  7. Bmi1 Regulates the Proliferation of Cochlear Supporting Cells Via the Canonical Wnt Signaling Pathway.

    PubMed

    Lu, Xiaoling; Sun, Shan; Qi, Jieyu; Li, Wenyan; Liu, Liman; Zhang, Yanping; Chen, Yan; Zhang, Shasha; Wang, Lei; Miao, Dengshun; Chai, Renjie; Li, Huawei

    2017-03-01

    Cochlear supporting cells (SCs), which include the cochlear progenitor cells, have been shown to be a promising resource for hair cell (HC) regeneration, but the mechanisms underlying the initiation and regulation of postnatal cochlear SC proliferation are not yet fully understood. Bmi1 is a member of the Polycomb protein family and has been reported to regulate the proliferation of stem cells and progenitor cells in multiple organs. In this study, we investigated the role of Bmi1 in regulating SC and progenitor cell proliferation in neonatal mice cochleae. We first showed that knockout of Bmi1 significantly inhibited the proliferation of SCs and Lgr5-positive progenitor cells after neomycin injury in neonatal mice in vitro, and we then showed that Bmi1 deficiency significantly reduced the sphere-forming ability of the organ of Corti and Lgr5-positive progenitor cells in neonatal mice. These results suggested that Bmi1 is required for the initiation of SC and progenitor cell proliferation in neonatal mice. Next, we found that DKK1 expression was significantly upregulated, while beta-catenin and Lgr5 expression were significantly downregulated in neonatal Bmi1(-/-) mice compared to wild-type controls. The observation that Bmi1 knockout downregulates Wnt signaling provides compelling evidence that Bmi1 is required for the Wnt signaling pathway. Furthermore, the exogenous Wnt agonist BIO overcame the downregulation of SC proliferation in Bmi1(-/-) mice, suggesting that Bmi1 knockout might inhibit the proliferation of SCs via downregulation of the canonical Wnt signaling pathway. Our findings demonstrate that Bmi1 plays an important role in regulating the proliferation of cochlear SCs and Lgr5-positive progenitor cells in neonatal mice through the Wnt signaling pathway, and this suggests that Bmi1 might be a new therapeutic target for HC regeneration.

  8. Interferon regulatory factor 8 regulates pathways for antigen presentation in myeloid cells and during tuberculosis.

    PubMed

    Marquis, Jean-François; Kapoustina, Oxana; Langlais, David; Ruddy, Rebecca; Dufour, Catherine Rosa; Kim, Bae-Hoon; MacMicking, John D; Giguère, Vincent; Gros, Philippe

    2011-06-01

    IRF8 (Interferon Regulatory Factor 8) plays an important role in defenses against intracellular pathogens, including several aspects of myeloid cells function. It is required for ontogeny and maturation of macrophages and dendritic cells, for activation of anti-microbial defenses, and for production of the Th1-polarizing cytokine interleukin-12 (IL-12) in response to interferon gamma (IFNγ) and protection against infection with Mycobacterium tuberculosis. The transcriptional programs and cellular pathways that are regulated by IRF8 in response to IFNγ and that are important for defenses against M. tuberculosis are poorly understood. These were investigated by transcript profiling and chromatin immunoprecipitation on microarrays (ChIP-chip). Studies in primary macrophages identified 368 genes that are regulated by IRF8 in response to IFNγ/CpG and that behave as stably segregating expression signatures (eQTLs) in F2 mice fixed for a wild-type or mutant allele at IRF8. A total of 319 IRF8 binding sites were identified on promoters genome-wide (ChIP-chip) in macrophages treated with IFNγ/CpG, defining a functional G/AGAAnTGAAA motif. An analysis of the genes bearing a functional IRF8 binding site, and showing regulation by IFNγ/CpG in macrophages and/or in M. tuberculosis-infected lungs, revealed a striking enrichment for the pathways of antigen processing and presentation, including multiple structural and enzymatic components of the Class I and Class II MHC (major histocompatibility complex) antigen presentation machinery. Also significantly enriched as IRF8 targets are the group of endomembrane- and phagosome-associated small GTPases of the IRG (immunity-related GTPases) and GBP (guanylate binding proteins) families. These results identify IRF8 as a key regulator of early response pathways in myeloid cells, including phagosome maturation, antigen processing, and antigen presentation by myeloid cells.

  9. Helicobacter pylori and the BMP pathway regulate CDX2 and SOX2 expression in gastric cells.

    PubMed

    Camilo, Vânia; Barros, Rita; Sousa, Sofia; Magalhães, Ana Maria; Lopes, Teresa; Mário Santos, António; Pereira, Teresa; Figueiredo, Céu; David, Leonor; Almeida, Raquel

    2012-10-01

    Helicobacter pylori infection is the main risk factor for intestinal metaplasia (IM) and gastric cancer development. IM is a pre-neoplastic lesion, induced by the transcription factor CDX2, where the gastric mucosa is converted to an intestinal phenotype. We previously demonstrated that key elements of the bone morphogenetic protein (BMP) pathway co-localize with CDX2 in IM and upregulate CDX2 expression in gastric cell lines. These observations, together with the hypothesis that CDX2 could be repressed by SOX2, led us to test whether H. pylori, through BMPs, SOX2 and CDX2 could participate in a molecular network critical for the development of IM. AGS cells with and without SMAD4 knock-down were co-cultured with H. pylori or BMP2 to assess the expression of BMP pathway members as well as CDX2 and SOX2 by qPCR and western blot. Proximity ligation assay (PLA) was also performed to evaluate SMAD proteins interaction. Immunohistochemistry and western blot were performed in gastric samples from mice infected with Helicobacter spp. to measure Smad4, pSmad1/5/8, Cdx2 and Sox2 expression in vivo. Increased expression and activity of the BMP pathway accompanied by CDX2 upregulation and SOX2 downregulation were observed in AGS cells co-cultured with H. pylori or BMP2. These effects were impaired by downregulation of the BMP pathway. Finally, infected mice present BMP pathway upregulation, focal Cdx2 expression and decreased Sox2. These results provide a novel link between H. pylori infection and the BMP pathway in the regulation of intestinal and gastric-specific genes that might be relevant for gastric IM.

  10. A Functional Genome-Wide In Vivo Screen Identifies New Regulators of Signalling Pathways during Early Xenopus Embryogenesis

    PubMed Central

    Zhang, Siwei; Li, Jingjing; Lea, Robert; Amaya, Enrique; Dorey, Karel

    2013-01-01

    Embryonic development requires exquisite regulation of several essential processes, such as patterning of tissues and organs, cell fate decisions, and morphogenesis. Intriguingly, these diverse processes are controlled by only a handful of signalling pathways, and mis-regulation in one or more of these pathways may result in a variety of congenital defects and diseases. Consequently, investigating how these signalling pathways are regulated at the molecular level is essential to understanding the mechanisms underlying vertebrate embryogenesis, as well as developing treatments for human diseases. Here, we designed and performed a large-scale gain-of-function screen in Xenopus embryos aimed at identifying new regulators of MAPK/Erk, PI3K/Akt, BMP, and TGF-β/Nodal signalling pathways. Our gain-of-function screen is based on the identification of gene products that alter the phosphorylation state of key signalling molecules, which report the activation state of the pathways. In total, we have identified 20 new molecules that regulate the activity of one or more signalling pathways during early Xenopus development. This is the first time that such a functional screen has been performed, and the findings pave the way toward a more comprehensive understanding of the molecular mechanisms regulating the activity of important signalling pathways under normal and pathological conditions. PMID:24244509

  11. GAME9 regulates the biosynthesis of steroidal alkaloids and upstream isoprenoids in the plant mevalonate pathway

    PubMed Central

    Cárdenas, Pablo D.; Sonawane, Prashant D.; Pollier, Jacob; Vanden Bossche, Robin; Dewangan, Veena; Weithorn, Efrat; Tal, Lior; Meir, Sagit; Rogachev, Ilana; Malitsky, Sergey; Giri, Ashok P.; Goossens, Alain; Burdman, Saul; Aharoni, Asaph

    2016-01-01

    Steroidal glycoalkaloids (SGAs) are cholesterol-derived molecules produced by solanaceous species. They contribute to pathogen defence but are toxic to humans and considered as anti-nutritional compounds. Here we show that GLYCOALKALOID METABOLISM 9 (GAME9), an APETALA2/Ethylene Response Factor, related to regulators of alkaloid production in tobacco and Catharanthus roseus, controls SGA biosynthesis. GAME9 knockdown and overexpression in tomato and potato alters expression of SGAs and upstream mevalonate pathway genes including the cholesterol biosynthesis gene STEROL SIDE CHAIN REDUCTASE 2 (SSR2). Levels of SGAs, C24-alkylsterols and the upstream mevalonate and cholesterol pathways intermediates are modified in these plants. Δ(7)-STEROL-C5(6)-DESATURASE (C5-SD) in the hitherto unresolved cholesterol pathway is a direct target of GAME9. Transactivation and promoter-binding assays show that GAME9 exerts its activity either directly or cooperatively with the SlMYC2 transcription factor as in the case of the C5-SD gene promoter. Our findings provide insight into the regulation of SGA biosynthesis and means for manipulating these metabolites in crops. PMID:26876023

  12. Dopamine is a key regulator in the signalling pathway underlying predator-induced defences in Daphnia.

    PubMed

    Weiss, Linda C; Leese, Florian; Laforsch, Christian; Tollrian, Ralph

    2015-10-07

    The waterflea Daphnia is a model to investigate the genetic basis of phenotypic plasticity resulting from one differentially expressed genome. Daphnia develops adaptive phenotypes (e.g. morphological defences) thwarting predators, based on chemical predator cue perception. To understand the genomic basis of phenotypic plasticity, the description of the precedent cellular and neuronal mechanisms is fundamental. However, key regulators remain unknown. All neuronal and endocrine stimulants were able to modulate but not induce defences, indicating a pathway of interlinked steps. A candidate able to link neuronal with endocrine responses is the multi-functional amine dopamine. We here tested its involvement in trait formation in Daphnia pulex and Daphnia longicephala using an induction assay composed of predator cues combined with dopaminergic and cholinergic stimulants. The mere application of both stimulants was sufficient to induce morphological defences. We determined dopamine localization in cells found in close association with the defensive trait. These cells serve as centres controlling divergent morphologies. As a mitogen and sclerotization agent, we anticipate that dopamine is involved in proliferation and structural formation of morphological defences. Furthermore, dopamine pathways appear to be interconnected with endocrine pathways, and control juvenile hormone and ecdysone levels. In conclusion, dopamine is suggested as a key regulator of phenotypic plasticity.

  13. Regulation of miR106b cluster through the RB pathway: mechanism and functional targets.

    PubMed

    Thangavel, Chellappagounder; Boopathi, Ettickan; Ertel, Adam; Lim, Meng; Addya, Sankar; Fortina, Paolo; Witkiewicz, Agnieszka K; Knudsen, Erik S

    2013-01-01

    The RB pathway plays a critical role in proliferation control that is commonly subverted in tumor development. However, restoration of RB pathway function can be elicited in many tumor cells by the inhibition of CDK4/6 activity that leads to dephosphorylation of RB and subsequent repression of E2F-mediated transcription. In this context, active RB/E2F complexes inhibit the expression of a critical program of coding genes that promote cell cycle progression. However, the non-coding RNA target genes downstream from RB that could be relevant for tumor biology remain obscure. Here, miRNA gene expression profiling identified the miR106b cluster as being efficiently repressed with CDK4/6 inhibition in an E2F and RB-dependent manner. Importantly, the miR106B-cluster is intragenic of MCM7, and through a series of functional studies, the basis of MCM7 regulation and concordant expression of the miRNA species within the 106b cluster was determined. Importantly, RB-mediated repression of the 106b cluster enhances the transcript levels of p21Cip1 and PTEN. These data provide a mechanistic basis for cross-talk between the RB pathway and p21 and PTEN through the regulation of the MCM7/miR106b locus.

  14. Distinct amino acid-sensing mTOR pathways regulate skeletal myogenesis.

    PubMed

    Yoon, Mee-Sup; Chen, Jie

    2013-12-01

    Signaling through the mammalian target of rapamycin (mTOR) in response to amino acid availability controls many cellular and developmental processes. mTOR is a master regulator of myogenic differentiation, but the pathways mediating amino acid signals in this process are not known. Here we examine the Rag GTPases and the class III phosphoinositide 3-kinase (PI3K) Vps34, two mediators of amino acid signals upstream of mTOR complex 1 (mTORC1) in cell growth regulation, for their potential involvement in myogenesis. We find that, although both Rag and Vps34 mediate amino acid activation of mTORC1 in C2C12 myoblasts, they have opposing functions in myogenic differentiation. Knockdown of RagA/B enhances, whereas overexpression of active RagB/C mutants impairs, differentiation, and this inhibitory function of Rag is mediated by mTORC1 suppression of the IRS1-PI3K-Akt pathway. On the other hand, Vps34 is required for myogenic differentiation. Amino acids activate a Vps34-phospholipase D1 (PLD1) pathway that controls the production of insulin-like growth factor II, an autocrine inducer of differentiation, through the Igf2 muscle enhancer. The product of PLD, phosphatidic acid, activates the enhancer in a rapamycin-sensitive but mTOR kinase-independent manner. Our results uncover amino acid-sensing mechanisms controlling the homeostasis of myogenesis and underline the versatility and context dependence of mTOR signaling.

  15. Interaction of PACAP with Sonic hedgehog reveals complex regulation of the hedgehog pathway by PKA.

    PubMed

    Niewiadomski, Pawel; Zhujiang, Annie; Youssef, Mary; Waschek, James A

    2013-11-01

    Sonic hedgehog (Shh) signaling is essential for proliferation of cerebellar granule cell progenitors (cGCPs) and its aberrant activation causes a cerebellar cancer medulloblastoma. Pituitary adenylate cyclase activating polypeptide (PACAP) inhibits Shh-driven proliferation of cGCPs and acts as tumor suppressor in murine medulloblastoma. We show that PACAP blocks canonical Shh signaling by a mechanism that involves activation of protein kinase A (PKA) and inhibition of the translocation of the Shh-dependent transcription factor Gli2 into the primary cilium. PKA is shown to play an essential role in inhibiting gene transcription in the absence of Shh, but global PKA activity levels are found to be a poor predictor of the degree of Shh pathway activation. We propose that the core Shh pathway regulates a small compartmentalized pool of PKA in the vicinity of primary cilia. GPCRs that affect global PKA activity levels, such as the PACAP receptor, cooperate with the canonical Shh signal to regulate Gli protein phosphorylation by PKA. This interaction serves to fine-tune the transcriptional and physiological function of the Shh pathway.

  16. GAME9 regulates the biosynthesis of steroidal alkaloids and upstream isoprenoids in the plant mevalonate pathway.

    PubMed

    Cárdenas, Pablo D; Sonawane, Prashant D; Pollier, Jacob; Vanden Bossche, Robin; Dewangan, Veena; Weithorn, Efrat; Tal, Lior; Meir, Sagit; Rogachev, Ilana; Malitsky, Sergey; Giri, Ashok P; Goossens, Alain; Burdman, Saul; Aharoni, Asaph

    2016-02-15

    Steroidal glycoalkaloids (SGAs) are cholesterol-derived molecules produced by solanaceous species. They contribute to pathogen defence but are toxic to humans and considered as anti-nutritional compounds. Here we show that GLYCOALKALOID METABOLISM 9 (GAME9), an APETALA2/Ethylene Response Factor, related to regulators of alkaloid production in tobacco and Catharanthus roseus, controls SGA biosynthesis. GAME9 knockdown and overexpression in tomato and potato alters expression of SGAs and upstream mevalonate pathway genes including the cholesterol biosynthesis gene STEROL SIDE CHAIN REDUCTASE 2 (SSR2). Levels of SGAs, C24-alkylsterols and the upstream mevalonate and cholesterol pathways intermediates are modified in these plants. Δ(7)-STEROL-C5(6)-DESATURASE (C5-SD) in the hitherto unresolved cholesterol pathway is a direct target of GAME9. Transactivation and promoter-binding assays show that GAME9 exerts its activity either directly or cooperatively with the SlMYC2 transcription factor as in the case of the C5-SD gene promoter. Our findings provide insight into the regulation of SGA biosynthesis and means for manipulating these metabolites in crops.

  17. Regulation of the PI3K pathway through a p85α monomer–homodimer equilibrium

    PubMed Central

    Cheung, Lydia WT; Walkiewicz, Katarzyna W; Besong, Tabot MD; Guo, Huifang; Hawke, David H; Arold, Stefan T; Mills, Gordon B

    2015-01-01

    The canonical action of the p85α regulatory subunit of phosphatidylinositol 3-kinase (PI3K) is to associate with the p110α catalytic subunit to allow stimuli-dependent activation of the PI3K pathway. We elucidate a p110α-independent role of homodimerized p85α in the positive regulation of PTEN stability and activity. p110α-free p85α homodimerizes via two intermolecular interactions (SH3:proline-rich region and BH:BH) to selectively bind unphosphorylated activated PTEN. As a consequence, homodimeric but not monomeric p85α suppresses the PI3K pathway by protecting PTEN from E3 ligase WWP2-mediated proteasomal degradation. Further, the p85α homodimer enhances the lipid phosphatase activity and membrane association of PTEN. Strikingly, we identified cancer patient-derived oncogenic p85α mutations that target the homodimerization or PTEN interaction surface. Collectively, our data suggest the equilibrium of p85α monomer–dimers regulates the PI3K pathway and disrupting this equilibrium could lead to disease development. DOI: http://dx.doi.org/10.7554/eLife.06866.001 PMID:26222500

  18. Neural mechanisms of emotion regulation: Evidence for two independent prefrontal-subcortical pathways

    PubMed Central

    Wager, Tor D.; Davidson, Matthew L.; Hughes, Brent L.; Lindquist, Martin A.; Ochsner, Kevin N.

    2008-01-01

    Although prefrontal cortex has been implicated in the cognitive regulation of emotion, the cortical-subcortical interactions that mediate this ability remain poorly understood. To address this issue, we identified a right ventrolateral prefrontal region (vlPFC) whose activity correlated with reduced negative emotional experience during cognitive reappraisal of aversive images. We then applied a novel pathway-mapping analysis on subcortical regions to locate mediators of the association between vlPFC activity and reappraisal success (i.e. reductions in reported emotion). Results identified two separable pathways that together explained ~50% of the reported variance in self-reported emotion: 1) a path through nucleus accumbens that predicted greater reappraisal success, and 2) a path through ventral amygdala that predicted reduced reappraisal success (i.e., more negative emotion). These results provide direct evidence that vlPFC is involved in both the generation and regulation of emotion through different subcortical pathways, suggesting a general role for this region in appraisal processes. PMID:18817740

  19. MicroRNAs regulate signaling pathways in osteogenic differentiation of mesenchymal stem cells (Review)

    PubMed Central

    PENG, SHUPING; GAO, DAN; GAO, CHENGDE; WEI, PINGPIN; NIU, MAN; SHUAI, CIJUN

    2016-01-01

    Osteogenesis is a complex multi-step process involving the differentiation of mesenchymal stem cells (MSCs) into osteoblast progenitor cells, preosteoblasts, osteoblasts and osteocytes, and the crosstalk between multiple cell types for the formation and remodeling of bone. The signaling regulatory networks during osteogenesis include various components, including growth factors, transcription factors, micro (mi)RNAs and effectors, a number of which form feedback loops controlling the balance of osteogenic differentiation by positive or negative regulation. miRNAs have been found to be important regulators of osteogenic signaling pathways in multiple aspects and multiple signaling pathways. The present review focusses on the progress in elucidating the role of miRNA in the osteogenesis signaling networks of MSCs as a substitute for bone implantation the the field of bone tissue engineering. In particular, the review classifies which miRNAs promote or suppress the osteogenic process, and summarizes which signaling pathway these miRNAs are involved in. Improvements in knowledge of the characteristics of miRNAs in osteogenesis provide an important step for their application in translational investigations of bone tissue engineering and bone disease. PMID:27222009

  20. Dopamine is a key regulator in the signalling pathway underlying predator-induced defences in Daphnia

    PubMed Central

    Weiss, Linda C.; Leese, Florian; Laforsch, Christian; Tollrian, Ralph

    2015-01-01

    The waterflea Daphnia is a model to investigate the genetic basis of phenotypic plasticity resulting from one differentially expressed genome. Daphnia develops adaptive phenotypes (e.g. morphological defences) thwarting predators, based on chemical predator cue perception. To understand the genomic basis of phenotypic plasticity, the description of the precedent cellular and neuronal mechanisms is fundamental. However, key regulators remain unknown. All neuronal and endocrine stimulants were able to modulate but not induce defences, indicating a pathway of interlinked steps. A candidate able to link neuronal with endocrine responses is the multi-functional amine dopamine. We here tested its involvement in trait formation in Daphnia pulex and Daphnia longicephala using an induction assay composed of predator cues combined with dopaminergic and cholinergic stimulants. The mere application of both stimulants was sufficient to induce morphological defences. We determined dopamine localization in cells found in close association with the defensive trait. These cells serve as centres controlling divergent morphologies. As a mitogen and sclerotization agent, we anticipate that dopamine is involved in proliferation and structural formation of morphological defences. Furthermore, dopamine pathways appear to be interconnected with endocrine pathways, and control juvenile hormone and ecdysone levels. In conclusion, dopamine is suggested as a key regulator of phenotypic plasticity. PMID:26423840

  1. LncRNA mediated regulation of aging pathways in Drosophila melanogaster during dietary restriction

    PubMed Central

    Tu, Jianbo; Gaur, Uma; Mao, Xueping; Fan, Xiaolan; Li, Diyan; Li, Ying; Yang, Mingyao

    2016-01-01

    Dietary restriction (DR) extends lifespan in many species which is a well-known phenomenon. Long non-coding RNAs (lncRNAs) play an important role in regulation of cell senescence and important age-related signaling pathways. Here, we profiled the lncRNA and mRNA transcriptome of fruit flies at 7 day and 42 day during DR and fully-fed conditions, respectively. In general, 102 differentially expressed lncRNAs and 1406 differentially expressed coding genes were identified. Most informatively we found a large number of differentially expressed lncRNAs and their targets enriched in GO and KEGG analysis. We discovered some new aging related signaling pathways during DR, such as hippo signaling pathway-fly, phototransduction-fly and protein processing in endoplasmic reticulum etc. Novel lncRNAs XLOC_092363 and XLOC_166557 are found to be located in 10 kb upstream sequences of hairy and ems promoters, respectively. Furthermore, tissue specificity of some novel lncRNAs had been analyzed at 7 day of DR in fly head, gut and fat body. Also the silencing of lncRNA XLOC_076307 resulted in altered expression level of its targets including Gadd45 (involved in FoxO signaling pathway). Together, the results implicated many lncRNAs closely associated with dietary restriction, which could provide a resource for lncRNA in aging and age-related disease field. PMID:27687893

  2. Improving fatty acids production by engineering dynamic pathway regulation and metabolic control

    PubMed Central

    Xu, Peng; Li, Lingyun; Zhang, Fuming; Stephanopoulos, Gregory; Koffas, Mattheos

    2014-01-01

    Global energy demand and environmental concerns have stimulated increasing efforts to produce carbon-neutral fuels directly from renewable resources. Microbially derived aliphatic hydrocarbons, the petroleum-replica fuels, have emerged as promising alternatives to meet this goal. However, engineering metabolic pathways with high productivity and yield requires dynamic redistribution of cellular resources and optimal control of pathway expression. Here we report a genetically encoded metabolic switch that enables dynamic regulation of fatty acids (FA) biosynthesis in Escherichia coli. The engineered strains were able to dynamically compensate the critical enzymes involved in the supply and consumption of malonyl-CoA and efficiently redirect carbon flux toward FA biosynthesis. Implementation of this metabolic control resulted in an oscillatory malonyl-CoA pattern and a balanced metabolism between cell growth and product formation, yielding 15.7- and 2.1-fold improvement in FA titer compared with the wild-type strain and the strain carrying the uncontrolled metabolic pathway. This study provides a new paradigm in metabolic engineering to control and optimize metabolic pathways facilitating the high-yield production of other malonyl-CoA–derived compounds. PMID:25049420

  3. Fanconi anemia pathway regulates convergent transcription-induced cell death at trinucleotide repeats in human cells

    PubMed Central

    Chatterjee, Nimrat; Lin, Yunfu; Wilson, John H.

    2016-01-01

    Almost 20 incurable neurodegenerative disorders are caused by trinucleotide repeat (TNR) expansion beyond a certain threshold, with disease time of onset and severity positively correlating with repeat length. Typically, long TNRs display a bias toward further expansion and repeats continue to expand not only during germline transmissions from parents to offspring, but also remain highly unstable in somatic tissues of patients. Hence, understanding TNR instability mechanisms sheds light on underlying disease pathology. Recently, we showed that activated ATR is the major signal for convergent-transcription-induced cell death at CAG repeats and is regulated by the mismatch repair (MMR) pathway. Additionally, components of other DNA repair pathways such as transcription-coupled nucleotide excision repair (TC-NER) and R-loop resolution by RNaseH reduce cell death. Because activated ATR signals the Fanconi anemia (FA) pathway of interstrand crosslink DNA repair, we asked whether the FA pathway also modulates convergent-transcription-induced cell death at expanded CAG repeats. We show here that siRNA knockdown of FA components—FANCI, FANCJ, FANCM, FANCA, and FANCD2—decreases cell death, suggesting that FA proteins, like MMR proteins, are activators of cell death during convergent transcription. PMID:27595121

  4. Effects of the Insulin-like Growth Factor Pathway on the Regulation of Mammary Gland Development

    PubMed Central

    Ha, Woo Tae; Jeong, Ha Yeon; Lee, Seung Yoon; Song, Hyuk

    2016-01-01

    The insulin-like growth factor (IGF) pathway is a key signal transduction pathway involved in cell proliferation, migration, and apoptosis. In dairy cows, IGF family proteins and binding receptors, including their intracellular binding partners, regulate mammary gland development. IGFs and IGF receptor interactions in mammary glands influence the early stages of mammogenesis, i.e., mammary ductal genesis until puberty. The IGF pathway includes three major components, IGFs (such as IGF-I, IGF-II, and insulin), their specific receptors, and their high-affinity binding partners (IGF binding proteins [IGFBPs]; i.e., IGFBP1–6), including specific proteases for each IGFBP. Additionally, IGFs and IGFBP interactions are critical for the bioactivities of various intracellular mechanisms, including cell proliferation, migration, and apoptosis. Notably, the interactions between IGFs and IGFBPs in the IGF pathway have been difficult to characterize during specific stages of bovine mammary gland development. In this review, we aim to describe the role of the interaction between IGFs and IGFBPs in overall mammary gland development in dairy cows. PMID:27795999

  5. Down-regulated Lotus japonicus GCR1 plants exhibit nodulation signalling pathways alteration.

    PubMed

    Rogato, Alessandra; Valkov, Vladimir Totev; Alves, Ludovico Martins; Apone, Fabio; Colucci, Gabriella; Chiurazzi, Maurizio

    2016-06-01

    G Protein Coupled Receptor (GPCRs) are integral membrane proteins involved in various signalling pathways by perceiving many extracellular signals and transducing them to heterotrimeric G proteins, which further transduce these signals to intracellular downstream effectors. GCR1 is the only reliable plant candidate as a member of the GPCRs superfamily. In the legume/rhizobia symbiotic interaction, G proteins are involved in signalling pathways controlling different steps of the nodulation program. In order to investigate the putative hierarchic role played by GCR1 in these symbiotic pathways we identified and characterized the Lotus japonicus gene encoding the seven transmembrane GCR1 protein. The detailed molecular and topological analyses of LjGCR1 expression patterns that are presented suggest a possible involvement in the early steps of nodule organogenesis. Furthermore, phenotypic analyses of independent transgenic RNAi lines, showing a significant LjGCR1 expression down regulation, suggest an epistatic action in the control of molecular markers of nodulation pathways, although no macroscopic symbiotic phenotypes could be revealed.

  6. The role of phosphatidylinositol signaling pathway in regulating serotonin-induced oocyte maturation in Mercenaria mercenaria

    NASA Astrophysics Data System (ADS)

    Wang, Qing; Zhang, Tao

    2011-05-01

    Serotonin (5-HT) has been found to stimulate meiotic maturation of oocytes in many molluscs. During maturation, several signaling pathways are involved, especially the phosphatidylinositol and cAMP pathways. In order to examine the possible role of the phosphatidylinositol signaling pathway in regulating oocyte maturation in Mercenaria mercenaria, the effects of the activator/inhibitor of phospholipase (PLC) and protein kinase C (PKC) on serotonin-induced maturation were studied. Results show that high-concentrations of neomycin (inhibitor of PLC) blocked oocyte maturation, while 9, 10-dimethyl-1, 2-benzanthracene (DMBA, activator of PLC) promoted oocyte maturation in the presence of serotonin. It was also found that in the presence of serotonin, phorbol 12-myristate 13-acetate (PMA, activator of PKC) inhibited oocyte maturation, while sphingosine (inhibitor of PKC) stimulated oocyte maturation. These results indicate that serotonin-induced oocyte maturation requires the activation of the phosphatidylinositol pathway. Decrease of PLC inhibited serotonin-induced oocyte maturation, whereas a decrease of PKC stimulated the maturation. Thus, our study indicates that serotonin promotes maturation of M. mercenaria oocytes through PLC stimulated increase in calcium ion concentration via inositol-1, 4, 5-trisphosphate (IP3) but not PKC.

  7. Berberine regulates neurite outgrowth through AMPK-dependent pathways by lowering energy status

    SciTech Connect

    Lu, Jiaqi; Cao, Yuanzhao; Cheng, Kuoyuan; Xu, Bo; Wang, Tianchang; Yang, Qi; Yang, Qin; Feng, Xudong; Xia, Qing

    2015-06-10

    As a widely used anti-bacterial agent and a metabolic inhibitor as well as AMP-activated protein kinase (AMPK) activator, berberine (BBR) has been shown to cross the blood–brain barrier. Its efficacy has been investigated in various disease models of the central nervous system. Neurite outgrowth is critical for nervous system development and is a highly energy-dependent process regulated by AMPK-related pathways. In the present study, we aimed to investigate the effects of BBR on AMPK activation and neurite outgrowth in neurons. The neurite outgrowth of primary rat cortical neurons at different stages of polarization was monitored after exposure of BBR. Intracellular energy level, AMPK activation and polarity-related pathways were also inspected. The results showed that BBR suppressed neurite outgrowth and affected cytoskeleton stability in the early stages of neuronal polarization, which was mediated by lowered energy status and AMPK activation. Liver kinase B1 and PI3K–Akt–GSK3β signaling pathways were also involved. In addition, mitochondrial dysfunction and endoplasmic reticulum stress contributed to the lowered energy status induced by BBR. This study highlighted the knowledge of the complex activities of BBR in neurons and corroborated the significance of energy status during the neuronal polarization. - Highlights: • BBR inhibited neurite outgrowth in early stages of neuronal development. • Lowered neuronal energy status was induced by BBR treatment. • Neuronal energy stress induced by BBR activated AMPK-related pathways. • BBR induced mitochondrial dysfunction and endoplasmic reticulum stress.

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

    PubMed

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

    2015-09-29

    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.

  9. RAS/ERK pathway transcriptional regulation through ETS/AP-1 binding sites.

    PubMed

    Hollenhorst, Peter C

    2012-01-01

    The RAS/RAF/MEK/ERK signaling pathway is activated by mutation in many cancers. Neighboring ETS and AP-1 DNA binding sequences can act as response elements for transcriptional activation by this pathway. ERK phosphorylation of an ETS transcription factor is one mechanism of activating the RAS/ERK gene expression program that can promote cancer cell phenotypes such as proliferation, invasion, and metastasis. Recent genome-wide mapping of ETS proteins over-expressed by chromosomal rearrangement in prostate cancer reveals a second mechanism for activation of this gene expression program. An oncogenic subset of ETS transcription factors can activate RAS/ERK target genes even in the absence of RAS/ERK pathway activation by binding ETS/AP-1 sequences. Thus, regulation of cancer cell invasion and metastasis via ETS/AP-1 sequence elements depends on which ETS protein is bound, and the status of the RAS/ERK pathway. This commentary will focus on what is known about the selectivity of ETS/AP-1 sequences for different ETS transcription factors and the transcriptional consequences of ETS protein selection.

  10. The planar cell polarity pathway drives pathogenesis of chronic lymphocytic leukemia by the regulation of B-lymphocyte migration.

    PubMed

    Kaucká, Markéta; Plevová, Karla; Pavlová, Sárka; Janovská, Pavlína; Mishra, Archana; Verner, Jan; Procházková, Jirina; Krejcí, Pavel; Kotasková, Jana; Ovesná, Petra; Tichy, Boris; Brychtová, Yvona; Doubek, Michael; Kozubík, Alois; Mayer, Jirí; Pospísilová, Sárka; Bryja, Vítezslav

    2013-03-01

    The planar cell polarity (PCP) pathway is a conserved pathway that regulates cell migration and polarity in various contexts. Here we show that key PCP pathway components such as Vangl2, Celsr1, Prickle1, FZD3, FZD7, Dvl2, Dvl3, and casein kinase 1 (CK1)-ε are upregulated in B lymphocytes of patients with chronic lymphocytic leukemia (CLL). Elevated levels of PCP proteins accumulate in advanced stages of the disease. Here, we show that PCP pathway is required for the migration and transendothelial invasion of CLL cells and that patients with high expression of PCP genes, FZD3, FZD7, and PRICKLE1, have a less favorable clinical prognosis. Our findings establish that the PCP pathway acts as an important regulator of CLL cell migration and invasion. PCP proteins represent an important class of molecules regulating pathogenic interaction of CLL cells with their microenvironment.

  11. Regulation of angiogenesis by a non-canonical Wnt-Flt1 pathway in myeloid cells.

    PubMed

    Stefater, James A; Lewkowich, Ian; Rao, Sujata; Mariggi, Giovanni; Carpenter, April C; Burr, Adam R; Fan, Jieqing; Ajima, Rieko; Molkentin, Jeffery D; Williams, Bart O; Wills-Karp, Marsha; Pollard, Jeffrey W; Yamaguchi, Terry; Ferrara, Napoleone; Gerhardt, Holger; Lang, Richard A

    2011-05-29

    Myeloid cells are a feature of most tissues. Here we show that during development, retinal myeloid cells (RMCs) produce Wnt ligands to regulate blood vessel branching. In the mouse retina, where angiogenesis occurs postnatally, somatic deletion in RMCs of the Wnt ligand transporter Wntless results in increased angiogenesis in the deeper layers. We also show that mutation of Wnt5a and Wnt11 results in increased angiogenesis and that these ligands elicit RMC responses via a non-canonical Wnt pathway. Using cultured myeloid-like cells and RMC somatic deletion of Flt1, we show that an effector of Wnt-dependent suppression of angiogenesis by RMCs is Flt1, a naturally occurring inhibitor of vascular endothelial growth factor (VEGF). These findings indicate that resident myeloid cells can use a non-canonical, Wnt-Flt1 pathway to suppress angiogenic branching.

  12. A chemical proteomic atlas of brain serine hydrolases identifies cell type-specific pathways regulating neuroinflammation.

    PubMed

    Viader, Andreu; Ogasawara, Daisuke; Joslyn, Christopher M; Sanchez-Alavez, Manuel; Mori, Simone; Nguyen, William; Conti, Bruno; Cravatt, Benjamin F

    2016-01-18

    Metabolic specialization among major brain cell types is central to nervous system function and determined in large part by the cellular distribution of enzymes. Serine hydrolases are a diverse enzyme class that plays fundamental roles in CNS metabolism and signaling. Here, we perform an activity-based proteomic analysis of primary mouse neurons, astrocytes, and microglia to furnish a global portrait of the cellular anatomy of serine hydrolases in the brain. We uncover compelling evidence for the cellular compartmentalization of key chemical transmission pathways, including the functional segregation of endocannabinoid (eCB) biosynthetic enzymes diacylglycerol lipase-alpha (DAGLα) and -beta (DAGLβ) to neurons and microglia, respectively. Disruption of DAGLβ perturbed eCB-eicosanoid crosstalk specifically in microglia and suppressed neuroinflammatory events in vivo independently of broader effects on eCB content. Mapping the cellular distribution of metabolic enzymes thus identifies pathways for regulating specialized inflammatory responses in the brain while avoiding global alterations in CNS function.

  13. CD47 Receptor Globally Regulates Metabolic Pathways That Control Resistance to Ionizing Radiation.

    PubMed

    Miller, Thomas W; Soto-Pantoja, David R; Schwartz, Anthony L; Sipes, John M; DeGraff, William G; Ridnour, Lisa A; Wink, David A; Roberts, David D

    2015-10-09

    Modulating tissue responses to stress is an important therapeutic objective. Oxidative and genotoxic stresses caused by ionizing radiation are detrimental to healthy tissues but beneficial for treatment of cancer. CD47 is a signaling receptor for thrombospondin-1 and an attractive therapeutic target because blocking CD47 signaling protects normal tissues while sensitizing tumors to ionizing radiation. Here we utilized a metabolomic approach to define molecular mechanisms underlying this radioprotective activity. CD47-deficient cells and cd47-null mice exhibited global advantages in preserving metabolite levels after irradiation. Metabolic pathways required for controlling oxidative stress and mediating DNA repair were enhanced. Some cellular energetics pathways differed basally in CD47-deficient cells, and the global declines in the glycolytic and tricarboxylic acid cycle metabolites characteristic of normal cell and tissue responses to irradiation were prevented in the absence of CD47. Thus, CD47 mediates signaling from the extracellular matrix that coordinately regulates basal metabolism and cytoprotective responses to radiation injury.

  14. The acid sphingomyelinase/ceramide pathway: biomedical significance and mechanisms of regulation.

    PubMed

    Zeidan, Y H; Hannun, Y A

    2010-07-01

    One of the most intriguing enzymes of sphingolipid biology is acid sphingomyelinase (ASMase). In a phospholipase C reaction, ASMase catalyzes the cleavage of the phosphocholine head group of sphingomyelin to generate ceramide. Cumulative efforts of various laboratories over the past 40 years have placed ASMase and its product ceramide at the forefront of lipid research. Activation of the ASMase/ceramide pathway is a shared response to an ever-growing list of receptor and non-receptor mediated forms of cellular stress including: death ligands (TNFalpha, TRAIL, Fas ligand), cytokines (IL-1, IFNgamma), radiation, pathogenic infections, cytotoxic agents and others. The strategic role of ASMase in lipid metabolism and cellular stress response has sparked interest in investigatig the molecular mechanisms underlying ASMase activation. In this article, we review the translational role of the ASMase/ceramide pathway and recent advances on its mechanisms of regulation.

  15. Down regulation of gene related sex hormone synthesis pathway in mouse testes by miroestrol and deoxymiroestrol.

    PubMed

    Udomsuk, Latiporn; Juengwatanatrakul, Thaweesak; Putalun, Waraporn; Jarukamjorn, Kanokwan

    2011-12-01

    Miroestrol and deoxymiroestrol are phytoestrogens isolated from tuberous root of Pueraria candollei var. mirifica. Modulatory effects of miroestrol and deoxymiroestrol on enzymes involved in sex-hormone synthesis pathway in male C57BL/6 mice were investigated using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). Miroestrol and deoxymiroestrol suppressed the expressions of 3β-HSD, 17β-HSD1, and CYP17 while CYP19 mRNA expression was slightly decreased. In addition, the expression of 17β-HSD2 was induced in correlation with those did by estradiol. These observations supported that miroestrol and deoxymiroestrol could exhibit the same effect as estradiol regarding regulation of testicular gene related sex hormone synthesis pathway.

  16. Signaling pathways regulating neuron-glia interaction and their implications in Alzheimer’s disease

    PubMed Central

    Lian, Hong; Zheng, Hui

    2015-01-01

    Astrocytes are the most abundant cells in the central nervous system. They play critical roles in neuronal homeostasis through their physical properties and neuron-glia signaling pathways. Astrocytes become reactive in response to neuronal injury and this process, referred to as reactive astrogliosis, is a common feature accompanying neurodegenerative conditions, particularly Alzheimer’s disease. Reactive astrogliosis represents a continuum of pathobiological processes and is associated with morphological, functional and gene expression changes of varying degrees. There has been a substantial growth of knowledge regarding the signaling pathways regulating glial biology and pathophysiology in recent years. Here we attempt to provide an unbiased review of some of the well-known players, namely calcium, proteoglycan, TGFβ, NFκB, and complement, in mediating neuron-glia interaction under physiological conditions as well as in Alzheimer’s disease. PMID:26546579

  17. [Dual-role regulations of canonical Wnt/beta-catenin signaling pathway].

    PubMed

    Liu, Yang; Zhang, Chen-guang; Zhou, Chun-yan

    2010-04-18

    In recent years, Wnt/beta-catenin signaling has been identified as a key player in embryogenesis and human diseases. Canonical Wnt signaling pathway is controlled by a variety of classic molecules like Wnt, beta-catenin, Axin, APC, GSK-3beta and CK1, which interact and coordinate to regulate the expressions of cell signaling molecules. The latest evidences suggest that some components of the Wnt/beta-catenin signaling, like APC, GSK-3beta, CK1, Dkk2 and WISE, play dual roles different from what they have been thought previously. Here we reviewed some recent discoveries on the canonical Wnt/beta-catenin signaling pathway to provide some new ideas and principles for signaling transduction studies.

  18. The Arabidopsis GAI gene defines a signaling pathway that negatively regulates gibberellin responses 

    PubMed Central

    Peng, Jinrong; Carol, Pierre; Richards, Donald E.; King, Kathryn E.; Cowling, Rachel J.; Murphy, George P.; Harberd, Nicholas P.

    1997-01-01

    The Arabidopsis gai mutant allele confers a reduction in gibberellin (GA) responsiveness. Here we report the molecular cloning of GAI and a closely related gene GRS. The predicted GAI (wild-type) and gai (mutant) proteins differ only by the deletion of a 17-amino-acid segment from within the amino-terminal region. GAI and GRS contain nuclear localization signals, a region of homology to a putative transcription factor, and motifs characteristic of transcriptional coactivators. Genetic analysis indicates that GAI is a repressor of GA responses, that GA can release this repression, and that gai is a mutant repressor that is relatively resistant to the effects of GA. Mutations at SPY and GAR2 suppress the gai phenotype, indicating the involvement of GAI, SPY, and GAR2 in a signaling pathway that regulates GA responses negatively. The existence of this pathway suggests that GA modulates plant growth through derepression rather than through simple stimulation. PMID:9389651

  19. Kinetics of early TCR signaling regulate the pathway of lytic granule delivery to the secretory domain

    PubMed Central

    Beal, Allison M.; Anikeeva, Nadia; Varma, Rajat; Cameron, Thomas O.; Vasiliver-Shamis, Gaia; Norris, Philip J.; Dustin, Michael L.; Sykulev, Yuri

    2009-01-01

    SUMMARY Cytolytic granule mediated killing of virus-infected cells is an essential function of cytotoxic T lymphocytes. Analysis of lytic granule delivery shows that the granules can take long or short paths to the secretory domain where they are released. Both paths utilize the same intracellular molecular events, which have different spatial and temporal arrangements in each path and are regulated by the kinetics of downstream Ca2+ mediated signaling. Rapid and robust signaling causes swift granule concentration near the MTOC and subsequent delivery by the polarized MTOC directly to the secretory domain - the shortest and fastest path. Indolent signaling leads to late recruitment of granules that move along microtubules to the periphery of the synapse and then move tangentially to fuse at the outer edge of the secretory domain - a longer path. The short pathway is associated with faster granule release and more efficient killing than the long pathway. PMID:19833088

  20. Melatonin Regulates Aging and Neurodegeneration through Energy Metabolism, Epigenetics, Autophagy and Circadian Rhythm Pathways

    PubMed Central

    Jenwitheesuk, Anorut; Nopparat, Chutikorn; Mukda, Sujira; Wongchitrat, Prapimpun; Govitrapong, Piyarat

    2014-01-01

    Brain aging is linked to certain types of neurodegenerative diseases and identifying new therapeutic targets has become critical. Melatonin, a pineal hormone, associates with molecules and signaling pathways that sense and influence energy metabolism, autophagy, and circadian rhythms, including insulin-like growth factor 1 (IGF-1), Forkhead box O (FoxOs), sirtuins and mammalian target of rapamycin (mTOR) signaling pathways. This review summarizes the current understanding of how melatonin, together with molecular, cellular and systemic energy metabolisms, regulates epigenetic processes in the neurons. This information will lead to a greater understanding of molecular epigenetic aging of the brain and anti-aging mechanisms to increase lifespan under healthy conditions. PMID:25247581

  1. Xenopus Cdc42 regulates convergent extension movements during gastrulation through Wnt/Ca2+ signaling pathway.

    PubMed

    Choi, Sun-Cheol; Han, Jin-Kwan

    2002-04-15

    Rho GTPases are molecular switches that regulate many essential cellular processes, including actin dynamics, cell adhesion, cell-cycle progression, and transcription. We have isolated the Xenopus homolog of Rho GTPase Cdc42 and examined its potential role during gastrulation movements in early Xenopus embryos. XCdc42 is expressed in tissues undergoing extensive morphogenetic changes, such as the deep layers of involuting mesoderm and posterior neuroectoderm during gastrulation, and somitic mesoderm at neurula stages. Overexpression of either wild-type (WT) or dominant-negative (DN) XCdc42 interferes with convergent extension movements in intact embryos, activin-stimulated animal caps, and dorsal marginal zone explants. These effects occur without affecting mesodermal specification. Overexpression of WT or DN XCdc42 leads to the decrease and increase of cell adhesiveness of blastomeres, respectively, as demonstrated by the cell adhesion assay. In addition, when overexpressed, PKC-alpha, XWnt-5a, and Mfz-3 inhibit activin-induced convergent extension in animal cap explants. This inhibition can be rescued by coexpression of DN XCdc42, implying that XCdc42 acts downstream of the Wnt/Ca2+ signaling pathway involving PKC activation. XCdc42 also lies downstream of XWnt-5a in the regulation of Ca2+-dependent cell adhesion. Taken together, our results suggest that XCdc42 plays a role in the regulation of convergent extension movements during gastrulation through the protein kinase C-mediated Wnt/Ca2+ pathway.

  2. Spatial Regulation of Root Growth: Placing the Plant TOR Pathway in a Developmental Perspective.

    PubMed

    Barrada, Adam; Montané, Marie-Hélène; Robaglia, Christophe; Menand, Benoît

    2015-08-19

    Plant cells contain specialized structures, such as a cell wall and a large vacuole, which play a major role in cell growth. Roots follow an organized pattern of development, making them the organs of choice for studying the spatio-temporal regulation of cell proliferation and growth in plants. During root growth, cells originate from the initials surrounding the quiescent center, proliferate in the division zone of the meristem, and then increase in length in the elongation zone, reaching their final size and differentiation stage in the mature zone. Phytohormones, especially auxins and cytokinins, control the dynamic balance between cell division and differentiation and therefore organ size. Plant growth is also regulated by metabolites and nutrients, such as the sugars produced by photosynthesis or nitrate assimilated from the soil. Recent literature has shown that the conserved eukaryotic TOR (target of rapamycin) kinase pathway plays an important role in orchestrating plant growth. We will summarize how the regulation of cell proliferation and cell expansion by phytohormones are at the heart of root growth and then discuss recent data indicating that the TOR pathway integrates hormonal and nutritive signals to orchestrate root growth.

  3. Circadian clock regulates response to pesticides in Drosophila via conserved Pdp1 pathway.

    PubMed

    Beaver, Laura Michelle; Hooven, Louisa Ada; Butcher, Shawn Michael; Krishnan, Natraj; Sherman, Katherine Alice; Chow, Eileen Shin-Yeu; Giebultowicz, Jadwiga Maria

    2010-06-01

    Daily rhythms generated by the circadian clock regulate many life functions, including responses to xenobiotic compounds. In Drosophila melanogaster, the circadian clock consists of positive elements encoded by cycle (cyc) and Clock (Clk) and negative elements encoded by period (per) and timeless (tim) genes. The epsilon-isoform of the PAR-domain protein 1 (Pdp1epsilon) transcription factor is controlled by positive clock elements and regulates daily locomotor activity rhythms. Pdp1 target genes have not been identified, and its involvement in other clock output pathways is not known. Mammalian orthologs of Pdp1 have been implicated in the regulation of xenobiotic metabolism; therefore, we asked whether Pdp1 has a similar role in the fly. Using pesticides as model toxicants, we determined that disruption of Pdp1epsilon increased pesticide-induced mortality in flies. Flies deficient for cyc also showed increased mortality, while disruption of per and tim had no effect. Day/night and Pdp1-dependent differences in the expression of xenobiotic-metabolizing enzymes Cyp6a2, Cyp6g1, and alpha-Esterase-7 were observed and likely contribute to impaired detoxification. DHR96, a homolog of constitutive androstane receptor and pregnane X receptor, is involved in pesticide response, and DHR96 expression decreased when Pdp1 was suppressed. Taken together, our data uncover a pathway from the positive arm of the circadian clock through Pdp1 to detoxification effector genes, demonstrating a conserved role of the circadian system in modulating xenobiotic toxicity.

  4. Autopalmitoylation of TEAD proteins regulates transcriptional output of the Hippo pathway.

    PubMed

    Chan, PuiYee; Han, Xiao; Zheng, Baohui; DeRan, Michael; Yu, Jianzhong; Jarugumilli, Gopala K; Deng, Hua; Pan, Duojia; Luo, Xuelian; Wu, Xu

    2016-04-01

    TEA domain (TEAD) transcription factors bind to the coactivators YAP and TAZ and regulate the transcriptional output of the Hippo pathway, playing critical roles in organ size control and tumorigenesis. Protein S-palmitoylation attaches a fatty acid, palmitate, to cysteine residues and regulates protein trafficking, membrane localization and signaling activities. Using activity-based chemical probes, we discovered that human TEADs possess intrinsic palmitoylating enzyme-like activities and undergo autopalmitoylation at evolutionarily conserved cysteine residues under physiological conditions. We determined the crystal structures of lipid-bound TEADs and found that the lipid chain of palmitate inserts into a conserved deep hydrophobic pocket. Strikingly, palmitoylation did not alter TEAD's localization, but it was required for TEAD's binding to YAP and TAZ and was dispensable for its binding to the Vgll4 tumor suppressor. Moreover, palmitoylation-deficient TEAD mutants impaired TAZ-mediated muscle differentiation in vitro and tissue overgrowth mediated by the Drosophila YAP homolog Yorkie in vivo. Our study directly links autopalmitoylation to the transcriptional regulation of the Hippo pathway.

  5. The CRTC1-SIK1 Pathway Regulates Entrainment of the Circadian Clock

    PubMed Central

    Jagannath, Aarti; Butler, Rachel; Godinho, Sofia I.H.; Couch, Yvonne; Brown, Laurence A.; Vasudevan, Sridhar R.; Flanagan, Kevin C.; Anthony, Daniel; Churchill, Grant C.; Wood, Matthew J.A.; Steiner, Guido; Ebeling, Martin; Hossbach, Markus; Wettstein, Joseph G.; Duffield, Giles E.; Gatti, Silvia; Hankins, Mark W.; Foster, Russell G.; Peirson, Stuart N.

    2013-01-01

    Summary Retinal photoreceptors entrain the circadian system to the solar day. This photic resetting involves cAMP response element binding protein (CREB)-mediated upregulation of Per genes within individual cells of the suprachiasmatic nuclei (SCN). Our detailed understanding of this pathway is poor, and it remains unclear why entrainment to a new time zone takes several days. By analyzing the light-regulated transcriptome of the SCN, we have identified a key role for salt inducible kinase 1 (SIK1) and CREB-regulated transcription coactivator 1 (CRTC1) in clock re-setting. An entrainment stimulus causes CRTC1 to coactivate CREB, inducing the expression of Per1 and Sik1. SIK1 then inhibits further shifts of the clock by phosphorylation and deactivation of CRTC1. Knockdown of Sik1 within the SCN results in increased behavioral phase shifts and rapid re-entrainment following experimental jet lag. Thus SIK1 provides negative feedback, acting to suppress the effects of light on the clock. This pathway provides a potential target for the regulation of circadian rhythms. PMID:23993098

  6. The CRTC1-SIK1 pathway regulates entrainment of the circadian clock.

    PubMed

    Jagannath, Aarti; Butler, Rachel; Godinho, Sofia I H; Couch, Yvonne; Brown, Laurence A; Vasudevan, Sridhar R; Flanagan, Kevin C; Anthony, Daniel; Churchill, Grant C; Wood, Matthew J A; Steiner, Guido; Ebeling, Martin; Hossbach, Markus; Wettstein, Joseph G; Duffield, Giles E; Gatti, Silvia; Hankins, Mark W; Foster, Russell G; Peirson, Stuart N

    2013-08-29

    Retinal photoreceptors entrain the circadian system to the solar day. This photic resetting involves cAMP response element binding protein (CREB)-mediated upregulation of Per genes within individual cells of the suprachiasmatic nuclei (SCN). Our detailed understanding of this pathway is poor, and it remains unclear why entrainment to a new time zone takes several days. By analyzing the light-regulated transcriptome of the SCN, we have identified a key role for salt inducible kinase 1 (SIK1) and CREB-regulated transcription coactivator 1 (CRTC1) in clock re-setting. An entrainment stimulus causes CRTC1 to coactivate CREB, inducing the expression of Per1 and Sik1. SIK1 then inhibits further shifts of the clock by phosphorylation and deactivation of CRTC1. Knockdown of Sik1 within the SCN results in increased behavioral phase shifts and rapid re-entrainment following experimental jet lag. Thus SIK1 provides negative feedback, acting to suppress the effects of light on the clock. This pathway provides a potential target for the regulation of circadian rhythms.

  7. Spatial Regulation of Root Growth: Placing the Plant TOR Pathway in a Developmental Perspective

    PubMed Central

    Barrada, Adam; Montané, Marie-Hélène; Robaglia, Christophe; Menand, Benoît

    2015-01-01

    Plant cells contain specialized structures, such as a cell wall and a large vacuole, which play a major role in cell growth. Roots follow an organized pattern of development, making them the organs of choice for studying the spatio-temporal regulation of cell proliferation and growth in plants. During root growth, cells originate from the initials surrounding the quiescent center, proliferate in the division zone of the meristem, and then increase in length in the elongation zone, reaching their final size and differentiation stage in the mature zone. Phytohormones, especially auxins and cytokinins, control the dynamic balance between cell division and differentiation and therefore organ size. Plant growth is also regulated by metabolites and nutrients, such as the sugars produced by photosynthesis or nitrate assimilated from the soil. Recent literature has shown that the conserved eukaryotic TOR (target of rapamycin) kinase pathway plays an important role in orchestrating plant growth. We will summarize how the regulation of cell proliferation and cell expansion by phytohormones are at the heart of root growth and then discuss recent data indicating that the TOR pathway integrates hormonal and nutritive signals to orchestrate root growth. PMID:26295391

  8. Nek2A/SuFu feedback loop regulates Gli-mediated Hedgehog signaling pathway

    PubMed Central

    Zhou, Fen; Huang, Dengliang; Li, Yong; Hu, Guanghui; Rao, Hai; Lu, Quqin; Luo, Shiwen; Wang, Yao

    2017-01-01

    Suppressor of Fused (SuFu), one of the most conserved components of the Hedgehog (Hh) signaling, binds Gli transcription factors and impedes activation of target gene expression in mammalian cells. Despite the central importance of SuFu in the Hh pathway, little is known about SuFu regulation. In a previous study, we identified NIMA-related expressed kinase 2A (Nek2A) as a SuFu-interacting protein. Here, we show that Nek2A stabilizes SuFu through impairing ubiquitin/proteasome degradation of SuFu. In addition, Nek2A negatively regulates target genes of Hh signaling as well as Gli2 transcriptional activity. In turn, inhibition of Hh signaling by GANT61 diminishes mRNA and protein levels of Nek2A, and Hh agonist promotes transcription of NEK2A gene. Chromatin immunoprecipitation assays revealed that Gli1 and Gli2 directly bind to the promoter regions of NEK2A gene and induced its transcription. Thus, we uncovered one of the mechanisms by which Nek2A acts as a modulator of the Hh signaling pathway in the context of a novel negative-feedback loop, which may offer new insights into Gli-mediated Hh signaling regulation in development and human diseases. PMID:28035348

  9. A nuclear receptor-like pathway regulating multidrug resistance in fungi.

    PubMed

    Thakur, Jitendra K; Arthanari, Haribabu; Yang, Fajun; Pan, Shih-Jung; Fan, Xiaochun; Breger, Julia; Frueh, Dominique P; Gulshan, Kailash; Li, Darrick K; Mylonakis, Eleftherios; Struhl, Kevin; Moye-Rowley, W Scott; Cormack, Brendan P; Wagner, Gerhard; Näär, Anders M

    2008-04-03

    Multidrug resistance (MDR) is a serious complication during treatment of opportunistic fungal infections that frequently afflict immunocompromised individuals, such as transplant recipients and cancer patients undergoing cytotoxic chemotherapy. Improved knowledge of the molecular pathways controlling MDR in pathogenic fungi should facilitate the development of novel therapies to combat these intransigent infections. MDR is often caused by upregulation of drug efflux pumps by members of the fungal zinc-cluster transcription-factor family (for example Pdr1p orthologues). However, the molecular mechanisms are poorly understood. Here we show that Pdr1p family members in Saccharomyces cerevisiae and the human pathogen Candida glabrata directly bind to structurally diverse drugs and xenobiotics, resulting in stimulated expression of drug efflux pumps and induction of MDR. Notably, this is mechanistically similar to regulation of MDR in vertebrates by the PXR nuclear receptor, revealing an unexpected functional analogy of fungal and metazoan regulators of MDR. We have also uncovered a critical and specific role of the Gal11p/MED15 subunit of the Mediator co-activator and its activator-targeted KIX domain in antifungal/xenobiotic-dependent regulation of MDR. This detailed mechanistic understanding of a fungal nuclear receptor-like gene regulatory pathway provides novel therapeutic targets for the treatment of multidrug-resistant fungal infections.

  10. MicroRNA-221 and -222 Regulate Radiation Sensitivity by Targeting the PTEN Pathway

    SciTech Connect

    Zhang Chunzhi; Kang Chunsheng; Wang Ping; Cao Yongzhen; Lv Zhonghong; Yu Shizhu; Wang Guangxiu; Zhang Anling; Jia Zhifan; Han Lei; Yang Chunying; Ishiyama, Hiromichi; Teh, Bin S.; Xu Bo; Pu Peiyu

    2011-05-01

    Purpose: MicroRNAs (miRNAs) are noncoding RNAs inhibiting expression of numerous target genes by posttranscriptional regulation. miRNA-221 and miRNA-222 (miRNA-221/-222) expression is elevated in radioresistant tumor cell lines; however, it is not known whether and how miRNAs control cellular responses to ionizing irradiation. Methods and Materials: We used bioinformatic analyses, luciferase reporter assay, and genetic knockdown and biochemical assays to characterize the regulation pathways of miRNA-221/-222 in response to radiation treatment. Results: We identified the PTEN gene as a target of miRNA-221/-222. Furthermore, we found that knocking down miRNA-221/-222 by antisense oligonucleotides upregulated PTEN expression. Upregulated PTEN expression suppressed AKT activity and increased radiation-induced apoptosis, resulting in enhancement of radiosensitivity in tumor cells. Conclusions: miRNA-221/-222 control radiation sensitivity by regulating the PTEN/AKT pathway and can be explored as novel targets for radiosensitization.

  11. Drosophila Mitf regulates the V-ATPase and the lysosomal-autophagic pathway

    PubMed Central

    Bouché, Valentina; Espinosa, Alma Perez; Leone, Luigi; Sardiello, Marco; Ballabio, Andrea; Botas, Juan

    2016-01-01

    ABSTRACT An evolutionarily conserved gene network regulates the expression of genes involved in lysosome biogenesis, autophagy, and lipid metabolism. In mammals, TFEB and other members of the MiTF-TFE family of transcription factors control this network. Here we report that the lysosomal-autophagy pathway is controlled by Mitf gene in Drosophila melanogaster. Mitf is the single MiTF-TFE family member in Drosophila and prior to this work was known only for its function in eye development. We show that Mitf regulates the expression of genes encoding V-ATPase subunits as well as many additional genes involved in the lysosomal-autophagy pathway. Reduction of Mitf function leads to abnormal lysosomes and impairs autophagosome fusion and lipid breakdown during the response to starvation. In contrast, elevated Mitf levels increase the number of lysosomes, autophagosomes and autolysosomes, and decrease the size of lipid droplets. Inhibition of Drosophila MTORC1 induces Mitf translocation to the nucleus, underscoring conserved regulatory mechanisms between Drosophila and mammalian systems. Furthermore, we show Mitf-mediated clearance of cytosolic and nuclear expanded ATXN1 (ataxin 1) in a cellular model of spinocerebellar ataxia type 1 (SCA1). This remarkable observation illustrates the potential of the lysosomal-autophagy system to prevent toxic protein aggregation in both the cytoplasmic and nuclear compartments. We anticipate that the genetics of the Drosophila model and the absence of redundant MIT transcription factors will be exploited to investigate the regulation and function of the lysosomal-autophagy gene network. PMID:26761346

  12. UHRF1 regulation of the Keap1-Nrf2 pathway in pancreatic cancer contributes to oncogenesis.

    PubMed

    Abu-Alainin, Wafa; Gana, Thompson; Liloglou, Triantafillos; Olayanju, Adedamola; Barrera, Lawrence N; Ferguson, Robert; Campbell, Fiona; Andrews, Timothy; Goldring, Christopher; Kitteringham, Neil; Park, Brian K; Nedjadi, Taoufik; Schmid, Michael C; Slupsky, Joseph R; Greenhalf, William; Neoptolemos, John P; Costello, Eithne

    2016-02-01

    The cellular defence protein Nrf2 is a mediator of oncogenesis in pancreatic ductal adenocarcinoma (PDAC) and other cancers. However, the control of Nrf2 expression and activity in cancer is not fully understood. We previously reported the absence of Keap1, a pivotal regulator of Nrf2, in ∼70% of PDAC cases. Here we describe a novel mechanism whereby the epigenetic regulator UHRF1 suppresses Keap1 protein levels. UHRF1 expression was observed in 20% (5 of 25) of benign pancreatic ducts compared to 86% (114 of 132) of pancreatic tumours, and an inverse relationship between UHRF1 and Keap1 levels in PDAC tumours (n = 124) was apparent (p = 0.002). We also provide evidence that UHRF1-mediated regulation of the Nrf2 pathway contributes to the aggressive behaviour of PDAC. Depletion of UHRF1 from PDAC cells decreased growth and enhanced apoptosis and cell cycle arrest. UHRF1 depletion also led to reduced levels of Nrf2-regulated downstream proteins and was accompanied by heightened oxidative stress, in the form of lower glutathione levels and increased reactive oxygen species. Concomitant depletion of Keap1 and UHRF1 restored Nrf2 levels and reversed cell cycle arrest and the increase in reactive oxygen species. Mechanistically, depletion of UHRF1 reduced global and tumour suppressor promoter methylation in pancreatic cancer cell lines, and KEAP1 gene promoter methylation was reduced in one of three cell lines examined. Thus, methylation of the KEAP1 gene promoter may contribute to the suppression of Keap1 protein levels by UHRF1, although our data suggest that additional mechanisms need to be explored. Finally, we demonstrate that K-Ras drives UHRF1 expression, establishing a novel link between this oncogene and Nrf2-mediated cellular protection. Since UHRF1 over-expression occurs in other cancers, its ability to regulate the Keap1-Nrf2 pathway may be critically important to the malignant behaviour of these cancers.

  13. Ellagic acid checks lymphoma promotion via regulation of PKC signaling pathway.

    PubMed

    Mishra, Sudha; Vinayak, Manjula

    2013-02-01

    Protein Kinase C (PKC) isozymes are key components involved in cell proliferation and their over activation leads to abnormal tumor growth. PKC follows signalling pathway by activation of downstream gene NF-kB and early transcription factor c-Myc. Over activation of NF-kB and c-Myc gene are also linked with unregulated proliferation of cancer cells. Therefore any agent which can inhibit the activation of Protein kinase C, NF-kB and c-Myc may be useful in reducing cancer progression. To investigate this hypothesis we have tested the effect of ellagic acid on these genes in Dalton's lymphoma bearing (DL). The role of ellagic acid was also tested in regulation of tumor suppressor gene Transforming growth factor-β1 (TGF-β1). DL mice were treated with three different doses (40, 60 and 80 mg/kg body weight) of ellagic acid. Ascites cells of mice were used for the experiments. Ellagic acid administration to DL mice decreased oxidative stress by reducing lipid peroxidation. Ellagic acid also down regulates the expression of classical isozymes of PKC i.e. PKCα, PKCβ, and PKCγ as well as activity of total PKC and NF-kB, indicating its antitumor action. The anticarcinogenic action of ellagic acid was also confirmed by up regulation of TGF-β1 and down regulation of c-Myc. Lymphoma prevention by ellagic acid is further supported by decrease in cell proliferation, cell viability, ascites fluid accumulation and increase in life span of DL mice. All these findings suggest that ellagic acid prevents the cancer progression by down regulation of PKC signaling pathway leading to cell proliferation.

  14. Oncogenic microRNA-4534 regulates PTEN pathway in prostate cancer

    PubMed Central

    Nip, Hannah; Dar, Altaf A.; Saini, Sharanjot; Colden, Melissa; Varahram, Shahryari; Chowdhary, Harshika; Yamamura, Soichiro; Mitsui, Yozo; Tanaka, Yuichiro; Kato, Taku; Hashimoto, Yutaka; Shiina, Marisa; Kulkarni, Priyanka; Dasgupta, Pritha; Imai-Sumida, Mitsuho; Tabatabai, Z. Laura; Greene, Kirsten; Deng, Guoren; Dahiya, Rajvir; Majid, Shahana

    2016-01-01

    Prostate carcinogenesis involves alterations in several signaling pathways, the most prominent being the PI3K/AKT pathway. This pathway is constitutively active and drives prostate cancer (PCa) progression to advanced metastatic disease. PTEN, a critical tumor and metastasis suppressor gene negatively regulates cell survival, proliferation, migration and angiogenesis via the PI3K/Akt pathway. PTEN is mutated, downregulated/dysfunctional in many cancers and its dysregulation correlates with poor prognosis in PCa. Here, we demonstrate that microRNA-4534 (miR-4534) is overexpressed in PCa and show that miR-4534 is hypermethylated in normal tissues and cell lines compared to PCa tissues/cells. miR-4534 exerts its oncogenic effects partly by downregulating the tumor suppressor PTEN gene. Knockdown of miR-4534 impaired cell proliferation, migration/invasion and induced G0/G1 cell cycle arrest and apoptosis in PCa. Suppression of miR-4534 and its effects on tumor growth was confirmed in a xenograft mouse model. We performed parallel experiments in non-cancer RWPE1 cells by overexpessing miR-4534 followed by functional assays. Overexpression of miR-4534 induced pro-cancerous characteristics in this non-cancer cell line. Statistical analyses revealed that miR-4534 has potential to independently distinguish malignant from normal tissues and positively correlated with poor overall and PSA recurrence free survival. Taken together, our results show that depletion of miR-4534 in PCa induces a tumor suppressor phenotype partly through induction of PTEN. These results have important implications for identifying and defining the role of new PTEN regulators such as microRNAs in prostate tumorigenesis. Understanding aberrantly overexpressed miR-4534 and its downregulation of PTEN will provide mechanistic insight and therapeutic targets for PCa therapy. PMID:27634912

  15. Multiple genetic pathways regulate replicative senescence in telomerase-deficient yeast

    PubMed Central

    Ballew, Bari J.; Lundblad, Victoria

    2013-01-01

    Summary Most human tissues express low levels of telomerase and undergo telomere shortening and eventual senescence; the resulting limitation on tissue renewal can lead to a wide range of age-dependent pathophysiologies. Increasing evidence indicates that the decline in cell division capacity in cells that lack telomerase can be influenced by numerous genetic factors. Here, we use telomerase-defective strains of budding yeast to probe whether replicative senescence can be attenuated or accelerated by defects in factors previously implicated in handling of DNA termini. We show that the MRX (Mre11-Rad50-Xrs2) complex, as well as negative (Rif2) and positive (Tel1) regulators of this complex, comprise a single pathway that promotes replicative senescence, in a manner that recapitulates how these proteins modulate resection of DNA ends. In contrast, the Rad51 recombinase, which acts downstream of the MRX complex in double-strand break (DSB) repair, regulates replicative senescence through a separate pathway operating in opposition to the MRX-Tel1-Rif2 pathway. Moreover, defects in several additional proteins implicated in DSB repair (Rif1 and Sae2) confer only transient effects during early or late stages of replicative senescence, respectively, further suggesting that a simple analogy between DSBs and eroding telomeres is incomplete. These results indicate that the replicative capacity of telomerase-defective yeast is controlled by a network comprised of multiple pathways. It is likely that telomere shortening in telomerase-depleted human cells is similarly under a complex pattern of genetic control; mechanistic understanding of this process should provide crucial information regarding how human tissues age in response to telomere erosion. PMID:23672410

  16. The Nuclear Factor-kB Pathway Regulates Cytochrome P450 3A4 Protein Stability

    SciTech Connect

    Zangar, Richard C.; Bollinger, Nikki; Verma, Seema; Karin, Norm J.; Lu, Yi

    2008-06-01

    We have previously observed that CYP3A4 protein levels are suppressed by inhibition of the proteasome in primary cultured hepatocytes. Because this result is opposite of what would be expected if CYP3A4 is degraded by the proteasome, it seems likely that there is another protein that is susceptible to proteasomal degradation that regulates CYP3A4 expression. In this study, we evaluate whether the nuclear factor kappa B (NF-kB) pathway is involved in that process. Our model system uses an adenovirus system to express CYP3A4 protein in HepG2 cells, which are derived from human cancer cells. Similar to results in primary hepatocytes, we found that inhibition of the proteasome with MG132 suppresses CYP3A4. Consistent with reports that proteasome inhibition suppresses the NF-kB pathway, we also observe a suppression of inhibitory kB kinase protein levels after treatment with MG132. Treatment of the HepG2 cells with NK-kB Activation Inhibitor also suppresses CYP3A4 proteins levels. In contrast, inhibition of either the proteasome or NF-kB pathways increases CYP3A4 mRNA levels. When the HepG2 cells are treated with cycloheximide, a general inhibitor of translation, the loss of CYP3A4 protein is accelerated by co-treatment with an NF-kB Activation Inhibitor. These results indicate that NF-kB activity regulates CYP3A4 protein stability and suggest that the NF-kB pathway is responsible for the decrease in CYP3A4 protein levels that results from the inhibition of proteasomal activity.

  17. Oncogenic microRNA-4534 regulates PTEN pathway in prostate cancer.

    PubMed

    Nip, Hannah; Dar, Altaf A; Saini, Sharanjot; Colden, Melissa; Varahram, Shahryari; Chowdhary, Harshika; Yamamura, Soichiro; Mitsui, Yozo; Tanaka, Yuichiro; Kato, Taku; Hashimoto, Yutaka; Shiina, Marisa; Kulkarni, Priyanka; Dasgupta, Pritha; Imai-Sumida, Mitsuho; Tabatabai, Z Laura; Greene, Kirsten; Deng, Guoren; Dahiya, Rajvir; Majid, Shahana

    2016-10-18

    Prostate carcinogenesis involves alterations in several signaling pathways, the most prominent being the PI3K/AKT pathway. This pathway is constitutively active and drives prostate cancer (PCa) progression to advanced metastatic disease. PTEN, a critical tumor and metastasis suppressor gene negatively regulates cell survival, proliferation, migration and angiogenesis via the PI3K/Akt pathway. PTEN is mutated, downregulated/dysfunctional in many cancers and its dysregulation correlates with poor prognosis in PCa. Here, we demonstrate that microRNA-4534 (miR-4534) is overexpressed in PCa and show that miR-4534 is hypermethylated in normal tissues and cell lines compared to PCa tissues/cells. miR-4534 exerts its oncogenic effects partly by downregulating the tumor suppressor PTEN gene. Knockdown of miR-4534 impaired cell proliferation, migration/invasion and induced G0/G1 cell cycle arrest and apoptosis in PCa. Suppression of miR-4534 and its effects on tumor growth was confirmed in a xenograft mouse model. We performed parallel experiments in non-cancer RWPE1 cells by overexpessing miR-4534 followed by functional assays. Overexpression of miR-4534 induced pro-cancerous characteristics in this non-cancer cell line. Statistical analyses revealed that miR-4534 has potential to independently distinguish malignant from normal tissues and positively correlated with poor overall and PSA recurrence free survival. Taken together, our results show that depletion of miR-4534 in PCa induces a tumor suppressor phenotype partly through induction of PTEN. These results have important implications for identifying and defining the role of new PTEN regulators such as microRNAs in prostate tumorigenesis. Understanding aberrantly overexpressed miR-4534 and its downregulation of PTEN will provide mechanistic insight and therapeutic targets for PCa therapy.

  18. p38 Mitogen-Activated Protein Kinase Pathway Regulates Genes during Proliferation and Differentiation in Oligodendrocytes

    PubMed Central

    Haines, Jeffery D.; Fulton, Debra L.; Richard, Stephane; Almazan, Guillermina

    2015-01-01

    We have previously shown that p38 mitogen-activated protein kinase (p38 MAPK) is important for oligodendrocyte (OLG) differentiation and myelination. However, the precise cellular mechanisms by which p38 regulates OLG differentiation remain largely unknown. To determine whether p38 functions in part through transcriptional events in regulating OLG identity, we performed microarray analysis on differentiating oligodendrocyte progenitors (OLPs) treated with a p38 inhibitor. Consistent with a role in OLG differentiation, pharmacological inhibition of p38 down-regulated the transcription of genes that are involved in myelin biogenesis, transcriptional control and cell cycle. Proliferation assays showed that OLPs treated with the p38 inhibitor retained a proliferative capacity which could be induced upon application of mitogens demonstrating that after two days of p38-inhibition OLGs remained poised to continue mitosis. Together, our results suggest that the p38 pathway regulates gene transcription which can coordinate OLG differentiation. Our microarray dataset will provide a useful resource for future studies investigating the molecular mechanisms by which p38 regulates oligodendrocyte differentiation and myelination. PMID:26714323

  19. THE E2/FRB PATHWAY REGULATION OF DNA REPLICATION AND PROTEIN BIOSYNTHESIS

    EPA Science Inventory

    The E2F/Rb pathway plays a pivotal role in the control of cell cycle progression and regulates the expression of genes required for Gl/S transition. Our study examines the genomic response in Drosophila embryos after overexpression and mutation of E2F/Rb pathway molecules. Hierar...

  20. Leptin regulated calcium channels of neuropeptide Y and proopiomelanocortin neurons by activation of different signal pathways.

    PubMed

    Wang, J-H; Wang, F; Yang, M-J; Yu, D-F; Wu, W-N; Liu, J; Ma, L-Q; Cai, F; Chen, J-G

    2008-09-22

    The fat-derived hormone leptin regulates food intake and body weight in part by modulating the activity of neuropeptide Y (NPY) and proopiomelanocortin (POMC) neurons in the hypothalamic arcuate nucleus (ARC). To investigate the electrophysiological activity of these neurons and their responses to leptin, we recorded whole-cell calcium currents on NPY and POMC neurons in the ARC of rats, which we identified by morphologic features and immunocytochemical identification at the end of recording. Leptin decreased the peak amplitude of high voltage-activated calcium currents (I(HVA)) in the isolated neurons from ARC, which were subsequently shown to be immunoreactive for NPY. The inhibition was prevented by pretreatment with inhibitors of Janus kinase 2 (JAK2) and mitogen-activated protein kinases (MAPK). In contrast, leptin increased the amplitude of I(HVA) in POMC-containing neurons. The stimulations of I(HVA) were inhibited by blockers of JAK2 and phosphatidylino 3-kinase (PI3-k). Both of these effects were counteracted by the L-type calcium channel antagonist nifedipine, suggesting that L-type calcium channels were involved in the regulation induced by leptin. These data indicated that leptin exerted opposite effects on these two classes of neurons. Leptin directly inhibited I(HVA) in NPY neurons via leptin receptor (LEPR) -JAK2-MAPK pathways, whereas evoked I(HVA) in POMC neurons by LEPR-JAK2-PI3-k pathways. These neural pathways and intracellular signaling mechanisms may play key roles in regulating NPY and POMC neuron activity, anorectic action of leptin and, thereby, feeding.

  1. Identification of cisplatin-regulated metabolic pathways in pluripotent stem cells.

    PubMed

    von Stechow, Louise; Ruiz-Aracama, Ainhoa; van de Water, Bob; Peijnenburg, Ad; Danen, Erik; Lommen, Arjen

    2013-01-01

    The chemotherapeutic compound, cisplatin causes various kinds of DNA lesions but also triggers other pertubations, such as ER and oxidative stress. We and others have shown that treatment of pluripotent stem cells with cisplatin causes a plethora of transcriptional and post-translational alterations that, to a major extent, point to DNA damage response (DDR) signaling. The orchestrated DDR signaling network is important to arrest the cell cycle and repair the lesions or, in case of damage beyond repair, eliminate affected cells. Failure to properly balance the various aspects of the DDR in stem cells contributes to ageing and cancer. Here, we performed metabolic profiling by mass spectrometry of embryonic stem (ES) cells treated for different time periods with cisplatin. We then integrated metabolomics with transcriptomics analyses and connected cisplatin-regulated metabolites with regulated metabolic enzymes to identify enriched metabolic pathways. These included nucleotide metabolism, urea cycle and arginine and proline metabolism. Silencing of identified proline metabolic and catabolic enzymes indicated that altered proline metabolism serves as an adaptive, rather than a toxic response. A group of enriched metabolic pathways clustered around the metabolite S-adenosylmethionine, which is a hub for methylation and transsulfuration reactions and polyamine metabolism. Enzymes and metabolites with pro- or anti-oxidant functions were also enriched but enhanced levels of reactive oxygen species were not measured in cisplatin-treated ES cells. Lastly, a number of the differentially regulated metabolic enzymes were identified as target genes of the transcription factor p53, pointing to p53-mediated alterations in metabolism in response to genotoxic stress. Altogether, our findings reveal interconnecting metabolic pathways that are responsive to cisplatin and may serve as signaling modules in the DDR in pluripotent stem cells.

  2. Regulation of the STARS signaling pathway in response to endurance and resistance exercise and training.

    PubMed

    Lamon, Séverine; Wallace, Marita A; Stefanetti, Renae J; Rahbek, Stine K; Vendelbo, Mikkel H; Russell, Aaron P; Vissing, Kristian

    2013-09-01

    The striated muscle activator of Rho signaling (STARS) protein and members of its downstream signaling pathway, including myocardin-related transcription factor-A (MRTF-A) and SRF, are increased in response to prolonged resistance exercise training but also following a single bout of endurance cycling. The aim of the present study was to measure and compare the regulation of STARS, MRTF-A and SRF mRNA and protein following 10 weeks of endurance training (ET) versus resistance training (RT), as well as before and following a single bout of endurance (EE) versus resistance exercise (RE). Following prolonged training, STARS, MRTF-A and SRF mRNA levels were all increased by similar magnitude, irrespective of training type. In the training-habituated state, STARS mRNA increased following a single-bout RE when measured 2.5 and 5 h post-exercise and had returned to resting level by 22 h following exercise. MRTF-A and SRF mRNA levels were decreased by 2.5, 5, and 22 h following a single bout of RE and EE exercise when compared to their respective basal levels, with no significant difference seen between the groups at any of the time points. No changes in protein levels were observed following the two modes of exercise training or a single bout of exercise. This study demonstrates that the stress signals elicited by ET and RT result in a comparable regulation of members of the STARS pathway. In contrast, a single bout of EE and RE, performed in the trained state, elicit different responses. These observations suggest that in the trained state, the acute regulation of the STARS pathway following EE or RE may be responsible for exercise-specific muscle adaptations.

  3. Salinity-induced regulation of the myo-inositol biosynthesis pathway in tilapia gill epithelium.

    PubMed

    Sacchi, Romina; Li, Johnathon; Villarreal, Fernando; Gardell, Alison M; Kültz, Dietmar

    2013-12-15

    The myo-inositol biosynthesis (MIB) pathway converts glucose-6-phosphate to the compatible osmolyte myo-inositol that protects cells from osmotic stress. Using proteomics, the enzymes that constitute the MIB pathway, myo-inositol phosphate synthase (MIPS) and inositol monophosphatase 1 (IMPA1), are identified in tilapia (Oreochromis mossambicus) gill epithelium. Targeted, quantitative, label-free proteomics reveals that they are both upregulated during salinity stress. Upregulation is stronger when fish are exposed to severe (34 ppt acute and 90 ppt gradual) relative to moderate (70 ppt gradual) salinity stress. IMPA1 always responds more strongly than MIPS, suggesting that MIPS is more stable during salinity stress. MIPS is N-terminally acetylated and the corresponding peptide increases proportionally to MIPS protein, while non-acetylated N-terminal peptide is not detectable, indicating that MIPS acetylation is constitutive and may serve to stabilize the protein. Hyperosmotic induction of MIPS and IMPA1 is confirmed using western blot and real-time qPCR and is much higher at the mRNA than at the protein level. Two distinct MIPS mRNA variants are expressed in the gill, but one is more strongly regulated by salinity than the other. A single MIPS gene is encoded in the tilapia genome whereas the zebrafish genome lacks MIPS entirely. The genome of euryhaline tilapia contains four IMPA genes, two of which are expressed, but only one is salinity regulated in gill epithelium. The genome of stenohaline zebrafish contains a single IMPA gene. We conclude that the MIB pathway represents a major salinity stress coping mechanism that is regulated at multiple levels in euryhaline fish but absent in stenohaline zebrafish.

  4. Reciprocal Transcriptional Regulation of Metabolic and Signaling Pathways Correlates with Disease Severity in Heart Failure

    PubMed Central

    Barth, Andreas S; Kumordzie, Ami; Frangakis, Constantine; Margulies, Kenneth B; Cappola, Thomas P; Tomaselli, Gordon F

    2012-01-01

    Background Systolic heart failure (HF) is a complex systemic syndrome that can result from a wide variety of clinical conditions and gene mutations. Despite phenotypic similarities, characterized by ventricular dilatation and reduced contractility, the extent of common and divergent gene expression between different forms of HF remains a matter of intense debate. Methods and Results Using a meta-analysis of 28 experimental (mouse, rat, dog) and human HF microarray studies, we demonstrate that gene expression changes are characterized by a coordinated and reciprocal regulation of major metabolic and signaling pathways. In response to a wide variety of stressors in animal models of HF, including ischemia, pressure overload, tachypacing, chronic isoproterenol infusion, Chagas disease, and transgenic mouse models, major metabolic pathways are invariably downregulated, while cell signaling pathways are upregulated. In contrast to this uniform transcriptional pattern observed in experimental animal models which recapitulates a fetal gene expression program, human HF microarray studies displayed a greater heterogeneity, with some studies even showing upregulation of metabolic and downregulation of signaling pathways in end-stage human hearts. These discrepant results between animal and human studies are due to a number of factors, prominently cardiac disease and variable exposure to cold cardioplegic solution in non-failing human samples which can downregulate transcripts involved in oxidative phosphorylation (OXPHOS) within the first 6h, thus mimicking gene expression patterns observed in failing samples. Additionally, beta-blockers and ACE-inhibitor use in end-stage human HF was associated with higher levels of myocardial OXPHOS transcripts, thus partially reversing the fetal gene expression pattern. In human failing samples, downregulation of metabolism was associated with hemodynamic markers of disease severity. Conclusions Irrespective of the etiology, gene

  5. MicroRNA regulatory pathway analysis identifies miR-142-5p as a negative regulator of TGF-β pathway via targeting SMAD3

    PubMed Central

    Huang, Wei; Liu, Hui; Zhang, Hong-Mei; Li, Qiubai; Chen, Zhichao; Guo, An-Yuan

    2016-01-01

    MicroRNAs (miRNAs) are non-coding RNAs with functions of posttranscriptional regulation. The abnormally expressed miRNAs have been shown to be crucial contributors and may serve as biomarkers in many diseases. However, determining the biological function of miRNAs is an ongoing challenge. By combining miRNA targets prediction, miRNA and mRNA expression profiles in TCGA cancers, and pathway data, we performed a miRNA-pathway regulation inference by Fisher's exact test for enrichment analysis. Then we constructed a database to show the cancer related miRNA-pathway regulatory network (http://bioinfo.life.hust.edu.cn/miR_path). As one of the miRNAs targeting many cancer related pathways, miR-142-5p potentially regulates the maximum number of genes in TGF-β signaling pathway. We experimentally confirmed that miR-142-5p directly targeted and suppressed SMAD3, a key component in TGF-β signaling. Ectopic overexpression of miR-142-5p significantly promoted tumor cell proliferation and inhibited apoptosis, while silencing of miR-142-5p inhibited the tumor cell proliferation and promoted apoptosis in vitro. These findings indicate that miR-142-5p plays as a negative regulator in TGF-β pathway by targeting SMAD3 and suppresses TGF-β-induced growth inhibition in cancer cells. Our study proved the feasibility of miRNA regulatory pathway analysis and shed light on combining bioinformatics with experiments in the research of complex diseases. PMID:27683030

  6. RNAseq Analysis of the Parasitic Nematode Strongyloides stercoralis Reveals Divergent Regulation of Canonical Dauer Pathways

    PubMed Central

    Stoltzfus, Jonathan D.; Minot, Samuel; Berriman, Matthew; Nolan, Thomas J.; Lok, James B.

    2012-01-01

    The infectious form of many parasitic nematodes, which afflict over one billion people globally, is a developmentally arrested third-stage larva (L3i). The parasitic nematode Strongyloides stercoralis differs from other nematode species that infect humans, in that its life cycle includes both parasitic and free-living forms, which can be leveraged to investigate the mechanisms of L3i arrest and activation. The free-living nematode Caenorhabditis elegans has a similar developmentally arrested larval form, the dauer, whose formation is controlled by four pathways: cyclic GMP (cGMP) signaling, insulin/IGF-1-like signaling (IIS), transforming growth factor β (TGFβ) signaling, and biosynthesis of dafachronic acid (DA) ligands that regulate a nuclear hormone receptor. We hypothesized that homologous pathways are present in S. stercoralis, have similar developmental regulation, and are involved in L3i arrest and activation. To test this, we undertook a deep-sequencing study of the polyadenylated transcriptome, generating over 2.3 billion paired-end reads from seven developmental stages. We constructed developmental expression profiles for S. stercoralis homologs of C. elegans dauer genes identified by BLAST searches of the S. stercoralis genome as well as de novo assembled transcripts. Intriguingly, genes encoding cGMP pathway components were coordinately up-regulated in L3i. In comparison to C. elegans, S. stercoralis has a paucity of genes encoding IIS ligands, several of which have abundance profiles suggesting involvement in L3i development. We also identified seven S. stercoralis genes encoding homologs of the single C. elegans dauer regulatory TGFβ ligand, three of which are only expressed in L3i. Putative DA biosynthetic genes did not appear to be coordinately regulated in L3i development. Our data suggest that while dauer pathway genes are present in S. stercoralis and may play a role in L3i development, there are significant differences between the two species

  7. ROS, Notch, and Wnt signaling pathways: crosstalk between three major regulators of cardiovascular biology.

    PubMed

    Caliceti, C; Nigro, P; Rizzo, P; Ferrari, R

    2014-01-01

    Reactive oxygen species (ROS), traditionally viewed as toxic by-products that cause damage to biomolecules, now are clearly recognized as key modulators in a variety of biological processes and pathological states. The development and regulation of the cardiovascular system require orchestrated activities; Notch and Wnt/β -catenin signaling pathways are implicated in many aspects of them, including cardiomyocytes and smooth muscle cells survival, angiogenesis, progenitor cells recruitment and differentiation, arteriovenous specification, vascular cell migration, and cardiac remodelling. Several novel findings regarding the role of ROS in Notch and Wnt/β-catenin modulation prompted us to review their emerging function in the cardiovascular system during embryogenesis and postnatally.

  8. The transcription factor AREB1 regulates primary metabolic pathways in tomato fruits.

    PubMed

    Bastías, Adriana; Yañez, Mónica; Osorio, Sonia; Arbona, Vicent; Gómez-Cadenas, Aurelio; Fernie, Alisdair R; Casaretto, José A

    2014-06-01

    Tomato fruit development is regulated both by the action of plant hormones and by tight genetic control. Recent studies suggest that abscisic acid (ABA) signalling may affect different aspects of fruit maturation. Previously, it was shown that SlAREB1, an ABA-regulated transcription factor involved in stress-induced responses, is expressed in seeds and in fruit tissues in tomato. Here, the role of SlAREB1 in regulating the expression of genes relevant for primary metabolic pathways and affecting the metabolic profile of the fruit was investigated using transgenic tomato lines. Metabolite profiling using gas chromatography-time of flight mass spectrometry (GC-TOF-MS) and non-targeted liquid chromatography-mass spectrometry (LC-MS) was performed on pericarp tissue from fruits harvested at three stages of fruit development. Principal component analysis of the data could distinguish the metabolite profiles of non-transgenic fruits from those that overexpress and down-regulate SlAREB1. Overexpression of SlAREB1 resulted in increased content of organic acids, hexoses, hexose-phosphates, and amino acids in immature green, mature green, and red ripe fruits, and these modifications correlated with the up-regulation of enzyme-encoding genes involved in primary carbohydrate and amino acid metabolism. A non-targeted LC-MS analysis indicated that the composition of secondary metabolites is also affected in transgenic lines. In addition, gene expression data revealed that some genes associated with fruit ripening are also up-regulated in SlAREB1-overexpressing lines compared with wild-type and antisense lines. Taken together, the results suggest that SlAREB1 participates in the regulation of the metabolic programming that takes place during fruit ripening and that may explain part of the role of ABA in fruit development in tomato.

  9. The transcription factor AREB1 regulates primary metabolic pathways in tomato fruits

    PubMed Central

    Bastías, Adriana; Osorio, Sonia; Casaretto, José A.

    2014-01-01

    Tomato fruit development is regulated both by the action of plant hormones and by tight genetic control. Recent studies suggest that abscisic acid (ABA) signalling may affect different aspects of fruit maturation. Previously, it was shown that SlAREB1, an ABA-regulated transcription factor involved in stress-induced responses, is expressed in seeds and in fruit tissues in tomato. Here, the role of SlAREB1 in regulating the expression of genes relevant for primary metabolic pathways and affecting the metabolic profile of the fruit was investigated using transgenic tomato lines. Metabolite profiling using gas chromatography–time of flight mass spectrometry (GC-TOF-MS) and non-targeted liquid chromatography–mass spectrometry (LC-MS) was performed on pericarp tissue from fruits harvested at three stages of fruit development. Principal component analysis of the data could distinguish the metabolite profiles of non-transgenic fruits from those that overexpress and down-regulate SlAREB1. Overexpression of SlAREB1 resulted in increased content of organic acids, hexoses, hexose-phosphates, and amino acids in immature green, mature green, and red ripe fruits, and these modifications correlated with the up-regulation of enzyme-encoding genes involved in primary carbohydrate and amino acid metabolism. A non-targeted LC-MS analysis indicated that the composition of secondary metabolites is also affected in transgenic lines. In addition, gene expression data revealed that some genes associated with fruit ripening are also up-regulated in SlAREB1-overexpressing lines compared with wild-type and antisense lines. Taken together, the results suggest that SlAREB1 participates in the regulation of the metabolic programming that takes place during fruit ripening and that may explain part of the role of ABA in fruit development in tomato. PMID:24659489

  10. Protein Kinase A Mediates Regulation of Gap Junctions Containing Connexin35 Through a Complex Pathway

    PubMed Central

    Ouyang, Xiaosen; Winbow, Virginia M.; Patel, Leena S.; Burr, Gary S.; Mitchell, Cheryl K.; O’Brien, John

    2008-01-01

    Connexin 35 (Cx35) is a major component of electrical synapses in the central nervous system. Many gap junctions containing Cx35 are regulated by dopamine receptor pathways that involve protein kinase A (PKA). To study the mechanism of PKA regulation, we analyzed direct phosphorylation of Cx35 by PKA in vitro, and studied the regulation of Neurobiotin tracer coupling in HeLa cells expressing Cx35 or Cx35 mutants that lack phosphorylation sites. In Cx35-transfected cells, application of the PKA activator Sp-8-cpt-cAMPS caused a significant decline in coupling, while a PKA inhibitor, Rp-8-cpt-cAMPS, significantly increased tracer coupling. In vitro phosphorylation and mutagenic analysis showed that PKA phosphorylates Cx35 directly at two major sites, Ser110 in the intracellular loop and Ser276 in the carboxyl terminus. In addition, a minor phosphorylation site in the C-terminus was identified by truncation of the last 7 amino acids at Ser298. The mutations Ser110Ala or Ser276Ala significantly reduced regulation of coupling by the PKA activator, while a combination of the two eliminated regulation. Truncation at Ser298 reversed the regulation such that the PKA activator significantly increased and the PKA inhibitor significantly decreased coupling. The activation was eliminated in the S110A,S276A,S298ter triple mutant. We conclude that PKA regulates Cx35 coupling in a complex manner that requires both major phosphorylation sites. Furthermore, the tip of the C-terminus acts as a “switch” that determines whether phosphorylation will inhibit or enhance coupling. Reliance on the combined states of three sites provides fine control over the degree of coupling through Cx35 gap junctions. PMID:15857663

  11. Characterization of the cis elements in the proximal promoter regions of the anthocyanin pathway genes reveals a common regulatory logic that governs pathway regulation

    PubMed Central

    Zhu, Zhixin; Wang, Hailong; Wang, Yiting; Guan, Shan; Wang, Fang; Tang, Jingyu; Zhang, Ruijuan; Xie, Lulu; Lu, Yingqing

    2015-01-01

    Cellular activities such as compound synthesis often require the transcriptional activation of an entire pathway; however, the molecular mechanisms underlying pathway activation have rarely been explained. Here, the cis regulatory architecture of the anthocyanin pathway genes targeted by the transcription factor (TF) complex including MYB, bHLH, and WDR was systematically analysed in one species and the findings extended to others. In Ipomoea purpurea, the IpMYB1-IpbHLH2-IpWDR1 (IpMBW) complex was found to be orthologous to the PAP1-GL3-TTG1 (AtPGT) complex of Arabidopsis thaliana, and interacted with a 7-bp MYB-recognizing element (MRE) and a 6-bp bHLH-recognizing element (BRE) at the proximal promoter region of the pathway genes. There was little transcription of the gene in the absence of the MRE or BRE. The cis elements identified experimentally converged on two syntaxes, ANCNNCC for MREs and CACN(A/C/T)(G/T) for BREs, and our bioinformatic analysis showed that these were present within anthocyanin gene promoters in at least 35 species, including both gymnosperms and angiosperms. For the anthocyanin pathway, IpMBW and AtPGT recognized the interspecific promoters of both early and later genes. In A. thaliana, the seed-specific TF complex (TT2, TT8, and TTG1) may regulate all the anthocyanin pathway genes, in addition to the proanthocyanidin-specific BAN. When multiple TF complexes in the anthocyanin pathway were compared, the cis architecture played a role larger than the TF complex in determining the variation in promoter activity. Collectively, a cis logic common to the pathway gene promoters was found, and this logic is essential for the trans factors to regulate the pathway. PMID:25911741

  12. Effective Strategies for Monitoring and Regulating Chemical Mixtures and Contaminants Sharing Pathways of Toxicity

    PubMed Central

    Venkatesan, Arjun K.; Halden, Rolf U.

    2015-01-01

    Traditionally, hazardous chemicals have been regulated in the U.S. on a one-by-one basis, an approach that is slow, expensive and can be inefficient, as illustrated by a decades-long succession of replacing one type of organohalogen flame retardants (OHFRs) with another one, without addressing the root cause of toxicity and associated public health threats posed. The present article expounds on the need for efficient monitoring strategies and pragmatic steps in reducing environmental pollution and adverse human health impacts. A promising approach is to combine specific bioassays with state-of-the-art chemical screening to identify chemicals and chemical mixtures sharing specific modes of action (MOAs) and pathways of toxicity (PoTs). This approach could be used to identify and regulate hazardous chemicals as classes or compound families, featuring similar biological end-points, such as endocrine disruption and mutagenicity. Opportunities and potential obstacles of implementing this approach are discussed. PMID:26343697

  13. Uncoupling JAK3 activation induces apoptosis in human lymphoid cancer cells via regulating critical survival pathways.

    PubMed

    Nagy, Zsuzsanna S; Ross, Jeremy A; Rodriguez, Georgialina; Bader, Julia; Dimmock, Jonathan; Kirken, Robert A

    2010-04-16

    In the current work, we report that specific inhibition of Janus tyrosine kinase (JAK3) via NC1153 induces apoptosis of certain leukemia/lymphoma cell lines. Affymetrix microarray profiling following NC1153 treatment unveiled JAK3 dependent survival modulating pathways (p53, TGF-beta, TNFR and ER stress) in Kit225 cells. IL-2 responsive NC1153 target genes were regulated in human JAK3 positive, but not in JAK3 negative lymphoid tumor cells. Moreover, primary lymphoma samples revealed that a number of these genes were reciprocally regulated during disease progression and JAK3 inhibition suggesting that downstream targets of JAK3 could be exploited in the development of novel cancer treatment regimes.

  14. Salmonella regulation of intestinal stem cells through the Wnt/β-catenin pathway

    PubMed Central

    Liu, Xingyin; Lu, Rong; Wu, Shaoping; Sun, Jun

    2010-01-01

    Recent studies have revealed that bacteria target stem cells for long-term survival in a Drosophila model. However, in mammalian models, little is known about bacterial infection and intestinal stem cells. Our study aims at understanding bacterial regulation of the intestinal stem cell in a Salmonella colitis mouse model. We found that Salmonella activates the Wnt/β-catenin signaling pathway that is known to regulate stem cells. We identified Salmonella protein AvrA that modulates Wnt signaling including upregulating Wnt expression, modifying β-catenin, increasing total β-catenin expression, and activating Wnt/β-catenin transcriptional activity in the intestinal epithelial cells. The numbers of stem cells and proliferative cells increased in the intestine infected with Salmonella expressing AvrA. Our study provides insights into bacterial infection and stem cell maintenance. PMID:20083111

  15. Cargo Sorting in the Endocytic Pathway: A Key Regulator of Cell Polarity and Tissue Dynamics

    PubMed Central

    Eaton, Suzanne; Martin-Belmonte, Fernando

    2014-01-01

    The establishment and maintenance of polarized plasma membrane domains is essential for cellular function and proper development of organisms. Epithelial cells polarize along two fundamental axes, the apicobasal and the planar, both depending on finely regulated protein trafficking mechanisms. Newly synthesized proteins destined for either surface domain are processed along the biosynthetic pathway and segregated into distinct subsets of transport carriers emanating from the trans-Golgi network or endosomes. This exocytic trafficking has been identified as essential for proper epithelial polarization. Accumulating evidence now reveals that endocytosis and endocytic recycling play an equally important role in epithelial polarization and the appropriate localization of key polarity proteins. Here, we review recent work in metazoan systems illuminating the connections between endocytosis, postendocytic trafficking, and cell polarity, both apicobasal and planar, in the formation of differentiated epithelial cells, and how these processes regulate tissue dynamics. PMID:25125399

  16. Genetic polymorphisms of RANTES, IL1-A, MCP-1 and TNF-A genes in patients with prostate cancer

    PubMed Central

    Sáenz-López, Pablo; Carretero, Rafael; Cózar, José Manuel; Romero, José Maria; Canton, Julia; Vilchez, José Ramón; Tallada, Miguel; Garrido, Federico; Ruiz-Cabello, Francisco

    2008-01-01

    Background Inflammation has been implicated as an etiological factor in several human cancers, including prostate cancer. Allelic variants of the genes involved in inflammatory pathways are logical candidates as genetic determinants of prostate cancer risk. The purpose of this study was to investigate whether single nucleotide polymorphisms of genes that lead to increased levels of pro-inflammatory cytokines and chemokines are associated with an increased prostate cancer risk. Methods A case-control study design was used to test the association between prostate cancer risk and the polymorphisms TNF-A-308 A/G (rs 1800629), RANTES-403 G/A (rs 2107538), IL1-A-889 C/T (rs 1800587) and MCP-1 2518 G/A (rs 1024611) in 296 patients diagnosed with prostate cancer and in 311 healthy controls from the same area. Results Diagnosis of prostate cancer was significantly associated with TNF-A GA + AA genotype (OR, 1.61; 95% CI, 1.09–2.64) and RANTES GA + AA genotype (OR, 1.44; 95% CI, 1.09–2.38). A alleles in TNF-A and RANTES influenced prostate cancer susceptibility and acted independently of each other in these subjects. No epistatic effect was found for the combination of different polymorphisms studied. Finally, no overall association was found between prostate cancer risk and IL1-A or MCP-1 polymorphisms. Conclusion Our results and previously published findings on genes associated with innate immunity support the hypothesis that polymorphisms in proinflammatory genes may be important in prostate cancer development. PMID:19099590

  17. Cytokinins negatively regulate the root iron uptake machinery in Arabidopsis through a growth-dependent pathway.

    PubMed

    Séguéla, Mathilde; Briat, Jean-François; Vert, Grégory; Curie, Catherine

    2008-07-01

    Plants display a number of biochemical and developmental responses to low iron availability in order to increase iron uptake from the soil. The ferric-chelate reductase FRO2 and the ferrous iron transporter IRT1 control iron entry from the soil into the root epidermis. In Arabidopsis, expression of IRT1 and FRO2 is tightly controlled to maintain iron homeostasis, and involves local and long-distance signals, as well as transcriptional and post-transcriptional events. FIT encodes a putative basic helix-loop-helix (bHLH) transcription factor that regulates iron uptake responses in Arabidopsis. Here, we uncover a new regulation of the root iron uptake genes. We show that IRT1, FRO2 and FIT are repressed by the exogenous addition of cytokinins (CKs), and that this repression acts at the level of transcript accumulation, and depends on the AHK3 and CRE1 CK receptors. The CKs and iron-deficiency signals act through distinct pathways to regulate the soil iron uptake genes, as (i) CK repression is independent of the iron status, (ii) IRT1 and FRO2 downregulation is unchanged in a fit loss-of-function mutant, indicating that FIT does not mediate CK repression, and (iii) the iron-regulated genes AtNRAMP3 and AtNRAMP4 are not downregulated by CKs. We show that root growth-inhibitory conditions, such as abiotic stresses (mannitol, NaCl) and hormonal treatments (auxin, abscissic acid), repress the iron starvation response genes. We propose that CKs control the root iron uptake machinery through a root growth dependent pathway in order to adapt nutrient uptake to the demand of the plant.

  18. YBX1 regulates tumor growth via CDC25a pathway in human lung adenocarcinoma

    PubMed Central

    Yu, Wendan; Li, Jinxiu; Tang, Zhipeng; Yu, Zhenlong; Zhao, Lei; Zhang, Yixiang; Wang, Ziyi; Wang, Peng; Li, Yechi; Li, Fengzhou; Sun, Zhe; Xuan, Yang; Tang, Ranran; Deng, Wu-guo; Guo, Wei; Gu, Chundong

    2016-01-01

    Y-box binding protein 1 (YBX1) is involved in the multi-tumor occurrence and development. However, the regulation of YBX1 in lung tumorigenesis and the underlying mechanisms, especially its relationship with CDC25a, was remains unclear. In this study, we analyzed the expression and clinical significance of YBX1 and CDC25a in lung adenocarcinoma and identified their roles in the regulation of lung cancer growth. The retrospective analysis of 116 patients with lung adenocarcinoma indicated that YBX1 was positively correlated with CDC25a expression. The Cox-regression analysis showed only high-ranking TNM stage and low CDC25a expression were an independent risk factor of prognosis in enrolled patients. High expression of YBX1 or CDC25a protein was also observed in lung adenocarcinoma cells compared with HLF cells. ChIP assay demonstrated the binding of endogenous YBX1 to the CDC25a promoter region. Overexpression of exogenous YBX1 up-regulated the expression of the CDC25a promoter-driven luciferase. By contrast, inhibition of YBX1 by siRNA markedly decreased the capability of YBX1 binding to CDC25a promoter in A549 and H322 cells. Inhibition of YBX1 expression also blocked cell cycle progression, suppressed cell proliferation and induced apoptosis via the CDC25a pathway in vitro. Moreover, inhibition of YBX1 by siRNA suppressed tumorigenesis in a xenograft mouse model and down-regulated the expression of YBX1, CDC25a, Ki67 and cleaved caspase 3 in the tumor tissues of mice. Collectively, these results demonstrate inhibition of YBX1 suppressed lung cancer growth partly via the CDC25a pathway and high expression of YBX1/CDC25a predicts poor prognosis in human lung adenocarcinoma. PMID:27384875

  19. Regulation of the Flavonoid Biosynthesis Pathway Genes in Purple and Black Grains of Hordeum vulgare

    PubMed Central

    Mock, Hans-Peter; Kukoeva, Tatjana V.; Börner, Andreas; Khlestkina, Elena K.

    2016-01-01

    Barley grain at maturity can have yellow, purple, blue, and black pigmentations which are suggested to play a protective role under stress conditions. The first three types of the colors are caused by phenolic compounds flavonoids; the last one is caused by phytomelanins, oxidized and polymerized phenolic compounds. Although the genetic basis of the flavonoid biosynthesis pathway in barley has been thoroughly studied, there is no data yet on its regulation in purple and black barley grains. In the current study, genetic model of Hordeum vulgare ‘Bowman’ near-isogenic lines (NILs) was used to investigate the regulation of the flavonoid biosynthesis in white, purple, and black barley grains. Microsatellite genotyping revealed donor segments in the purple- and black-grained lines on chromosomes 2H (in region of the Ant2 gene determining purple color of grains) and 1H (in region of the Blp gene determining black lemma and pericarp), respectively. The isolated dominant Ant2 allele of the purple-grained line has high level of sequence similarity with the recessive Bowman’s ant2 in coding region, whereas an insertion of 179 bp was detected in promoter region of ant2. This structural divergence between Ant2 and ant2 alleles may underlie their different expression in grain pericarp: Bowman’s Ant2 is not transcribed, whereas it was up-regulated in the purple-grained line with coordinately co-expressed flavonoid biosynthesis structural genes (Chs, Chi, F3h, F3’h, Dfr, Ans). This led to total anthocyain content increase in purple-grained line identified by ultra-performance liquid chromatography (HPLC). Collectively, these results proved the regulatory function of the Ant2 gene in anthocyanin biosynthesis in barley grain pericarp. In the black-grained line, the specific transcriptional regulation of the flavonoid biosynthesis pathway genes was not detected, suggesting that flavonoid pigments are not involved in development of black lemma and pericarp trait. PMID

  20. AcSDKP Regulates Cell Proliferation through the PI3KCA/Akt Signaling Pathway

    PubMed Central

    Hu, Ping; Li, Bin; Zhang, Wenhua; Li, Yijian; Li, Guang; Jiang, Xinnong; Wdzieczak-Bakala, Joanna; Liu, Jianmiao

    2013-01-01

    The natural tetrapeptide acetyl-N-Ser-Asp-Lys-Pro (AcSDKP) is generated from the N-terminus of thymosin-β4 through enzymatic cleavage by prolyl oligopeptidase (POP). AcSDKP regulation of proliferation of different cells is implicated in hematopoiesis and angiogenesis. This tetrapeptide present in almost all cells was recently detected at elevated concentrations in neoplastic diseases. However, previously reported in vitro and in vivo studies indicate that AcSDKP does not contribute to the pathogenesis of cancers. Here we show that exogenous AcSDKP exerts no effect on the proliferation of actively dividing malignant cells. Using S17092, a specific POP inhibitor (POPi), to suppress the biosynthesis of AcSDKP in U87-MG glioblastoma cells characterized by high intracellular levels of this peptide, we found that all tested doses of POPi resulted in an equally effective depletion of AcSDKP, which was not correlated with the dose-dependent decreases in the proliferation rate of treated cells. Interestingly, addition of exogenous AcSDKP markedly reversed the reduction in the proliferation of U87-MG cells treated with the highest dose of POPi, and this effect was associated with activation of the phosphatidylinositol-3 kinase (PI3K)/Akt pathway. However, extracellular-regulated protein kinase (ERK) activation was unaltered by S17092 and AcSDKP co-treatment. Knockdown of individual PI3K catalytic subunits revealed that p110α and p110β contributed differently to AcSDKP regulation of U87-MG cell proliferation. Disruption of p110α expression by small interfering RNA (siRNA) abrogated AcSDKP-stimulated Akt phosphorylation, whereas knockdown of p110β expression exhibited no such effect. Our findings indicate for the first time that the PI3KCA/Akt pathway mediates AcSDKP regulation of cell proliferation and suggest a role for this ubiquitous intracellular peptide in cell survival. PMID:24244481

  1. Chromatin landscaping in algae reveals novel regulation pathway for biofuels production

    SciTech Connect

    Ngan, Chew Yee; Wong, Chee-Hong; Choi, Cindy; Pratap, Abhishek; Han, James; Wei, Chia-Lin

    2013-02-19

    The diminishing reserve of fossil fuels calls for the development of biofuels. Biofuels are produced from renewable resources, including photosynthetic organisms, generating clean energy. Microalgae is one of the potential feedstock for biofuels production. It grows easily even in waste water, and poses no competition to agricultural crops for arable land. However, little is known about the algae lipid biosynthetic regulatory mechanisms. Most studies relied on the homology to other plant model organisms, in particular Arabidopsis or through low coverage expression analysis to identify key enzymes. This limits the discovery of new components in the biosynthetic pathways, particularly the genetic regulators and effort to maximize the production efficiency of algal biofuels. Here we report an unprecedented and de novo approach to dissect the algal lipid pathways through disclosing the temporal regulations of chromatin states during lipid biosynthesis. We have generated genome wide chromatin maps in chlamydomonas genome using ChIP-seq targeting 7 histone modifications and RNA polymerase II in a time-series manner throughout conditions activating lipid biosynthesis. To our surprise, the combinatory profiles of histone codes uncovered new regulatory mechanism in gene expression in algae. Coupled with matched RNA-seq data, chromatin changes revealed potential novel regulators and candidate genes involved in the activation of lipid accumulations. Genetic perturbation on these candidate regulators further demonstrated the potential to manipulate the regulatory cascade for lipid synthesis efficiency. Exploring epigenetic landscape in microalgae shown here provides powerful tools needed in improving biofuel production and new technology platform for renewable energy generation, global carbon management, and environmental survey.

  2. Comparative genomics reveals candidate carotenoid pathway regulators of ripening watermelon fruit

    PubMed Central

    2013-01-01

    Background Many fruits, including watermelon, are proficient in carotenoid accumulation during ripening. While most genes encoding steps in the carotenoid biosynthetic pathway have been cloned, few transcriptional regulators of these genes have been defined to date. Here we describe the identification of a set of putative carotenoid-related transcription factors resulting from fresh watermelon carotenoid and transcriptome analysis during fruit development and ripening. Our goal is to both clarify the expression profiles of carotenoid pathway genes and to identify candidate regulators and molecular targets for crop improvement. Results Total carotenoids progressively increased during fruit ripening up to ~55 μg g-1 fw in red-ripe fruits. Trans-lycopene was the carotenoid that contributed most to this increase. Many of the genes related to carotenoid metabolism displayed changing expression levels during fruit ripening generating a metabolic flux toward carotenoid synthesis. Constitutive low expression of lycopene cyclase genes resulted in lycopene accumulation. RNA-seq expression profiling of watermelon fruit development yielded a set of transcription factors whose expression was correlated with ripening and carotenoid accumulation. Nineteen putative transcription factor genes from watermelon and homologous to tomato carotenoid-associated genes were identified. Among these, six were differentially expressed in the flesh of both species during fruit development and ripening. Conclusions Taken together the data suggest that, while the regulation of a common set of metabolic genes likely influences carotenoid synthesis and accumulation in watermelon and tomato fruits during development and ripening, specific and limiting regulators may differ between climacteric and non-climacteric fruits, possibly related to their differential susceptibility to and use of ethylene during ripening. PMID:24219562

  3. The Phosphatidylinositol 3-Kinase/Akt Pathway Negatively Regulates Nod2-Mediated NF-kB Pathway

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Nucleotide-binding oligomerization domain containing proteins (Nods) are intracellular pattern recognition receptors (PRRs) that recognize conserved moieties of bacterial peptidoglycan and activate downstream signaling pathways including NF-kB pathway. Here, we show that Nod2 agonist MDP induces Ak...

  4. Flexible metabolic pathway construction using modular and divisible selection gene regulators.

    PubMed

    Rugbjerg, Peter; Myling-Petersen, Nils; Sommer, Morten O A

    2015-09-01

    Genetic selections are important to biological engineering. Although selectable traits are limited, currently each trait only permits simultaneous introduction of a single DNA fragment. Complex pathway and strain construction however depends on rapid, combinatorial introduction of many genes that encode putative pathway candidates and homologs. To triple the utility of existing selection genes, we have developed divisible selection in Saccharomyces cerevisiae. Here, independent DNA fragments can be introduced and selected for simultaneously using a set of split hybrid transcription factors composed of parts from Escherichia coli LexA and Herpes simplex VP16 to regulate one single selectable phenotype of choice. Only when co-expressed, these split hybrid transcription factors promote transcription of a selection gene, causing tight selection of transformants containing all desired DNA fragments. Upon transformation, 94% of the selected colonies resulted strictly from transforming all three modules based on ARS/CEN plasmids. Similarly when used for chromosome integration, 95% of the transformants contained all three modules. The divisible selection system acts dominantly and thus expands selection gene utility from one to three without any genomic pre-modifications of the strain. We demonstrate the approach by introducing the fungal rubrofusarin polyketide pathway at a gene load of 11 kb distributed on three different plasmids, using a single selection trait and one yeast transformation step. By tripling the utility of existing selection genes, the employment of divisible selection improves flexibility and freedom in the strain engineering process.

  5. Dentate Gyrus Development Requires ERK Activity to Maintain Progenitor Population and MAPK Pathway Feedback Regulation

    PubMed Central

    Vithayathil, Joseph; Pucilowska, Joanna; Goodnough, L. Henry; Atit, Radhika P.

    2015-01-01

    The ERK/MAPK pathway is an important developmental signaling pathway. Mutations in upstream elements of this pathway result in neuro-cardio-facial cutaneous (NCFC) syndromes, which are typified by impaired neurocognitive abilities that are reliant upon hippocampal function. The role of ERK signaling during hippocampal development has not been examined and may provide critical insight into the cause of hippocampal dysfunction in NCFC syndromes. In this study, we have generated ERK1 and conditional ERK2 compound knock-out mice to determine the role of ERK signaling during development of the hippocampal dentate gyrus. We found that loss of both ERK1 and ERK2 resulted in 60% fewer granule cells and near complete absence of neural progenitor pools in the postnatal dentate gyrus. Loss of ERK1/2 impaired maintenance of neural progenitors as they migrate from the dentate ventricular zone to the dentate gyrus proper, resulting in premature depletion of neural progenitor cells beginning at E16.5, which prevented generation of granule cells later in development. Finally, loss of ERK2 alone does not impair development of the dentate gyrus as animals expressing only ERK1 developed a normal hippocampus. These findings establish that ERK signaling regulates maintenance of progenitor cells required for development of the dentate gyrus. PMID:25926459

  6. BDNF Selectively Regulates GABAA Receptor Transcription by Activation of the JAK/STAT Pathway

    PubMed Central

    Lund, Ingrid V.; Hu, Yinghui; Raol, YogendraSinh H.; Benham, Rebecca S.; Faris, Ramona; Russek, Shelley J.; Brooks-Kayal, Amy R.

    2009-01-01

    The γ-aminobutyric acid (GABA) type A receptor (GABAAR) is the major inhibitory neurotransmitter receptor in the brain. Its multiple subunits show regional, developmental, and disease-related plasticity of expression; however, the regulatory networks controlling GABAAR subunit expression remain poorly understood. We report that the seizure-induced decrease in GABAAR α1 subunit expression associated with epilepsy is mediated by the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway regulated by brain-derived neurotrophic factor (BDNF). BDNF- and seizure-dependent phosphorylation of STAT3 cause the adenosine 3′,5′-monophosphate (cAMP) response element–binding protein (CREB) family member ICER (inducible cAMP early repressor) to bind with phosphorylated CREB at the Gabra1:CRE site. JAK/STAT pathway inhibition prevents the seizure-induced decrease in GABAAR α1 abundance in vivo and, given that BDNF is known to increase the abundance of GABAAR α4 in a JAK/STAT-independent manner, indicates that BDNF acts through at least two distinct pathways to influence GABAAR-dependent synaptic inhibition. PMID:18922788

  7. ERK1/2/MAPK pathway-dependent regulation of the telomeric factor TRF2

    PubMed Central

    Picco, Vincent; Coste, Isabelle; Giraud-Panis, Marie-Josèphe; Renno, Toufic; Gilson, Eric; Pagès, Gilles

    2016-01-01

    Telomere stability is a hallmark of immortalized cells, including cancer cells. While the telomere length is maintained in most cases by the telomerase, the activity of a protein complex called Shelterin is required to protect telomeres against unsuitable activation of the DNA damage response pathway. Within this complex, telomeric repeat binding factor 2 (TRF2) plays an essential role by blocking the ataxia telangiectasia-mutated protein (ATM) signaling pathway at telomeres and preventing chromosome end fusion. We showed that TRF2 was phosphorylated in vitro and in vivo on serine 323 by extracellular signal-regulated kinase (ERK1/2) in both normal and cancer cells. Moreover, TRF2 and activated ERK1/2 unexpectedly interacted in the cytoplasm of tumor cells and human tumor tissues. The expression of non-phosphorylatable forms of TRF2 in melanoma cells induced the DNA damage response, leading to growth arrest and tumor reversion. These findings revealed that the telomere stability is under direct control of one of the major pro-oncogenic signaling pathways (RAS/RAF/MEK/ERK) via TRF2 phosphorylation. PMID:27366950

  8. The Potential of Vitamin D-Regulated Intracellular Signaling Pathways as Targets for Myeloid Leukemia Therapy

    PubMed Central

    Gocek, Elzbieta; Studzinski, George P.

    2015-01-01

    The current standard regimens for the treatment of acute myeloid leukemia (AML) are curative in less than half of patients; therefore, there is a great need for innovative new approaches to this problem. One approach is to target new treatments to the pathways that are instrumental to cell growth and survival with drugs that are less harmful to normal cells than to neoplastic cells. In this review, we focus on the MAPK family of signaling pathways and those that are known to, or potentially can, interact with MAPKs, such as PI3K/AKT/FOXO and JAK/STAT. We exemplify the recent studies in this field with specific relevance to vitamin D and its derivatives, since they have featured prominently in recent scientific literature as having anti-cancer properties. Since microRNAs also are known to be regulated by activated vitamin D, this is also briefly discussed here, as are the implications of the emerging acquisition of transcriptosome data and potentiation of the biological effects of vitamin D by other compounds. While there are ongoing clinical trials of various compounds that affect signaling pathways, more studies are needed to establish the clinical utility of vitamin D in the treatment of cancer. PMID:26239344

  9. Hydroxyeicosatetraenoic acids released through the cytochrome P-450 pathway regulate 3T6 fibroblast growth.

    PubMed

    Nieves, Diana; Moreno, Juan José

    2006-12-01

    Eicosanoids participate in the regulation of cellular proliferation. Thus, we observed that prostaglandin E(2) interaction with membrane receptors is involved in the control of 3T6 fibroblast growth induced by serum. However, our results suggested that another arachidonic acid pathway might be implicated in these events. Our results show that 3T6 fibroblasts synthesized hydroxyeicosatetraenoic acids (HETEs) such as 12-HETE through the cytochrome P-450 (CYP450) pathway. However, 3T6 fibroblasts did not produce leukotriene B(4) (LTB(4)), and lipoxygenase inhibitors and LT antagonists failed to inhibit 3T6 fibroblast growth induced by FBS. In contrast, we observed that CYP450 inhibitors such as SKF-525A, 17-octadecynoic acid, 1-aminobenzotriazole, and 6-(2-propargyloxyphenyl)hexanoic acid reduced 12(S)-HETE levels, 3T6 fibroblast growth, and DNA synthesis induced by FBS. The impairment of DNA synthesis and 3T6 fibroblast growth induced by SKF-525A were reversed by exogenous addition of HETEs. Moreover, we report that 5-HETE, 12(S)-HETE, and 15(S)-HETE are mitogenic on 3T6 fibroblast in the absence of another growth factor, and this effect was dependent on the activation of the phosphatidylinositol-3-kinase pathway. In conclusion, our results show that HETEs, probably produced by CYP450, are involved in the control of 3T6 fibroblast growth.

  10. PORCN moonlights in a Wnt-independent pathway that regulates cancer cell proliferation.

    PubMed

    Covey, Tracy M; Kaur, Simran; Tan Ong, Tina; Proffitt, Kyle D; Wu, Yonghui; Tan, Patrick; Virshup, David M

    2012-01-01

    Porcupine (PORCN) is a membrane-bound O-acyl transferase that is required for the palmitoylation of Wnt proteins, and that is essential in diverse Wnt pathways for Wnt-Wntless (WLS) binding, Wnt secretion, and Wnt signaling activity. We tested if PORCN was required for the proliferation of transformed cells. Knockdown of PORCN by multiple independent siRNAs results in a cell growth defect in a subset of epithelial cancer cell lines. The growth defect is transformation-dependent in human mammary epithelial (HMEC) cells. Additionally, inducible PORCN knockdown by two independent shRNAs markedly reduces the growth of established MDA-MB-231 cancers in orthotopic xenografts in immunodeficient mice. Unexpectedly, the proliferation defect resulting from loss of PORCN occurs in a Wnt-independent manner, as it is rescued by re-expression of catalytically inactive PORCN, and is not seen after RNAi-mediated knockdown of the Wnt carrier protein WLS, nor after treatment with the PORCN inhibitor IWP. Consistent with a role in a Wnt-independent pathway, knockdown of PORCN regulates a distinct set of genes that are not altered by other inhibitors of Wnt signaling. PORCN protein thus appears to moonlight in a novel signaling pathway that is rate-limiting for cancer cell growth and tumorigenesis independent of its enzymatic function in Wnt biosynthesis and secretion.

  11. An endocytosis pathway initiated through neuropilin-1 and regulated by nutrient availability

    PubMed Central

    Pang, Hong-Bo; Braun, Gary B.; Friman, Tomas; Aza-Blanc, Pedro; Ruidiaz, Manuel E.; Sugahara, Kazuki N.; Teesalu, Tambet; Ruoslahti, Erkki

    2014-01-01

    Neuropilins (NRPs) are trans-membrane receptors involved in axon guidance and vascular development. Many growth factors and other signaling molecules bind to NRPs through a C-terminal, basic sequence motif (C-end Rule or CendR motif). Peptides with this motif (CendR peptides) are taken up into cells by endocytosis. Tumor-homing CendR peptides penetrate through tumor tissue and have shown utility in enhancing drug delivery into tumors. Here we show, using RNAi screening and subsequent validation studies, that NRP1-mediated endocytosis of CendR peptides is distinct from known endocytic pathways. Ultrastructurally, CendR endocytosis resembles macropinocytosis, but is mechanistically different. We also show that nutrient-sensing networks such as mTOR signaling regulate CendR endocytosis and subsequent intercellular transport of CendR cargo, both of which are stimulated by nutrient depletion. As CendR is a bulk transport pathway, our results suggest a role for it in nutrient transport; CendR-enhanced drug delivery then makes use of this natural pathway. PMID:25277522

  12. Kaempferol Attenuates Cardiac Hypertrophy via Regulation of ASK1/MAPK Signaling Pathway and Oxidative Stress.

    PubMed

    Feng, Hong; Cao, Jianlei; Zhang, Guangyu; Wang, Yanggan

    2017-02-20

    Kaempferol has been demonstrated to provide benefits for the treatment of atherosclerosis, coronary heart disease, hyperlipidemia, and diabetes through its antioxidant and anti-inflammatory properties. However, its role in cardiac hypertrophy remains to be elucidated. The aim of our study was to investigate the effects of kaempferol on cardiac hypertrophy and the underlying mechanism. Mice subjected to aorta banding were treated with or without kaempferol (100 mg/kg/d, p. o.) for 6 weeks. Echocardiography was performed to evaluate cardiac function. Mice hearts were collected for pathological observation and molecular mechanism investigation. H9c2 cardiomyocytes were stimulated with or without phenylephrine for in vitro study. Kaempferol significantly attenuated cardiac hypertrophy induced by aorta banding as evidenced by decreased cardiomyocyte areas and interstitial fibrosis, accompanied with improved cardiac functions and decreased apoptosis. The ASK1/MAPK signaling pathways (JNK1/2 and p38) were markedly activated in the aorta banding mouse heart but inhibited by kaempferol treatment. In in vitro experiments, kaempferol also inhibited the activity of ASK1/JNK1/2/p38 signaling pathway and the enlargement of H9c2 cardiomyocytes. Furthermore, our study revealed that kaempferol could protect the mouse heart and H9c2 cells from pathological oxidative stress. Our investigation indicated that treatment with kaempferol protects against cardiac hypertrophy, and its cardioprotection may be partially explained by the inhibition of the ASK1/MAPK signaling pathway and the regulation of oxidative stress.

  13. New insights into the role of the ubiquitin-proteasome pathway in the regulation of apoptosis.

    PubMed

    Liu, Cui-Hua; Goldberg, Alfred L; Qiu, Xiao-Bo

    2007-01-01

    The ubiquitin-proteasome pathway (UPP) is the major system responsible for degradation of intracellular proteins in eukaryotes. By controlling the levels of key proteins, it regulates almost all of the cellular activities, including cell cycle progression, DNA replication and repair, transcription, protein quality control, immune response, and apoptosis. UPP is composed of the ubiquitination system that marks proteins for degradation and the proteasome which degrades the ubiquitinated proteins. The 26S proteasome is a 2400 kDa complex consisting of more than 40 subunits. Following ubiquitination catalyzed by the ubiquitin activating enzyme (El), a ubiquitin-carrier protein (E2), and one of the cell's many ubiquitin-protein ligases (E3s), the protein substrates are targeted to the proteasome for degradation into small peptides. E3s regulate the degradation of protein substrates indirectly by determining both the specificity and timing of substrate ubiquitination, whereas the deubiquitinating enzymes can inhibit this process by releasing ubiquitin from substrates. In this review, we attempt to highlight the recent progress in research on UPP and its role in the regulation of apoptosis by focusing on several of its important components, including the ubiqutin ligase Nrdp 1, which regulates ErbB/EGFR family of receptor tyrosine kinases, the ubiquitin-carrier protein BRUCE/Apollon (an Inhibitor of Apoptosis Protein), and the novel proteasome subunit hRpnl3 (a binding site for the deubiquitinating enzyme, UCH37).

  14. A Mesh-Duox pathway regulates homeostasis in the insect gut.

    PubMed

    Xiao, Xiaoping; Yang, Lijuan; Pang, Xiaojing; Zhang, Rudian; Zhu, Yibin; Wang, Penghua; Gao, Guanjun; Cheng, Gong

    2017-03-01

    The metazoan gut harbours complex communities of commensal and symbiotic bacterial microorganisms. The quantity and quality of these microorganisms fluctuate dynamically in response to physiological changes. The mechanisms that hosts have developed to respond to and manage such dynamic changes and maintain homeostasis remain largely unknown. Here, we identify a dual oxidase (Duox)-regulating pathway that contributes to maintaining homeostasis in the gut of both Aedes aegypti and Drosophila melanogaster. We show that a gut-membrane-associated protein, named Mesh, plays an important role in controlling the proliferation of gut bacteria by regulating Duox expression through an Arrestin-mediated MAPK JNK/ERK phosphorylation cascade. Expression of both Mesh and Duox is correlated with the gut bacterial microbiome, which, in mosquitoes, increases dramatically soon after a blood meal. Ablation of Mesh abolishes Duox induction, leading to an increase of the gut microbiome load. Our study reveals that the Mesh-mediated signalling pathway is a central homeostatic mechanism of the insect gut.

  15. Target of Rapamycin Complex 2 Regulates Actin Polarization and Endocytosis via Multiple Pathways*

    PubMed Central

    Rispal, Delphine; Eltschinger, Sandra; Stahl, Michael; Vaga, Stefania; Bodenmiller, Bernd; Abraham, Yann; Filipuzzi, Ireos; Movva, N. Rao; Aebersold, Ruedi; Helliwell, Stephen B.; Loewith, Robbie

    2015-01-01

    Target of rapamycin is a Ser/Thr kinase that operates in two conserved multiprotein complexes, TORC1 and TORC2. Unlike TORC1, TORC2 is insensitive to rapamycin, and its functional characterization is less advanced. Previous genetic studies demonstrated that TORC2 depletion leads to loss of actin polarization and loss of endocytosis. To determine how TORC2 regulates these readouts, we engineered a yeast strain in which TORC2 can be specifically and acutely inhibited by the imidazoquinoline NVP-BHS345. Kinetic analyses following inhibition of TORC2, supported with quantitative phosphoproteomics, revealed that TORC2 regulates these readouts via distinct pathways as follows: rapidly through direct protein phosphorylation cascades and slowly through indirect changes in the tensile properties of the plasma membrane. The rapid signaling events are mediated in large part through the phospholipid flippase kinases Fpk1 and Fpk2, whereas the slow signaling pathway involves increased plasma membrane tension resulting from a gradual depletion of sphingolipids. Additional hits in our phosphoproteomic screens highlight the intricate control TORC2 exerts over diverse aspects of eukaryote cell physiology. PMID:25882841

  16. Identification and Characterization of Multiple Intermediate Alleles of the Key Genes Regulating Brassinosteroid Biosynthesis Pathways

    PubMed Central

    Du, Junbo; Zhao, Baolin; Sun, Xin; Sun, Mengyuan; Zhang, Dongzhi; Zhang, Shasha; Yang, Wenyu

    2017-01-01

    Most of the early identified brassinosteroid signaling and biosynthetic mutants are null mutants, exhibiting extremely dwarfed phenotypes and male sterility. These null mutants are usually unable to be directly transformed via a routinely used Agrobacterium-mediated gene transformation system and therefore are less useful for genetic characterization of the brassinosteroid (BR)-related pathways. Identification of intermediate signaling mutants such as bri1–5 and bri1–9 has contributed drastically to the elucidation of BR signaling pathway using both genetic and biochemical approaches. However, intermediate mutants of key genes regulating BR biosynthesis have seldom been reported. Here we report identification of several intermediate BR biosynthesis mutants mainly resulted from leaky transcriptions due to the insertions of T-DNAs in the introns. These mutants are semi-dwarfed and fertile and capable to be transformed. These intermediate mutants could be useful tools for future discovery and analyses of novel components regulating BR biosynthesis and catabolism via genetic modifier screen. PMID:28138331

  17. Molecular Mechanism for the Regulation of Microcystin Toxicity to Protein Phosphatase 1 by Glutathione Conjugation Pathway

    PubMed Central

    Wang, Xiaoning; Du, Yonggang; Zhang, Shuhan; Zhang, Ying; Teng, Yue

    2017-01-01

    Glutathione (GSH) conjugation was an important pathway to regulate the toxicity of microcystins (MCs) targeted to protein phosphatases. To explore the specific molecular mechanism for GSH detoxification, two typical MC-GSHs (derived from MCLR and MCRR) were synthesized, prepared, and purified according to previous research. Then, the reduced inhibition effect for MC-GSHs on protein phosphatase 1 was verified by comparing with their original toxins. To further clarify the molecular mechanism for MC-GSHs detoxification, we evaluated the interactions between MCs/MC-GSHs and PP1 with the assistance of MOE molecule simulation. When GSH was introduced to MCs, the covalent binding (Mdha7 to Cys273), the hydrophobic interaction (Adda5 with PP1), the hydrogen bonds (especially for Lys2-Arg96 and Glu6-Tyr272), the covalent combination (between Mdha7 and Cys273), and the ion bonds (between Mn2+ and Asn124/His248/Asp64/His66) of MCLR/MCRR-PP1 complexes weakened to a certain extent, while the ion bonds between Mn2+ and His173/Asp92 residues increased. It was not difficult to find that the toxicity of MCs was closely related to the above sites/interactions and the above key information for MCs-PP1; MC-GSHs-PP1 complexes were important for clarifying the detoxification mechanism of MC-GSHs pathway. This study offers a comprehensive cognition on MCs toxicity regulation and provides valid theoretical support to control their potential risk. PMID:28337461

  18. Pak1 regulates the orientation of apical polarization and lumen formation by distinct pathways.

    PubMed

    deLeon, Orlando; Puglise, Jason M; Liu, Fengming; Smits, Jos; ter Beest, Martin B; Zegers, Mirjam M

    2012-01-01

    The development of the basic architecture of branching tubules enclosing a central lumen that characterizes most epithelial organs crucially depends on the apico-basolateral polarization of epithelial cells. Signals from the extracellular matrix control the orientation of the apical surface, so that it faces the lumen interior, opposite to cell-matrix adhesion sites. This orientation of the apical surface is thought to be intrinsically linked to the formation of single lumens. We previously demonstrated in three-dimensional cyst cultures of Madin-Darby canine kidney (MDCK) cells that signaling by β1 integrins regulates the orientation of the apical surface, via a mechanism that depends on the activity of the small GTPase Rac1. Here, we investigated whether the Rac1 effector Pak1 is a downstream effector in this pathway. Expression of constitutive active Pak1 phenocopies the effect of β1 integrin inhibition in that it misorients the apical surface and induces a multilumen phenotype. The misorientation of apical surfaces depends on the interaction of active Pak1 with PIX proteins and is linked to defects in basement membrane assembly. In contrast, the multilumen phenotype was independent of PIX and the basement membrane. Therefore, Pak1 likely regulates apical polarization and lumen formation by two distinct pathways.

  19. Pak1 Regulates the Orientation of Apical Polarization and Lumen Formation by Distinct Pathways

    PubMed Central

    Smits, Jos; ter Beest, Martin B.; Zegers, Mirjam M.

    2012-01-01

    The development of the basic architecture of branching tubules enclosing a central lumen that characterizes most epithelial organs crucially depends on the apico-basolateral polarization of epithelial cells. Signals from the extracellular matrix control the orientation of the apical surface, so that it faces the lumen interior, opposite to cell-matrix adhesion sites. This orientation of the apical surface is thought to be intrinsically linked to the formation of single lumens. We previously demonstrated in three-dimensional cyst cultures of Madin-Darby canine kidney (MDCK) cells that signaling by β1 integrins regulates the orientation of the apical surface, via a mechanism that depends on the activity of the small GTPase Rac1. Here, we investigated whether the Rac1 effector Pak1 is a downstream effector in this pathway. Expression of constitutive active Pak1 phenocopies the effect of β1 integrin inhibition in that it misorients the apical surface and induces a multilumen phenotype. The misorientation of apical surfaces depends on the interaction of active Pak1 with PIX proteins and is linked to defects in basement membrane assembly. In contrast, the multilumen phenotype was independent of PIX and the basement membrane. Therefore, Pak1 likely regulates apical polarization and lumen formation by two distinct pathways. PMID:22815903

  20. Glycitin regulates osteoblasts through TGF-β or AKT signaling pathways in bone marrow stem cells

    PubMed Central

    Zhang, Liyan; Chen, Jiying; Chai, Wei; Ni, Min; Sun, Xin; Tian, Dan

    2016-01-01

    The aim of the present study was to examine the effect of glycitin on the regulation of osteoblasts from bone marrow stem cells (BMSCs) through transforming growth factor (TGF)-β or protein kinase B (AKT) signaling pathways. BMSCs were extracted from New Zealand white rabbits and used to analyze the effect of glycitin on BMSCs. BMSCs were cleared using xylene and observed via light microscopy. BMSCs were subsequently induced with glycitin (0.01, 0.5, 1, 5 and 10 µM) for 7 days, and stained with Oil Red O. The mechanism of action of glycitin on BMSCs was investigated, in which contact with collagen type I (Col I), alkaline phosphatase (ALP), TGF-β and AKT was studied. Firstly, BMSCs appeared homogeneously mazarine blue, and which showed that BMSCs were successful extracted. Administration of glycitin increased cell proliferation and promoted osteoblast formation from BMSCs. Furthermore, glycitin activated the gene expression of Col I and ALP in BMSCs. Notably, glycitin suppressed protein expression of TGF-β and AKT in BMSCs. These results indicated that glycitin may regulate osteoblasts through TGF-β or AKT signaling pathways in BMSCs. PMID:27882117

  1. The RhoA-ROCK pathway in the regulation of T and B cell responses

    PubMed Central

    Ricker, Edd; Chowdhury, Luvana; Yi, Woelsung; Pernis, Alessandra B.

    2016-01-01

    Effective immune responses require the precise regulation of dynamic interactions between hematopoietic and non-hematopoietic cells. The Rho subfamily of GTPases, which includes RhoA, is rapidly activated downstream of a diverse array of biochemical and biomechanical signals, and is emerging as an important mediator of this cross-talk. Key downstream effectors of RhoA are the Rho kinases, or ROCKs. The ROCKs are two serine-threonine kinases that can act as global coordinators of a tissue’s response to stress and injury because of their ability to regulate a wide range of biological processes. Although the RhoA-ROCK pathway has been extensively investigated in the non-hematopoietic compartment, its role in the immune system is just now becoming appreciated. In this commentary, we provide a brief overview of recent findings that highlight the contribution of this pathway to lymphocyte development and activation, and the impact that dysregulation in the activation of RhoA and/or the ROCKs may exert on a growing list of autoimmune and lymphoproliferative disorders. PMID:27785353

  2. Regulation of Bovine Papillomavirus Replicative Helicase E1 by the Ubiquitin-Proteasome Pathway

    PubMed Central

    Malcles, Marie-Helene; Cueille, Nathalie; Mechali, Francisca; Coux, Olivier; Bonne-Andrea, Catherine

    2002-01-01

    Papillomaviruses maintain their genomes in a relatively constant copy number as stable extrachromosomal plasmids in the nuclei of dividing host cells. The viral initiator of replication, E1, is not detected in papillomavirus-infected cells. Here, we present evidence that E1 encoded by bovine papillomavirus type 1 is an unstable protein that is degraded through the ubiquitin-proteasome pathway. In a cell-free system derived from Xenopus egg extracts, E1 degradation is regulated by both cyclin E/Cdk2 binding and E1 replication activity. Free E1 is readily ubiquitinated and degraded by the proteasome, while it becomes resistant to this degradation pathway when bound to cyclin E/Cdk2 complexes before the start of DNA synthesis. This stabilization is reversed in a process involving E1-dependent replication activity. In transiently transfected cells, E1 is also polyubiquitinated and accumulates when proteasome activity is inhibited. Thus, the establishment and maintenance of a stable number of papillomavirus genomes in latently infected cells are in part a function of regulated ubiquitin-mediated degradation of E1. PMID:12388695

  3. ANGUSTIFOLIA mediates one of the multiple SCRAMBLED signaling pathways regulating cell growth pattern in Arabidopsis thaliana.

    PubMed

    Kwak, Su-Hwan; Song, Sang-Kee; Lee, Myeong Min; Schiefelbein, John

    2015-09-25

    In Arabidopsis thaliana, an atypical leucine-rich repeat receptor-like kinase, SCRAMBLED (SCM), is required for multiple developmental processes including root epidermal cell fate determination, silique dehiscence, inflorescence growth, ovule morphogenesis, and tissue morphology. Previous work suggested that SCM regulates these multiple pathways using distinct mechanisms via interactions with specific downstream factors. ANGUSTIFOLIA (AN) is known to regulate cell and tissue morphogenesis by influencing cortical microtubule arrangement, and recently, the AN protein was reported to interact with the SCM protein. Therefore, we examined whether AN might be responsible for mediating some of the SCM-dependent phenotypes. We discovered that both scm and an mutant lines cause an abnormal spiral or twisting growth of roots, but only the scm mutant affected root epidermal patterning. The siliques of the an and scm mutants also exhibited spiral growth, as previously reported, but only the scm mutant altered silique dehiscence. Interestingly, we discovered that the spiral growth of roots and siliques of the scm mutant is rescued by a truncated SCM protein that lacks its kinase domain, and that a juxtamembrane domain of SCM was sufficient for AN binding in the yeast two-hybrid analysis. These results suggest that the AN protein is one of the critical downstream factors of SCM pathways specifically responsible for mediating its effects on cell/tissue morphogenesis through cortical microtubule arrangement.

  4. Construction of an miRNA-regulated drug-pathway network reveals drug repurposing candidates for myasthenia gravis

    PubMed Central

    Cao, Yuze; Lu, Xiaoyan; Wang, Jianjian; Zhang, Huixue; Liu, Zhaojun; Xu, Si; Wang, Tianfeng; Ning, Shangwei; Xiao, Bo; Wang, Lihua

    2017-01-01

    Myasthenia gravis (MG) is a rare debilitating autoimmune neuromuscular disorder. Many studies have focused on the mechanism and treatment strategies of MG. However, the exact pathogenesis of MG and effective treatment strategies remain unclear. Recent studies have indicated that microRNAs (miRNAs or miRs) can regulate the pathological pathways of MG, suggesting their potential role in novel treatments. In the present study, we created a comprehensive catalog of experimentally confirmed MG risk genes and miRNAs by manually mining published literature and public databases. Based on these genes and miRNAs, we identified 41 MG risk pathways and 105 approved drugs that can affect these pathways. Some important MG-related pathways, such as hsa04060 (cytokine-cytokine receptor interaction) and hsa05200 (pathway in cancer), were found to be regulated by MG risk miRNAs and drugs. Furthermore, we constructed an miRNA-regulated drug-pathway network and identified miRNAs and drugs that synergistically regulate key MG pathways and biological processes. We developed a drug repurposing strategy to identify 25 drug repurposing candidates for MG; several of these drugs, such as rituximab, adalimumab, sunitinib, and muromonab, have the potential to be novel MG treatment drugs. This study provides novel insight into the pathogenesis of MG and potential drug candidates for MG were identified. PMID:28075449

  5. Regulation of the Omega-3 Fatty Acid Biosynthetic Pathway in Atlantic Salmon Hepatocytes

    PubMed Central

    Ruyter, Bente; Berge, Gerd Marit; Sun, Yajing; Østbye, Tone-Kari Knutsdatter

    2016-01-01

    Limited availability of the n-3 fatty acids EPA and DHA have led to an interest in better understanding of the n-3 biosynthetic pathway and its regulation. The biosynthesis of alpha-linolenic acid to EPA and DHA involves several complex reaction steps including desaturation-, elongation- and peroxisomal beta-oxidation enzymes. The aims of the present experiments were to gain more knowledge on how this biosynthesis is regulated over time by different doses and fatty acid combinations. Hepatocytes isolated from salmon were incubated with various levels and combinations of oleic acid, EPA and DHA. Oleic acid led to a higher expression of the Δ6 fatty acid desaturase (fad) genes Δ6fad_a, Δ6fad_b, Δ6fad_c and the elongase genes elovl2 compared with cells cultured in medium enriched with DHA. Further, the study showed rhythmic variations in expression over time. Levels were reached where a further increase in specific fatty acids given to the cells not stimulated the conversion further. The gene expression of Δ6fad_a_and Δ6fad_b responded similar to fatty acid treatment, suggesting a co-regulation of these genes, whereas Δ5fad and Δ6fad_c showed a different regulation pattern. EPA and DHA induced different gene expression patterns, especially of Δ6fad_a. Addition of radiolabelled alpha-linolenic acid to the hepatocytes confirmed a higher degree of elongation and desaturation in cells treated with oleic acid compared to cells treated with DHA. This study suggests a complex regulation of the conversion process of n-3 fatty acids. Several factors, such as that the various gene copies are differently regulated, the gene expression show rhythmic variations and gene expression only affected to a certain level, determines when you get the maximum conversion of the beneficial n-3 fatty acids. PMID:27973547

  6. Mechanisms regulating neuronal excitability and seizure development following mTOR pathway hyperactivation

    PubMed Central

    LaSarge, Candi L.; Danzer, Steve C.

    2014-01-01

    The phosphatidylinositol-3-kinase/phosphatase and tensin homolog (PTEN)-mammalian target of rapamycin (mTOR) pathway regulates a variety of neuronal functions, including cell proliferation, survival, growth, and plasticity. Dysregulation of the pathway is implicated in the development of both genetic and acquired epilepsies. Indeed, several causal mutations have been identified in patients with epilepsy, the most prominent of these being mutations in PTEN and tuberous sclerosis complexes 1 and 2 (TSC1, TSC2). These genes act as negative regulators of mTOR signaling, and mutations lead to hyperactivation of the pathway. Animal models deleting PTEN, TSC1, and TSC2 consistently produce epilepsy phenotypes, demonstrating that increased mTOR signaling can provoke neuronal hyperexcitability. Given the broad range of changes induced by altered mTOR signaling, however, the mechanisms underlying seizure development in these animals remain uncertain. In transgenic mice, cell populations with hyperactive mTOR have many structural abnormalities that support recurrent circuit formation, including somatic and dendritic hypertrophy, aberrant basal dendrites, and enlargement of axon tracts. At the functional level, mTOR hyperactivation is commonly, but not always, associated with enhanced synaptic transmission and plasticity. Moreover, these populations of abnormal neurons can affect the larger network, inducing secondary changes that may explain paradoxical findings reported between cell and network functioning in different models or at different developmental time points. Here, we review the animal literature examining the link between mTOR hyperactivation and epileptogenesis, emphasizing the impact of enhanced mTOR signaling on neuronal form and function. PMID:24672426

  7. Coenzyme Q regulates the expression of essential genes of the pathogen- and xenobiotic-associated defense pathway in C. elegans

    PubMed Central

    Fischer, Alexandra; Niklowitz, Petra; Menke, Thomas; Döring, Frank

    2015-01-01

    Coenzyme Q (CoQ) is necessary for mitochondrial energy production and modulates the expression of genes that are important for inflammatory processes, growth and detoxification reactions. A cellular surveillance-activated detoxification and defenses (cSADDs) pathway has been recently identified in C. elegans. The down-regulation of the components of the cSADDs pathway initiates an aversion behavior of the nematode. Here we hypothesized that CoQ regulates genes of the cSADDs pathway. To verify this we generated CoQ-deficient worms (“CoQ-free”) and performed whole-genome expression profiling. We found about 30% (120 genes) of the cSADDs pathway genes were differentially regulated under CoQ-deficient condition. Remarkably, 83% of these genes were down-regulated. The majority of the CoQ-sensitive cSADDs pathway genes encode for proteins involved in larval development (enrichment score (ES) = 38.0, p = 5.0E−37), aminoacyl-tRNA biosynthesis, proteasome function (ES 8.2, p = 5.9E−31) and mitochondria function (ES 3.4, p = 1.7E−5). 67% (80 genes) of these genes are categorized as lethal. Thus it is shown for the first time that CoQ regulates a substantial number of essential genes that function in the evolutionary conserved cellular surveillance-activated detoxification and defenses pathway in C. elegans. PMID:26566301

  8. Maternal Undernourished Fetal Kidneys Exhibit Differential Regulation of Nephrogenic Genes Including Downregulation of the Notch Signaling Pathway

    PubMed Central

    Magee, Thomas R.; Tafti, Sanaz A.; Desai, Mina; Liu, Qinghai; Ross, Michael G.; Nast, Cynthia C

    2011-01-01

    Maternal undernutrition results in offspring nephron number reduction and hypertension that are hypothesized to begin as compensatory changes in fetal gene expression during gestation. To evaluate mechanisms of dysregulated nephrogenesis, pregnant Sprague Dawley rats were 50% food restricted from embryonic day (E) 10 to E20. At E20, fetal male kidneys were examined by microarray analysis. A total of 476 differentially expressed transcripts were detected including those regulating development and differentiation, mitosis and cell cycle, chromatin assembly, and steroid hormone regulation. Differentially regulated genes were detected in MAPK/ERK, Wnt, and Notch signaling pathways. Validation of the microarray results was performed for the Notch signaling pathway, an important pathway in nephron formation. Protein expression of Notch pathway factors by Western blotting showed significantly decreased Notch2 and downstream effector Hey1 protein expression, while Ctbp1 co-repressor was increased. These data together show that maternal undernutrition results in developmental disruption in fetal nephrogenesis gene expression signaling. PMID:21273641

  9. The canonical Notch pathway effector RBP-J regulates neuronal plasticity and expression of GABA transporters in hippocampal networks.

    PubMed

    Liu, Shuxi; Wang, Yue; Worley, Paul F; Mattson, Mark P; Gaiano, Nicholas

    2015-05-01

    Activation of the Notch pathway in neurons is essential for learning and memory in various species from invertebrates to mammals. However, it remains unclear how Notch signaling regulates neuronal plasticity, and whether the transcriptional regulator and canonical pathway effector RBP-J plays a role. Here, we report that conditional disruption of RBP-J in the postnatal hippocampus leads to defects in long-term potentiation, long-term depression, and in learning and memory. Using gene expression profiling and chromatin immunoprecipitation, we identified two GABA transporters, GAT2 and BGT1, as putative Notch/RBP-J pathway targets, which may function downstream of RBP-J to limit the accumulation of GABA in the Schaffer collateral pathway. Our results reveal an essential role for canonical Notch/RBP-J signaling in hippocampal synaptic plasticity and suggest that role, at least in part, is mediated by the regulation of GABAergic signaling.

  10. The mRNA Decay Pathway Regulates the Expression of the Flo11 Adhesin and Biofilm Formation in Saccharomyces cerevisiae

    PubMed Central

    Lo, Tricia L.; Qu, Yue; Uwamahoro, Nathalie; Quenault, Tara; Beilharz, Traude H.; Traven, Ana

    2012-01-01

    Regulation of the FLO11 adhesin is a model for gene expression control by extracellular signals and developmental switches. We establish that the major mRNA decay pathway regulates FLO11 expression. mRNA deadenylation of transcriptional repressors of FLO11 by the exonuclease Ccr4 keeps their levels low, thereby allowing FLO11 transcription. PMID:22595243

  11. Differential regulation of hepatic apoptotic pathways by dietary olive and sunflower oils in the aging rat.

    PubMed

    Bello, Rosario I; Gómez-Díaz, Consuelo; Burón, María I; Navas, Plácido; Villalba, José M

    2006-11-01

    In this work we have studied how dietary fat affects aging-related changes in a number of factors that regulate rat hepatic apoptosis. Animals were fed lifelong with two experimental diets containing either virgin olive oil or sunflower oil as dietary fat. Caspases of the intrinsic and extrinsic pathways of apoptosis, Bcl-2 and Bax polypeptide levels, and plasma membrane neutral sphingomyelinase activity were determined at 6, 12, and 24 months of age. Caspase-8/10 activity (a marker of the extrinsic pathway) was not affected by either aging or dietary fat, but activities of both caspase-9 (a marker of the intrinsic pathway) and caspase-3 (an executioner caspase) were significantly depressed in liver from animals fed on a sunflower oil-based diet. These decreases were not observed in animals fed with a diet based on virgin olive oil, which also resulted in significantly lower Bcl-2/Bax ratios. On the other hand, in comparison with sunflower, dietary olive oil decreased oxidative stress in liver from aged rats, resulting in lower levels of membrane hydroperoxides and higher coenzyme Q levels in plasma membrane. Plasma membrane Mg(2+)-dependent neutral sphingomyelinase was strongly activated in aged rats fed on the sunflower oil diet, but no aging-related increase was observed in animals fed on the olive oil diet. Our results support that dietary oil can alter significantly the susceptibility of hepatocytes to different apoptotic stimuli by altering both pro- and anti-apoptotic mediators, which reinforces the importance of the diet in aging studies. Because virgin olive oil may increase susceptibility of hepatocytes to apoptosis induced through the intrinsic pathway under conditions of decreased oxidative stress, our results may have important implications to understand the potential beneficial effects of that edible oil against liver carcinogenesis during aging.

  12. C-Fos Regulation by the MAPK and PKC Pathways in Intervertebral Disc Cells

    PubMed Central

    Yokoyama, Katsuya; Hiyama, Akihiko; Arai, Fumiyuki; Nukaga, Tadashi; Sakai, Daisuke; Mochida, Joji

    2013-01-01

    Background The gene encoding c-fos is an important factor in the pathogenesis of joint disease in patients with osteoarthritis. However, it is unknown whether the signal mechanism of c-fos acts in intervertebral disc (IVD) cells. We investigated whether c-fos is activated in relation to mitogen-activated protein kinases (MAPKs) and the protein kinase C (PKC) pathway in nucleus pulposus (NP) cells. Methodology/Results Reverse transcription-polymerase chain reaction and western blotting analyses were used to measure the expression of c-fos in rat IVD cells. Transfections were performed to determine the effects of c-fos on target gene activity. The effect of c-fos protein expression was examined in transfection experiments and in a 3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide cell viability assay. Phorbol 12-myristate 13-acetate (PMA), the most commonly used phorbol ester, binds to and activates protein kinase C (PKC), causing a wide range of effects in cells and tissues. PMA induced the expression of c-fos gene transcription and protein expression, and led to activation of the MAPK pathways in NP cells. The c-fos promoter was suppressed completely in the presence of the MAPK inhibitor PD98059, an inhibitor of the MEK/ERK kinase cascade, but not in the presence of SKF86002, SB202190, or SP600125. The effects of the PKC pathway on the transcriptional activity of the c-fos were evaluated. PKCγ and PKCδ suppressed the promoter activity of c-fos. Treatment with c-fos inhibited aggrecan and Col2 promoter activities and the expression of these genes in NP cells. Conclusions This study demonstrated, for the first time, that the MAPK and PKC pathways had opposing effects on the regulation of c-fos in NP cells. Thus, the expression of c-fos can be suppressed in the extracellular matrix of NP cells. PMID:24023832

  13. 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

  14. TRPM7 channels regulate glioma stem cell through STAT3 and Notch signaling pathways.

    PubMed

    Liu, Mingli; Inoue, Koichi; Leng, Tiandong; Guo, Shanchun; Xiong, Zhi-gang

    2014-12-01

    Glioblastoma multiforme (GBM) is the most common and aggressive primary brain tumor in adults with median survival time of 14.6 months. A small fraction of cancer stem cells (CSC) initiate and maintain tumors thus driving glioma tumorigenesis and being responsible for resistance to classical chemo- and radio-therapies. It is desirable to identify signaling pathways related to CSC to develop novel therapies to selectively target them. Transient receptor potential cation channel, subfamily M, member 7, also known as TRPM7 is a ubiquitous, Ca(2+) and Mg(2+) permeable ion channels that are special in being both an ion channel and a serine/threonine kinase. In studies of glioma cells silenced for TRPM7, we demonstrated that Notch (Notch1, JAG1, Hey2, and Survivin) and STAT3 pathways are down regulated in glioma cells grown in monolayer. Furthermore, phospho-STAT3, Notch target genes and CSC markers (ALDH1 and CD133) were significantly higher in spheroid glioma CSCs when compared with monolayer cultures. The results further show that tyrosine-phosphorylated STAT3 binds and activates the ALDH1 promoters in glioma cells. We found that TRMP7-induced upregulation of ALDH1 expression is associated with increases in ALDH1 activity and is detectable in stem-like cells when expanded as spheroid CSCs. Finally, TRPM7 promotes proliferation, migration and invasion of glioma cells. These demonstrate that TRPM7 activates JAK2/STAT3 and/or Notch signaling pathways and leads to increased cell proliferation and migration. These findings for the first time demonstrates that TRPM7 (1) activates a previously unrecognized STAT3→ALDH1 pathway, and (2) promotes the induction of ALDH1 activity in glioma cells.

  15. Computational inference of the structure and regulation of the lignin pathway in Panicum virgatum

    SciTech Connect

    Faraji, Mojdeh; Fonseca, Luis L.; Escamilla-Treviño, Luis; Dixon, Richard A.; Voit, Eberhard O.

    2015-09-17

    Switchgrass is a prime target for biofuel production from inedible plant parts and has been the subject of numerous investigations in recent years. Yet, one of the main obstacles to effective biofuel production remains to be the major problem of recalcitrance. Recalcitrance emerges in part from the 3-D structure of lignin as a polymer in the secondary cell wall. Lignin limits accessibility of the sugars in the cellulose and hemicellulose polymers to enzymes and ultimately decreases ethanol yield. Monolignols, the building blocks of lignin polymers, are synthesized in the cytosol and translocated to the plant cell wall, where they undergo polymerization. The biosynthetic pathway leading to monolignols in switchgrass is not completely known, and difficulties associated with in vivo measurements of these intermediates pose a challenge for a true understanding of the functioning of the pathway. In this study, a systems biological modeling approach is used to address this challenge and to elucidate the structure and regulation of the lignin pathway through a computational characterization of alternate candidate topologies. The analysis is based on experimental data characterizing stem and tiller tissue of four transgenic lines (knock-downs of genes coding for key enzymes in the pathway) as well as wild-type switchgrass plants. These data consist of the observed content and composition of monolignols. The possibility of a G-lignin specific metabolic channel associated with the production and degradation of coniferaldehyde is examined, and the results support previous findings from another plant species. The computational analysis suggests regulatory mechanisms of product inhibition and enzyme competition, which are well known in biochemistry, but so far had not been reported in switchgrass. By including these mechanisms, the pathway model is able to represent all observations. In conclusion, the results show that the presence of the coniferaldehyde channel is necessary

  16. Computational inference of the structure and regulation of the lignin pathway in Panicum virgatum

    DOE PAGES

    Faraji, Mojdeh; Fonseca, Luis L.; Escamilla-Treviño, Luis; ...

    2015-09-17

    Switchgrass is a prime target for biofuel production from inedible plant parts and has been the subject of numerous investigations in recent years. Yet, one of the main obstacles to effective biofuel production remains to be the major problem of recalcitrance. Recalcitrance emerges in part from the 3-D structure of lignin as a polymer in the secondary cell wall. Lignin limits accessibility of the sugars in the cellulose and hemicellulose polymers to enzymes and ultimately decreases ethanol yield. Monolignols, the building blocks of lignin polymers, are synthesized in the cytosol and translocated to the plant cell wall, where they undergomore » polymerization. The biosynthetic pathway leading to monolignols in switchgrass is not completely known, and difficulties associated with in vivo measurements of these intermediates pose a challenge for a true understanding of the functioning of the pathway. In this study, a systems biological modeling approach is used to address this challenge and to elucidate the structure and regulation of the lignin pathway through a computational characterization of alternate candidate topologies. The analysis is based on experimental data characterizing stem and tiller tissue of four transgenic lines (knock-downs of genes coding for key enzymes in the pathway) as well as wild-type switchgrass plants. These data consist of the observed content and composition of monolignols. The possibility of a G-lignin specific metabolic channel associated with the production and degradation of coniferaldehyde is examined, and the results support previous findings from another plant species. The computational analysis suggests regulatory mechanisms of product inhibition and enzyme competition, which are well known in biochemistry, but so far had not been reported in switchgrass. By including these mechanisms, the pathway model is able to represent all observations. In conclusion, the results show that the presence of the coniferaldehyde channel is

  17. MiR-223/PAX6 Axis Regulates Glioblastoma Stem Cell Proliferation and the Chemo Resistance to TMZ via Regulating PI3K/Akt Pathway.

    PubMed

    Huang, Bai-Sheng; Luo, Qi-Zhi; Han, Yang; Huang, Dong; Tang, Qing-Ping; Wu, Li-Xiang

    2017-03-23

    Chemotherapy is a standard strategy for glioma, while chemoresistance remains a major therapeutic challenge in current clinical practice. Our present study was aimed to determine whether inhibition of the miR-223/ paired box 6 (PAX6) pathway could increase the sensitivity of glioma to Temozolomide. An elevated level of miR-223 was observed in glioma tissues. Exogenous miR-223 promoted cell survival when exposed to Temozolomide (TMZ), while miR-223 inhibition could reverse this process. The RNA and protein levels of PAX6 were significantly decreased by exogenous miR-223, and the 3'-untranslated region of PAX6 was shown to be a target of miR-223. Besides, it has also been reported that PI3K/Akt signaling pathway is pivotal to regulate glioma growth and proliferation. In the present study, we revealed that miR-223/PAX6 axis regulated the growth, invasion and chemo resistance of glioblastoma stem cells to TMZ via regulating PI3K/Akt signaling pathway, which present a novel potential therapy for intervention of glioblastoma. Taken together, our findings shed new light on the miR-223/PAX6 pathway in glioma and this pathway might modulate the sensitivity of glioma to TMZ via regulating PI3K/Akt signaling pathway. This article is protected by copyright. All rights reserved.

  18. The ORCA2 transcription factor plays a key role in regulation of the terpenoid indole alkaloid pathway

    PubMed Central

    2013-01-01

    Background The terpenoid indole alkaloid (TIA) pathway leads to the production of pharmaceutically important drugs, such as the anticancer compounds vinblastine and vincristine. Unfortunately, these drugs are produced in trace amounts, causing them to be very costly. To increase production of these drugs, an improved understanding of the TIA regulatory pathway is needed. Towards this end, transgenic Catharanthus roseus hairy roots that overexpress the ORCA2 TIA transcriptional activator were generated and characterized. Results Transcriptional profiling experiments revealed that overexpression of ORCA2 results in altered expression of key genes from the indole and terpenoid pathways, which produce precursors for the TIA pathway, and from the TIA pathway itself. In addition, metabolite-profiling experiments revealed that overexpression of ORCA2 significantly affects the levels of several TIA metabolites. ORCA2 overexpression also causes significant increases in transcript levels of several TIA regulators, including TIA transcriptional repressors. Conclusions Results presented here indicate that ORCA2 plays a critical role in regulation of TIA metabolism. ORCA2 regulates expression of key genes from both feeder pathways, as well as the genes (STR and SGD) encoding the enzymes that catalyze the first two steps in TIA biosynthesis. ORCA2 may play an especially important role in regulation of the downstream branches of the TIA pathway, as it regulates four out of five genes characterized from this part of the pathway. Regulation of TIA transcriptional repressors by ORCA2 may provide a mechanism whereby increases in TIA metabolite levels in response to external stimuli are transient and limited in magnitude. PMID:24099172

  19. Notch1 Regulates Hippocampal Plasticity Through Interaction with the Reelin Pathway, Glutamatergic Transmission and CREB Signaling

    PubMed Central

    Brai, Emanuele; Marathe, Swananda; Astori, Simone; Fredj, Naila Ben; Perry, Elisabeth; Lamy, Christophe; Scotti, Alessandra; Alberi, Lavinia

    2015-01-01

    Notch signaling plays a crucial role in adult brain function such as synaptic plasticity, memory and olfaction. Several reports suggest an involvement of this pathway in neurodegenerative dementia. Yet, to date, the mechanism underlying Notch activity in mature neurons remains unresolved. In this work, we investigate how Notch regulates synaptic potentiation and contributes to the establishment of memory in mice. We observe that Notch1 is a postsynaptic receptor with functional interactions with the Reelin receptor, apolipoprotein E receptor 2 (ApoER2) and the ionotropic receptor, N-methyl-D-aspartate receptor (NMDAR). Targeted loss of Notch1 in the hippocampal CA fields affects Reelin signaling by influencing Dab1 expression and impairs the synaptic potentiation achieved through Reelin stimulation. Further analysis indicates that loss of Notch1 affects the expression and composition of the NMDAR but not AMPAR. Glutamatergic signaling is further compromised through downregulation of CamKII and its secondary and tertiary messengers resulting in reduced cAMP response element-binding (CREB) signaling. Our results identify Notch1 as an important regulator of mechanisms involved in synaptic plasticity and memory formation. These findings emphasize the possible involvement of this signaling receptor in dementia. Highlights In this paper, we propose a mechanism for Notch1-dependent plasticity that likely underlies the function of Notch1 in memory formation: Notch1 interacts with another important developmental pathway, the Reelin cascade. Notch1 regulates both NMDAR expression and composition. Notch1 influences a cascade of cellular events culminating in CREB activation. PMID:26635527

  20. Respiratory Substrates Regulate S-Nitrosylation of Mitochondrial Proteins through a Thiol-Dependent Pathway

    PubMed Central

    2015-01-01

    S-Nitrosylation is a reversible post-translational modification on cysteinyl thiols that can modulate the function of redox-sensitive proteins. The S-nitrosylation of mitochondrial proteins has been shown to regulate various mitochondrial activities involved in energy-transducing systems and mitochondrion-driven apoptosis. In isolated rat brain mitochondria, we demonstrate that mitochondrial protein S-nitrosylation is regulated by respiratory substrates (glutamate/malate) through a thiol-dependent pathway. Mitochondrial proteins become susceptible to S-nitrosoglutathione (GSNO)-induced S-nitrosylation in mitochondria with an oxidized environment (low glutathione (GSH), NADH, and NADPH, and high GSSG, NAD+, and NADP+) caused by isolation of mitochondria using a discontinuous Percoll gradient. Activation of mitochondrial respiration by respiratory substrates leads to increased NAD(P)H and GSH levels, which in turn reduces mitochondrial S-nitrosylated proteins. 1-Chloro-2,4-dinitrobenzene (CDNB), which depletes mitochondrial GSH and inhibits the thioredoxin–thioredoxin reductase system, prevented the denitrosylation of mitochondrial proteins caused by respiratory substrate treatment. Using biotin-switch coupled with LC-MS/MS, several mitochondrial proteins were identified as targets of S-nitrosylation including adenine nucleotide translocase (ANT) and voltage-dependent anion channel (VDAC), important components of the mitochondria permeability transition pore (MPTP), as well as ATP synthase. The S-nitrosylation of ATP synthase by GSNO was found to inhibit its activity. These findings emphasize the importance of respiratory substrates in regulating S-nitrosylation through a thiol-dependent (GSH and/or thioredoxin) pathway, with implications for mitochondrial bioenergetics and mitochondrion-driven apoptosis. PMID:24716714

  1. Cyp1b1 Regulates Ocular Fissure Closure Through a Retinoic Acid–Independent Pathway

    PubMed Central

    Williams, Antionette L.; Eason, Jessica; Chawla, Bahaar; Bohnsack, Brenda L.

    2017-01-01

    Purpose Mutations in the CYP1B1 gene are the most commonly identified genetic causes of primary infantile-onset glaucoma. Despite this disease association, the role of CYP1B1 in eye development and its in vivo substrate remain unknown. In the present study, we used zebrafish to elucidate the mechanism by which cyp1b1 regulates eye development. Methods Zebrafish eye and neural crest development were analyzed using live imaging of transgenic zebrafish embryos, in situ hybridization, immunostaining, TUNEL assay, and methylacrylate sections. Cyp1b1 and retinoic acid (RA) levels were genetically (morpholino oligonucleotide antisense and mRNA) and pharmacologically manipulated to examine gene function. Results Using zebrafish, we observed that cyp1b1 was expressed in a specific spatiotemporal pattern in the ocular fissures of the developing zebrafish retina and regulated fissure patency. Decreased Cyp1b1 resulted in the premature breakdown of laminin in the ventral fissure and altered subsequent neural crest migration into the anterior segment. In contrast, cyp1b1 overexpression inhibited cell survival in the ventral ocular fissure and prevented fissure closure via an RA-independent pathway. Cyp1b1 overexpression also inhibited the ocular expression of vsx2, pax6a, and pax6b and increased the extraocular expression of shha. Importantly, embryos injected with human wild-type but not mutant CYP1B1 mRNA also showed colobomas, demonstrating the evolutionary and functional conservation of gene function between species. Conclusions Cyp1b1 regulation of ocular fissure closure indirectly affects neural crest migration and development through an RA-independent pathway. These studies provide insight into the role of Cyp1b1 in eye development and further elucidate the pathogenesis of primary infantile-onset glaucoma. PMID:28192799

  2. The Che4 pathway of Myxococcus xanthus regulates type IV pilus-mediated motility.

    PubMed

    Vlamakis, Hera C; Kirby, John R; Zusman, David R

    2004-06-01

    Myxococcus xanthus co-ordinates cell movement during its complex life cycle using multiple chemotaxis-like signal transduction pathways. These pathways regulate both type IV pilus-mediated social (S) motility and adventurous (A) motility. During a search for new chemoreceptors, we identified the che4 operon, which encodes homologues to a MCP (methyl-accepting chemotaxis protein), two CheWs, a hybrid CheA-CheY, a response regulator and a CheR. Deletion of the che4 operon did not cause swarming or developmental defects in either the wild-type (A(+)S(+)) strain or in a strain sustaining only A motility (A(+)S(-)). However, in a strain displaying only S motility (A(-)S(+)), deletion of the che4 operon or the gene encoding the response regulator, cheY4, caused enhanced vegetative swarming and prevented aggregation and sporulation. In contrast, deletion of mcp4 caused reduced vegetative swarming and enhanced development compared with the parent strain. Single-cell analysis of the motility of the A(-)S(+) parent strain revealed a previously unknown inverse correlation between velocity and reversal frequency. Thus, cells that moved at higher velocities showed a reduced reversal frequency. This co-ordination of reversal frequency and velocity was lost in the mcp4 and cheY4 mutants. The structural components of the S motility apparatus were unaffected in the che4 mutants, suggesting that the Che4 system affects reversal frequency of cells by modulating the function of the type IV pilus.

  3. Glucose induction pathway regulates meiosis in Saccharomyces cerevisiae in part by controlling turnover of Ime2p meiotic kinase.

    PubMed

    Gray, Misa; Piccirillo, Sarah; Purnapatre, Kedar; Schneider, Brandt L; Honigberg, Saul M

    2008-08-01

    Several components of the glucose induction pathway, namely the Snf3p glucose sensor and the Rgt1p and Mth1p transcription factors, were shown to be involved in inhibition of sporulation by glucose. The glucose sensors had only a minor role in regulating transcript levels of the two key regulators of meiotic initiation, the Ime1p transcription factor and the Ime2p kinase, but a major role in regulating Ime2p stability. Interestingly, Rgt1p was involved in glucose inhibition of spore formation but not inhibition of Ime2p stability. Thus, the glucose induction pathway may regulate meiosis through both RGT1-dependent and RGT1-independent pathways.

  4. Secretory granule biogenesis and neuropeptide sorting to the regulated secretory pathway in neuroendocrine cells.

    PubMed

    Loh, Y Peng; Kim, Taeyoon; Rodriguez, Yazmin M; Cawley, Niamh X

    2004-01-01

    Neuropeptide precursors synthesized at the rough endoplasmic reticulum are transported and sorted at the trans-Golgi network (TGN) to the granules of the regulated secretory pathway (RSP) of neuroendocrine cells. They are then processed into active peptides and stored in large dense-core granules (LDCGs) until secreted upon stimulation. We have studied the regulation of biogenesis of the LDCGs and the mechanism by which neuropeptide precursors, such as pro-opiomelanocortin (POMC), are sorted into these LDCGs of the RSP in neuroendocrine and endocrine cells. We provide evidence that chromogranin A (CgA), one of the most abundant acidic glycoproteins ubiquitously present in neuroendocrine/endocrine cells, plays an important role in the regulation of LDCG biogenesis. Specific depletion of CgA expression by antisense RNAs in PC12 cells led to a profound loss of secretory granule formation. Exogenously expressed POMC was neither stored nor secreted in a regulated manner in these CgA-deficient PC12 cells. Overexpression of CgA in a CgA- and LDCG-deficient endocrine cell line, 6T3, restored regulated secretion of transfected POMC and the presence of immunoreactive CgA at the tips of the processes of these cells. Unlike CgA, CgB, another granin protein, could not substitute for the role of CgA in regulating LDCG biogenesis. Thus, we conclude that CgA is a key player in the regulation of the biogenesis of LDCGs in neuroendocrine cells. To examine the mechanism of sorting POMC to the LDCGs, we carried out site-directed mutagenesis, transfected the POMC mutants into PC12 cells, and assayed for regulated secretion. Our previous molecular modeling studies predicted a three-dimensional sorting motif in POMC that can bind to a sorting receptor, membrane carboxypeptidase E (CPE). The sorting signal consists of four conserved residues at the N-terminal loop structure of POMC: two acidic residues and two hydrophobic residues. The two acidic residues were predicted to bind to a

  5. Epigenetic Pathways Regulating Bone Homeostasis: Potential Targeting for Intervention of Skeletal Disorders

    PubMed Central

    Gordon, Jonathan A. R.; Montecino, Martin A.; Aqeilan, Rami I.; Stein, Janet L.; Stein, Gary S.; Lian, Jane B.

    2014-01-01

    Epigenetic regulation utilizes different mechanisms to convey heritable traits to progeny cells that are independent of DNA sequence, including DNA silencing, post-translational modifications of histone proteins and the post-transcriptional modulation of RNA transcript levels by non-coding RNAs. Although long non-coding RNAs have recently emerged as important regulators of gene imprinting, but their functions during osteogenesis are as yet unexplored. In contrast, microRNAs (miRNAs) are well characterized for their control of osteogenic and osteoclastic pathways; thus, further defining how gene regulatory networks essential for skeleton functions are coordinated and finely tuned through the activities of miRNAs. Roles of miRNAs are constantly expanding as new studies uncover associations with skeletal disorders. The distinct functions of epigenetic regulators and evidence for integrating their activities to control normal bone gene expression and bone disease will be presented. In addition, potential for using “signature microRNAs” to identify, manage and therapeutically treat osteosarcoma will be discussed in this review. PMID:25260661

  6. Epigenetic pathways regulating bone homeostasis: potential targeting for intervention of skeletal disorders.

    PubMed

    Gordon, Jonathan A R; Montecino, Martin A; Aqeilan, Rami I; Stein, Janet L; Stein, Gary S; Lian, Jane B

    2014-12-01

    Epigenetic regulation utilizes different mechanisms to convey heritable traits to progeny cells that are independent of DNA sequence, including DNA silencing, post-translational modifications of histone proteins, and the post-transcriptional modulation of RNA transcript levels by non-coding RNAs. Although long non-coding RNAs have recently emerged as important regulators of gene imprinting, their functions during osteogenesis are as yet unexplored. In contrast, microRNAs (miRNAs) are well characterized for their control of osteogenic and osteoclastic pathways; thus, further defining how gene regulatory networks essential for skeleton functions are coordinated and finely tuned through the activities of miRNAs. Roles of miRNAs are constantly expanding as new studies uncover associations with skeletal disorders. The distinct functions of epigenetic regulators and evidence for integrating their activities to control normal bone gene expression and bone disease will be presented. In addition, potential for using "signature miRNAs" to identify, manage, and therapeutically treat osteosarcoma will be discussed in this review.

  7. TLR ligands and butyrate increase Pyy expression through two distinct but inter-regulated pathways.

    PubMed

    Larraufie, Pierre; Doré, Joël; Lapaque, Nicolas; Blottière, Hervé M

    2017-02-01

    The intestinal epithelium is an active barrier separating the host from its microbiota. It senses microbial compounds through expression of a wide range of receptors including the Toll-like receptors (TLRs). TLRs have been shown to regulate epithelium permeability or secretion of defensin by Paneth cells. However, the expression and function of TLRs in enteroendocrine L-cells, a specific subtype of intestinal cells secreting PYY and GLP-1, have not yet been assessed. PYY and GLP-1 are implicated in regulation of gut motility, food intake and insulin secretion, and are of great interest regarding obesity and type 2 diabetes. Using a cellular model of human L-cells and a reporter system for NF-κB activation pathway, we reported functional expression of TLRs in these cells. Stimulation with specific TLR-agonists increased expression of Pyy but not Proglucagon in an NF-κB-dependent manner. Moreover, the effect of TLR stimulation was additive to butyrate, a product of bacterial fermentation, on Pyy expression. Additionally, butyrate also increased Tlr expression, including Tlr4, and the NF-κB response to TLR stimulation. Altogether, our results demonstrated a role of TLRs in the modulation of Pyy expression and the importance of butyrate, a product of bacterial fermentation in regulation of microbial TLR-dependent sensing.

  8. Lxr regulates lipid metabolic and visual perception pathways during zebrafish development.

    PubMed

    Pinto, Caroline Lucia; Kalasekar, Sharanya Maanasi; McCollum, Catherine W; Riu, Anne; Jonsson, Philip; Lopez, Justin; Swindell, Eric C; Bouhlatouf, Abdel; Balaguer, Patrick; Bondesson, Maria; Gustafsson, Jan-Åke

    2016-01-05

    The Liver X Receptors (LXRs) play important roles in multiple metabolic pathways, including fatty acid, cholesterol, carbohydrate and energy metabolism. To expand the knowledge of the functions of LXR signaling during embryonic development, we performed a whole-genome microarray analysis of Lxr target genes in zebrafish larvae treated with either one of the synthetic LXR ligands T0901317 or GW3965. Assessment of the biological processes enriched by differentially expressed genes revealed a prime role for Lxr in regulating lipid metabolic processes, similarly to the function of LXR in mammals. In addition, exposure to the Lxr ligands induced changes in expression of genes in the neural retina and lens of the zebrafish eye, including the photoreceptor guanylate cyclase activators and lens gamma crystallins, suggesting a potential novel role for Lxr in modulating the transcription of genes associated with visual function in zebrafish. The regulation of expression of metabolic genes was phenotypically reflected in an increased absorption of yolk in the zebrafish larvae, and changes in the expression of genes involved in visual perception were associated with morphological alterations in the retina and lens of the developing zebrafish eye. The regulation of expression of both lipid metabolic and eye specific genes was sustained in 1 month old fish. The transcriptional networks demonstrated several conserved effects of LXR activation between zebrafish and mammals, and also identified potential novel functions of Lxr, supporting zebrafish as a promising model for investigating the role of Lxr during development.

  9. Lxr regulates lipid metabolic and visual perception pathways during zebrafish development

    PubMed Central

    Pinto, Caroline Lucia; Kalasekar, Sharanya Maanasi; McCollum, Catherine W.; Riu, Anne; Jonsson, Philip; Lopez, Justin; Swindell, Eric; Bouhlatouf, Abdel; Balaguer, Patrick; Bondesson, Maria; Gustafsson, Jan-Åke

    2015-01-01

    The Liver X Receptors (LXRs) play important roles in multiple metabolic pathways, including fatty acid, cholesterol, carbohydrate and energy metabolism. To expand the knowledge of the functions of LXR signaling during embryonic development, we performed a whole-genome microarray analysis of Lxr target genes in zebrafish larvae treated with either one of the synthetic LXR ligands T0901317 or GW3965. Assessment of the biological processes enriched by differentially expressed genes revealed a prime role for Lxr in regulating lipid metabolic processes, similarly to the function of LXR in mammals. In addition, exposure to the Lxr ligands induced changes in expression of genes in the neural retina and lens of the zebrafish eye, including the photoreceptor guanylate cyclase activators and lens gamma crystallins, suggesting a potential novel role for Lxr in modulating the transcription of genes associated with visual function in zebrafish. The regulation of expression of metabolic genes was phenotypically reflected in an increased absorption of yolk in the zebrafish larvae, and changes in the expression of genes involved in visual perception were associated with morphological alterations in the retina and lens of the developing zebrafish eye. The regulation of expression of both lipid metabolic and eye specific genes was sustained in 1 month old fish. The transcriptional networks demonstrated several conserved effects of LXR activation between zebrafish and mammals, and also identified potential novel functions of Lxr, supporting zebrafish as a promising model for investigating the role of Lxr during development. PMID:26427652

  10. Hedgehog signaling pathway regulates ovarian cancer invasion and migration via adhesion molecule CD24

    PubMed Central

    Zeng, Chunyan; Chen, Tingtao; Zhang, Yan; Chen, Qi

    2017-01-01

    Hedgehog (Hh) signalling plays an important role in cancer; however, its mechanism in ovarian cancer migration and invasion remains unclear. In the present study, we aimed to clarify the effect of the Hh signalling pathway on ovarian cancer migration and invasion through the regulation of CD24 expression, both in vitro and in vivo. Patients with ovarian cancer (n = 97) were recruited for this study. Evaluation of the explored the role parameters of patients indicated that CD24 expression was negatively associated with age, histological type and lymph node metastasis (p>0.05), but was positively associated with the clinical stage and pathological grading (p<0.05).The in vitro results indicated that the activator (sonic hedgehog, Shh) and inhibitor (GANT61) of Hh signalling significantly enhanced and reduced CD24 expression, respectively, at both the gene and protein levels (p<0.05).The addition of Shh significantly enhanced cellular migration and invasion of SKOV3 cells in vitro (p<0.05) Down regulation of CD24 using siRNA inhibited the tumour-promoting effects of Shh, and the in vivo results confirmed that GANT61 significantly inhibited CD24 expression and reduced tumour growth (p<0.01). In conclusion, the expression of CD24 can be regulated by Hh signalling, and downregulation of CD24 could play an important role in inhibiting ovarian cancer progression. PMID:28382140

  11. Protein cofactor competition regulates the action of a multifunctional RNA helicase in different pathways

    PubMed Central

    Heininger, Annika U.; Hackert, Philipp; Andreou, Alexandra Z.; Boon, Kum-Loong; Memet, Indira; Prior, Mira; Clancy, Anne; Schmidt, Bernhard; Urlaub, Henning; Schleiff, Enrico; Sloan, Katherine E.; Deckers, Markus; Lührmann, Reinhard; Enderlein, Jörg; Klostermeier, Dagmar; Rehling, Peter; Bohnsack, Markus T.

    2016-01-01

    ABSTRACT A rapidly increasing number of RNA helicases are implicated in several distinct cellular processes, however, the modes of regulation of multifunctional RNA helicases and their recruitment to different target complexes have remained unknown. Here, we show that the distribution of the multifunctional DEAH-box RNA helicase Prp43 between its diverse cellular functions can be regulated by the interplay of its G-patch protein cofactors. We identify the orphan G-patch protein Cmg1 (YLR271W) as a novel cofactor of Prp43 and show that it stimulates the RNA binding and ATPase activity of the helicase. Interestingly, Cmg1 localizes to the cytoplasm and to the intermembrane space of mitochondria and its overexpression promotes apoptosis. Furthermore, our data reveal that different G-patch protein cofactors compete for interaction with Prp43. Changes in the expression levels of Prp43-interacting G-patch proteins modulate the cellular localization of Prp43 and G-patch protein overexpression causes accumulation of the helicase in the cytoplasm or nucleoplasm. Overexpression of several G-patch proteins also leads to defects in ribosome biogenesis that are consistent with withdrawal of the helicase from this pathway. Together, these findings suggest that the availability of cofactors and the sequestering of the helicase are means to regulate the activity of multifunctional RNA helicases and their distribution between different cellular processes. PMID:26821976

  12. Regulation of the Phenylpropanoid Pathway: A Mechanism of Selenium Tolerance in Peanut (Arachis hypogaea L.) Seedlings.

    PubMed

    Wang, Guang; Wu, Liying; Zhang, Hong; Wu, Wenjia; Zhang, Mengmeng; Li, Xiaofeng; Wu, Hui

    2016-05-11

    To clarify the mechanisms of selenium (Se) tolerance in peanut seedlings, we grew peanut seedlings with sodium selenite (0, 3, and 6 mg/L), and investigated the phenylpropanoids metabolism in seedling roots. The results showed that selenite up-regulated the expression of genes and related enzyme activities involving in the phenylpropanoids biosynthesis cascade, such as phenylalanine ammonia-lyase, trans-cinnamate-4-hydroxylase, chalcone synthase, chalcone isomerase, and cinnamyl-alcohol dehydrogenase. Selenite significantly increased phenolic acids and flavonoids, which contributed to the alleviation of selenite-induced stress. Moreover, selenite enhanced the formation of endodermis in roots, which may be attributed to the up-regulation of lignin biosynthesis mediated by the selenite-induced changes of H2O2 and NO, which probably regulated the selenite uptake from an external medium. Accumulation of polyphenolic compounds via the phenylpropanoid pathway may be one of the mechanisms of the increasing selenite tolerance in plants, by which peanut seedlings survived in seleniferous soil, accompanied by accumulation of Se.

  13. Perturbation-based analysis and modeling of combinatorial regulation in the yeast sulfur assimilation pathway.

    PubMed

    McIsaac, R Scott; Petti, Allegra A; Bussemaker, Harmen J; Botstein, David

    2012-08-01

    In yeast, the pathways of sulfur assimilation are combinatorially controlled by five transcriptional regulators (three DNA-binding proteins [Met31p, Met32p, and Cbf1p], an activator [Met4p], and a cofactor [Met28p]) and a ubiquitin ligase subunit (Met30p). This regulatory system exerts combinatorial control not only over sulfur assimilation and methionine biosynthesis, but also on many other physiological functions in the cell. Recently we characterized a gene induction system that, upon the addition of an inducer, results in near-immediate transcription of a gene of interest under physiological conditions. We used this to perturb levels of single transcription factors during steady-state growth in chemostats, which facilitated distinction of direct from indirect effects of individual factors dynamically through quantification of the subsequent changes in genome-wide patterns of gene expression. We were able to show directly that Cbf1p acts sometimes as a repressor and sometimes as an activator. We also found circumstances in which Met31p/Met32p function as repressors, as well as those in which they function as activators. We elucidated and numerically modeled feedback relationships among the regulators, notably feedforward regulation of Met32p (but not Met31p) by Met4p that generates dynamic differences in abundance that can account for the differences in function of these two proteins despite their identical binding sites.

  14. Metabolic pathways regulated by TAp73 in response to oxidative stress

    PubMed Central

    Agostini, Massimiliano; Annicchiarico-Petruzzelli, Margherita; Melino, Gerry; Rufini, Alessandro

    2016-01-01

    Reactive oxygen species are involved in both physiological and pathological processes including neurodegeneration and cancer. Therefore, cells have developed scavenging mechanisms to maintain redox homeostasis under control. Tumor suppressor genes play a critical role in the regulation of antioxidant genes. Here, we investigated whether the tumor suppressor gene TAp73 is involved in the regulation of metabolic adaptations triggered in response to oxidative stress. H2O2 treatment resulted in numerous biochemical changes in both control and TAp73 knockout (TAp73−/−) mouse embryonic fibroblasts, however the extent of these changes was more pronounced in TAp73−/− cells when compared to control cells. In particular, loss of TAp73 led to alterations in glucose, nucleotide and amino acid metabolism. In addition, H2O2 treatment resulted in increased pentose phosphate pathway (PPP) activity in null mouse embryonic fibroblasts. Overall, our results suggest that in the absence of TAp73, H2O2 treatment results in an enhanced oxidative environment, and at the same time in an increased pro-anabolic phenotype. In conclusion, the metabolic profile observed reinforces the role of TAp73 as tumor suppressor and indicates that TAp73 exerts this function, at least partially, by regulation of cellular metabolism. PMID:27119504

  15. hCLP46 regulates U937 cell proliferation via Notch signaling pathway

    SciTech Connect

    Ma, Wenzhan; Du, Jie; Chu, Qiaoyun; Wang, Youxin; Liu, Lixin; Song, Manshu; Wang, Wei

    2011-04-29

    Highlights: {yields} Knock down of hCLP46 by RNAi impairs mammalian Notch signaling. {yields} hCLP46 affects neither cell surface Notch1 expression nor ligand-receptor binding. {yields} Knock down of hCLP46 inhibits U937 cell-growth by up-regulation of CDKN1B. -- Abstract: Human CAP10-like protein 46 kDa (hCLP46) is the homolog of Rumi, which is the first identified protein O-glucosyltransferase that modifies Notch receptor in Drosophila. Dysregulation of hCLP46 occurs in many hematologic diseases, but the role of hCLP46 remains unclear. Knockdown of hCLP46 by RNA interference resulted in decreased protein levels of endogenous Notch1, Notch intracellular domain (NICD) and Notch target gene Hes-1, suggesting the impairment of the Notch signaling. However, neither cell surface Notch expression nor ligand binding activities were affected. In addition, down-regulated expression of hCLP46 inhibited the proliferation of U937 cells, which was correlated with increased cyclin-dependent kinase inhibitor (CDKI) CDKN1B (p27) and decreased phosphorylation of retinoblastoma (RB) protein. We showed that lack of hCLP46 results in impaired ligand induced Notch activation in mammalian cell, and hCLP46 regulates the proliferation of U937 cell through CDKI-RB signaling pathway, which may be important for the pathogenesis of leukemia.

  16. REGγ regulates ERα degradation via ubiquitin–proteasome pathway in breast cancer

    SciTech Connect

    Chai, Fan; Liang, Yan; Bi, Jiong; Chen, Li; Zhang, Fan; Cui, Youhong; Jiang, Jun

    2015-01-02

    Highlights: • High expression of REGγ is correlated with ERα status and poor clinical features. • Cell growth, mobility and invasion are significantly impaired by REGγ knockdown. • REGγ indirectly regulates ERα protein expression. - Abstract: REGγ is a proteasome coactivator which regulates proteolytic activity in eukaryotic cells. Abundant lines of evidence have showed that REGγ is over expressed in a number of human carcinomas. However, its precise role in the pathogenesis of cancer is still unclear. In this study, by examining 200 human breast cancer specimens, we demonstrated that REGγ was highly expressed in breast cancers, and the expression of REGγ was positively correlated with breast cancer patient estrogen receptor alpha (ERα) status. Moreover, the expression of REGγ was found positively associated with poor clinical features and low survival rates in ERα positive breast cancer patients. Further cell culture studies using MCF7 and BT474 breast cancer cell lines showed that cell proliferation, motility, and invasion capacities were decreased significantly by REGγ knockdown. Lastly, we demonstrated that REGγ indirectly regulates the degradation of ERα protein via ubiquitin–proteasome pathway. In conclusion, our findings provide the evidence that REGγ expression was positively correlated with ERα status and poor clinical prognosis in ERα positive breast cancer patients. As well, we disclose a new connection between the two molecules that are both highly expressed in most breast cancer cases.

  17. A TRPC5-regulated calcium signaling pathway controls dendrite patterning in the mammalian brain.

    PubMed

    Puram, Sidharth V; Riccio, Antonio; Koirala, Samir; Ikeuchi, Yoshiho; Kim, Albert H; Corfas, Gabriel; Bonni, Azad

    2011-12-15

    Transient receptor potential (TRP) channels have been implicated as sensors of diverse stimuli in mature neurons. However, developmental roles for TRP channels in the establishment of neuronal connectivity remain largely unexplored. Here, we identify an essential function for TRPC5, a member of the canonical TRP subfamily, in the regulation of dendrite patterning in the mammalian brain. Strikingly, TRPC5 knockout mice harbor long, highly branched granule neuron dendrites with impaired dendritic claw differentiation in the cerebellar cortex. In vivo RNAi analyses suggest that TRPC5 regulates dendrite morphogenesis in the cerebellar cortex in a cell-autonomous manner. Correlating with impaired dendrite patterning in the cerebellar cortex, behavioral analyses reveal that TRPC5 knockout mice have deficits in gait and motor coordination. Finally, we uncover the molecular basis of TRPC5's function in dendrite patterning. We identify the major protein kinase calcium/calmodulin-dependent kinase II β (CaMKIIβ) as a critical effector of TRPC5 function in neurons. Remarkably, TRPC5 forms a complex specifically with CaMKIIβ, but not the closely related kinase CaMKIIα, and thereby induces the CaMKIIβ-dependent phosphorylation of the ubiquitin ligase Cdc20-APC at the centrosome. Accordingly, centrosomal CaMKIIβ signaling mediates the ability of TRPC5 to regulate dendrite morphogenesis in neurons. Our findings define a novel function for TRPC5 that couples calcium signaling to a ubiquitin ligase pathway at the centrosome and thereby orchestrates dendrite patterning and connectivity in the brain.

  18. Transcriptional regulation of the Drosophila moira and osa genes by the DREF pathway.

    PubMed

    Nakamura, Kumi; Ida, Hiroyuki; Yamaguchi, Masamitsu

    2008-07-01

    The DNA replication-related element binding factor (DREF) plays an important role in regulation of cell proliferation in Drosophila, binding to DRE and activating transcription of genes carrying this element in their promoter regions. Overexpression of DREF in eye imaginal discs induces a rough eye phenotype in adults, which can be suppressed by half dose reduction of the osa or moira (mor) genes encoding subunits of the BRM complex. This ATP-dependent chromatin remodeling complex is known to control gene expression and the cell cycle. In the 5' flanking regions of the osa and mor genes, DRE and DRE-like sequences exist which contribute to their promoter activities. Expression levels and promoter activities of osa and mor are decreased in DREF knockdown cells and our results in vitro and in cultured cells indicate that transcription of osa and mor is regulated by the DRE/DREF regulatory pathway. In addition, mRNA levels of other BRM complex subunits and a target gene, string/cdc25, were found to be decreased by knockdown of DREF. These results indicate that DREF is involved in regulation of the BRM complex and thereby the cell cycle.

  19. Pds5 regulators segregate cohesion and condensation pathways in Saccharomyces cerevisiae.

    PubMed

    Tong, Kevin; Skibbens, Robert V

    2015-06-02

    Cohesins are required both for the tethering together of sister chromatids (termed cohesion) and subsequent condensation into discrete structures-processes fundamental for faithful chromosome segregation into daughter cells. Differentiating between cohesin roles in cohesion and condensation would provide an important advance in studying chromatin metabolism. Pds5 is a cohesin-associated factor that is essential for both cohesion maintenance and condensation. Recent studies revealed that ELG1 deletion suppresses the temperature sensitivity of pds5 mutant cells. However, the mechanisms through which Elg1 may regulate cohesion and condensation remain unknown. Here, we report that ELG1 deletion from pds5-1 mutant cells results in a significant rescue of cohesion, but not condensation, defects. Based on evidence that Elg1 unloads the DNA replication clamp PCNA from DNA, we tested whether PCNA overexpression would similarly rescue pds5-1 mutant cell cohesion defects. The results indeed reveal that elevated levels of PCNA rescue pds5-1 temperature sensitivity and cohesion defects, but do not rescue pds5-1 mutant cell condensation defects. In contrast, RAD61 deletion rescues the condensation defect, but importantly, neither the temperature sensitivity nor cohesion defects exhibited by pds5-1 mutant cells. In combination, these findings reveal that cohesion and condensation are separable pathways and regulated in nonredundant mechanisms. These results are discussed in terms of a new model through which cohesion and condensation are spatially regulated.

  20. Regulation of mammalian Gli proteins by Costal 2 and PKA in Drosophila reveals Hedgehog pathway conservation.

    PubMed

    Marks, Steven A; Kalderon, Daniel

    2011-06-01

    Hedgehog (Hh) signaling activates full-length Ci/Gli family transcription factors and prevents Ci/Gli proteolytic processing to repressor forms. In the absence of Hh, Ci/Gli processing is initiated by direct Pka phosphorylation. Despite those fundamental similarities between Drosophila and mammalian Hh pathways, the differential reliance on cilia and some key signal transduction components had suggested a major divergence in the mechanisms that regulate Ci/Gli protein activities, including the role of the kinesin-family protein Costal 2 (Cos2), which directs Ci processing in Drosophila. Here, we show that Cos2 binds to three regions of Gli1, just as for Ci, and that Cos2 functions to silence mammalian Gli1 in Drosophila in a Hh-regulated manner. Cos2 and the mammalian kinesin Kif7 can also direct Gli3 and Ci processing in fly, underscoring a fundamental conserved role for Cos2 family proteins in Hh signaling. We also show that direct PKA phosphorylation regulates the activity, rather than the proteolysis of Gli in Drosophilia, and we provide evidence for an analogous action of PKA on Ci.

  1. The Hippo Pathway Targets Rae1 to Regulate Mitosis and Organ Size and to Feed Back to Regulate Upstream Components Merlin, Hippo, and Warts

    PubMed Central

    Jenny, Andreas; Pfleger, Cathie M.

    2016-01-01

    Hippo signaling acts as a master regulatory pathway controlling growth, proliferation, and apoptosis and also ensures that variations in proliferation do not alter organ size. How the pathway coordinates restricting proliferation with organ size control remains a major unanswered question. Here we identify Rae1 as a highly-conserved target of the Hippo Pathway integrating proliferation and organ size. Genetic and biochemical studies in Drosophila cells and tissues and in mammalian cells indicate that Hippo signaling promotes Rae1 degradation downstream of Warts/Lats. In proliferating cells, Rae1 loss restricts cyclin B levels and organ size while Rae1 over-expression increases cyclin B levels and organ size, similar to Hippo Pathway over-activation or loss-of-function, respectively. Importantly, Rae1 regulation by the Hippo Pathway is crucial for its regulation of cyclin B and organ size; reducing Rae1 blocks cyclin B accumulation and suppresses overgrowth caused by Hippo Pathway loss. Surprisingly, in addition to suppressing overgrowth, reducing Rae1 also compromises survival of epithelial tissue overgrowing due to loss of Hippo signaling leading to a tissue “synthetic lethality” phenotype. Excitingly, Rae1 plays a highly conserved role to reduce the levels and activity of the Yki/YAP oncogene. Rae1 increases activation of the core kinases Hippo and Warts and plays a post-transcriptional role to increase the protein levels of the Merlin, Hippo, and Warts components of the pathway; therefore, in addition to Rae1 coordinating organ size regulation with proliferative control, we propose that Rae1 also acts in a feedback circuit to regulate pathway homeostasis. PMID:27494403

  2. Coordinated Regulation of Species-Specific Hydroxycinnamic Acid Degradation and Siderophore Biosynthesis Pathways in Agrobacterium fabrum

    PubMed Central

    Baude, Jessica; Vial, Ludovic; Villard, Camille; Campillo, Tony; Lavire, Céline; Nesme, Xavier

    2016-01-01

    ABSTRACT The rhizosphere-inhabiting species Agrobacterium fabrum (genomospecies G8 of the Agrobacterium tumefaciens species complex) is known to degrade hydroxycinnamic acids (HCAs), especially ferulic acid and p-coumaric acid, via the novel A. fabrum HCA degradation pathway. Gene expression profiles of A. fabrum strain C58 were investigated in the presence of HCAs, using a C58 whole-genome oligoarray. Both ferulic acid and p-coumaric acid caused variations in the expression of more than 10% of the C58 genes. Genes of the A. fabrum HCA degradation pathway, together with the genes involved in iron acquisition, were among the most highly induced in the presence of HCAs. Two operons coding for the biosynthesis of a particular siderophore, as well as genes of the A. fabrum HCA degradation pathway, have been described as being specific to the species. We demonstrate here their coordinated expression, emphasizing the interdependence between the iron concentration in the growth medium and the rate at which ferulic acid is degraded by cells. The coordinated expression of these functions may be advantageous in HCA-rich but iron-starved environments in which microorganisms have to compete for both iron and carbon sources, such as in plant roots. The present results confirm that there is cooperation between the A. fabrum-specific genes, defining a particular ecological niche. IMPORTANCE We previously identified seven genomic regions in Agrobacterium fabrum that were specifically present in all of the members of this species only. Here we demonstrated that two of these regions, encoding the hydroxycinnamic acid degradation pathway and the iron acquisition pathway, were regulated in a coordinated manner. The coexpression of these functions may be advantageous in hydroxycinnamic acid-rich but iron-starved environments in which microorganisms have to compete for both iron and carbon sources, such as in plant roots. These data support the view that bacterial genomic species

  3. Pathways from marital aggression to infant emotion regulation: the development of withdrawal in infancy.

    PubMed

    Crockenberg, Susan C; Leerkes, Esther M; Lekka, Shamila K

    2007-02-01

    Associations between marital conflict and infant emotion regulation exist, but explanatory pathways have not been explored. For older children, parental behavior partially mediates this association through a "spillover" process. We test: associations between mothers' and fathers' verbally aggressive marital conflict, infant temperament, and infant withdrawal; mediating effects of negative maternal behavior, and moderating effects of infant temperament, exposure to marital arguments, and contact with father. Eighty mothers, 73 fathers, and their 6-month-old infants participated; parents reported marital aggression prenatally, mothers reported infant exposure to arguments, direct caregiving by father, and infant temperament at 5 months. Negative maternal behavior, infant withdrawal, distress to novelty, activity, and look away were observed at 6 months. Mothers' and fathers' aggressive marital conflict predicted infant withdrawal, interactively with exposure to marital arguments and extent of father caregiving, as did infant temperament and negative maternal behavior. Maternal behavior did not mediate between marital conflict and withdrawal.

  4. Rhodopsin-Regulated Insulin Receptor Signaling Pathway in Rod Photoreceptor Neurons

    PubMed Central

    Rajala, Raju V.S.; Anderson, Robert E.

    2010-01-01

    The retina is an integral part of the central nervous system and retinal cells are known to express insulin receptors (IR), although their function is not known. This article describes recent studies that link the photoactivation of rhodopsin to tyrosine phosphorylation of the IR and subsequent activation of phosphoinositide 3-kinase (PI3K), a neuron survival factor. Our studies suggest that the physiological role of this process is to provide neuroprotection of the retina against light-damage by activating proteins that protect against stress-induced apoptosis. We focus mainly on our recently identified regulation of the IR pathway through the G-protein-coupled receptor rhodopsin. Various mutant and knockout proteins of phototransduction cascade have been used to study the light-induced activation of the retinal IR. Our studies suggest that rhodopsin may have additional previously uncharacterized signaling functions in photoreceptors. PMID:20407846

  5. Genome Engineering of the 2,3-Butanediol Biosynthetic Pathway for Tight Regulation in Cyanobacteria.

    PubMed

    Nozzi, Nicole E; Atsumi, Shota

    2015-11-20

    Cyanobacteria have gained popularity among the metabolic engineering community as a tractable photosynthetic host for renewable chemical production. However, though a number of successfully engineered production systems have been reported, long-term genetic stability remains an issue for cyanobacterial systems. The genetic engineering toolbox for cyanobacteria is largely lacking inducible systems for expression control. The characterization of tight regulation systems for use in cyanobacteria may help to alleviate this problem. In this work we explore the function of the IPTG inducible promoter P(L)lacO1 in the model cyanobacterium Synechococcus elongatus PCC 7942 as well as the effect of gene order within an operon on pathway expression. According to our experiments, P(L)lacO1 functions well as an inducible promoter in S. elongatus. Additionally, we found that gene order within an operon can strongly influence control of expression of each gene.

  6. Current advances in DNA repair: regulation of enzymes and pathways involved in maintaining genomic stability.

    PubMed

    Neher, Tracy M; Turchi, John J

    2011-06-15

    Novel discoveries in the DNA repair field have lead to continuous and rapid advancement of our understanding of not only DNA repair but also DNA replication and recombination. Research in the field transcends numerous areas of biology, biochemistry, physiology, and medicine, making significant connections across these broad areas of study. From early studies conducted in bacterial systems to current analyses in eukaryotic systems and human disease, the innovative research into the mechanisms of repair machines and the consequences of ineffective DNA repair has impacted a wide scientific community. This Forum contains a select mix of primary research articles in addition to a number of timely reviews covering a subset of DNA repair pathways where recent advances and novel discoveries are improving our understanding of DNA repair, its regulation, and implications to human disease.

  7. S-nitrosothiols regulate nitric oxide production and storage in plants through the nitrogen assimilation pathway.

    PubMed

    Frungillo, Lucas; Skelly, Michael J; Loake, Gary J; Spoel, Steven H; Salgado, Ione

    2014-11-11

    Nitrogen assimilation plays a vital role in plant metabolism. Assimilation of nitrate, the primary source of nitrogen in soil, is linked to the generation of the redox signal nitric oxide (NO). An important mechanism by which NO regulates plant development and stress responses is through S-nitrosylation, that is, covalent attachment of NO to cysteine residues to form S-nitrosothiols (SNO). Despite the importance of nitrogen assimilation and NO signalling, it remains largely unknown how these pathways are interconnected. Here we show that SNO signalling suppresses both nitrate uptake and reduction by transporters and reductases, respectively, to fine tune nitrate homeostasis. Moreover, NO derived from nitrate assimilation suppresses the redox enzyme S-nitrosoglutathione Reductase 1 (GSNOR1) by S-nitrosylation, preventing scavenging of S-nitrosoglutathione, a major cellular bio-reservoir of NO. Hence, our data demonstrates that (S)NO controls its own generation and scavenging by modulating nitrate assimilation and GSNOR1 activity.

  8. HJURP regulates cellular senescence in human fibroblasts and endothelial cells via a p53-dependent pathway.

    PubMed

    Heo, Jong-Ik; Cho, Jung Hee; Kim, Jae-Ryong

    2013-08-01

    Holliday junction recognition protein (HJURP), a centromere protein-A (CENP-A) histone chaperone, mediates centromere-specific assembly of CENP-A nucleosome, contributing to high-fidelity chromosome segregation during cell division. However, the role of HJURP in cellular senescence of human primary cells remains unclear. We found that the expression levels of HJURP decreased in human dermal fibroblasts and umbilical vein endothelial cells in replicative or premature senescence. Ectopic expression of HJURP in senescent cells partially overcame cell senescence. Conversely, downregulation of HJURP in young cells led to premature senescence. p53 knockdown, but not p16 knockdown, abolished senescence phenotypes caused by HJURP reduction. These data suggest that HJURP plays an important role in the regulation of cellular senescence through a p53-dependent pathway and might contribute to tissue or organismal aging and protection of cellular transformation.

  9. Regulation of human myometrial contractility during pregnancy and labour: are calcium homeostatic pathways important?

    PubMed

    Tribe, R M

    2001-03-01

    If we are to develop new strategies for the treatment and management of preterm and dysfunctional term labour, it is imperative that we improve current understanding of the control of human uterine activity. Despite many studies of animal pregnancy, there is a paucity of knowledge relating to the complex control of human myometrium during pregnancy. It is hypothesized that human myometrium is relatively quiescent during the majority of pregnancy and that as term approaches there is cascade of molecular events that prepare the uterus for labour. This review will consider the cellular mechanisms involved in the regulation of human myometrial activity and the modulation of these by hormonal and mechanical signals. In particular, the contribution of calcium homeostatic pathways to the control of human myometrial contractility during gestation will be discussed. Experimental Physiology (2001) 86.2, 247-254.

  10. MicroRNAs mediated regulation of MAPK signaling pathways in chronic myeloid leukemia

    PubMed Central

    Sharma, Garima; Lee, Sang-Soo

    2016-01-01

    Chronic myeloid leukemia (CML) is a severe problem throughout the world and requires identification of novel targets for its treatment. This multifactorial disease accounts for about 15% of the all diagnosed leukemia cases. Mitogen-activated protein kinase (MAPK) signaling pathway is crucial for the cell survival and its dysregulation is being implicated in various types of cancers. In here, we have discussed the potential role of various miRNAs that are found involved in regulating the proteins cascades of MAPK signaling pathway associated with CML. An emphasis has been paid to summarize the influence of various miRNAs in elevating or suppressing the expression level of significant proteins such as miR-203, miR-196a, miR-196b, miR-30a, miR-29b, miR-138 in BCR-ABL tyrosine kinase; miR-126, miR-221, miR-128, miR-15a, miR-188-5p, miR-17 in CRK family proteins; miR-155, miR-181a with SOS proteins; miR-155, miR-19a, with KRAS proteins; miR-19a with RAF1 protein; and miR-17, miR-19a, miR-17-92 cluster with MAPK/ERK proteins. In light of ever-increasing importance and ever-widening regulatory roles of miRNAs in cells, we have reviewed the recent progress in the field of miRNAs and have tried to suggest them as controlling targets for various protein cascades of MAPK signaling pathway. An understanding of the supervisory mechanism of MAPK by miRNAs might provide novel targets for treating CML. PMID:26967056

  11. BMP9 Crosstalk with the Hippo Pathway Regulates Endothelial Cell Matricellular and Chemokine Responses

    PubMed Central

    Young, Kira; Tweedie, Eric; Conley, Barbara; Ames, Jacquelyn; FitzSimons, MaryLynn; Brooks, Peter; Liaw, Lucy; Vary, Calvin P. H.

    2015-01-01

    Endoglin is a type III TGFβ auxiliary receptor that is upregulated in endothelial cells during angiogenesis and, when mutated in humans, results in the vascular disease hereditary hemorrhagic telangiectasia (HHT). Though endoglin has been implicated in cell adhesion, the underlying molecular mechanisms are still poorly understood. Here we show endoglin expression in endothelial cells regulates subcellular localization of zyxin in focal adhesions in response to BMP9. RNA knockdown of endoglin resulted in mislocalization of zyxin and altered formation of focal adhesions. The mechanotransduction role of focal adhesions and their ability to transmit regulatory signals through binding of the extracellular matrix are altered by endoglin deficiency. BMP/TGFβ transcription factors, SMADs, and zyxin have recently been implicated in a newly emerging signaling cascade, the Hippo pathway. The Hippo transcription coactivator, YAP1 (yes-associated protein 1), has been suggested to play a crucial role in mechanotransduction and cell-cell contact. Identification of BMP9-dependent nuclear localization of YAP1 in response to endoglin expression suggests a mechanism of crosstalk between the two pathways. Suppression of endoglin and YAP1 alters BMP9-dependent expression of YAP1 target genes CCN1 (cysteine-rich 61, CYR61) and CCN2 (connective tissue growth factor, CTGF) as well as the chemokine CCL2 (monocyte chemotactic protein 1, MCP-1). These results suggest a coordinate effect of endoglin deficiency on cell matrix remodeling and local inflammatory responses. Identification of a direct link between the Hippo pathway and endoglin may reveal novel mechanisms in the etiology of HHT. PMID:25909848

  12. Structurally similar estradiol analogs uniquely alter the regulation of intracellular signaling pathways.

    PubMed

    Yarger, James G; Babine, Robert E; Bittner, Michael; Shanle, Erin; Xu, Wei; Hershberger, Pamela; Nye, Steven H

    2013-02-01

    Ligand structure can affect the activation of nuclear receptors, such as estrogen receptors (ERs), and their control of signaling pathways for cellular responses including death and differentiation. We hypothesized that distinct biological functions of similar estradiol (E(2)) analogs could be identified by integrating gene expression patterns obtained from human tumor cell lines with receptor binding and functional data for the purpose of developing compounds for treatment of a variety of diseases. We compared the estrogen receptor subtype selectivity and impact on signaling pathways for three distinct, but structurally similar, analogs of E(2). Modifications in the core structure of E(2) led to pronounced changes in subtype selectivity for estrogen receptors, ER-α or ER-β, along with varying degrees of ER dimerization and activation. While all three E(2) analogs are predominantly ER-β agonists, the cell growth inhibitory activity commonly associated with this class of compounds was detected for only two of the analogs and might be explained by a ligand-specific pattern of gene transcription. Microarray studies using three different human tumor cell lines demonstrated that the analogs distinctly affect the transcription of genes in signaling pathways for chromosome replication, cell death, and oligodendrocyte progenitor cell differentiation. That the E(2) analogs could lower tumor cell viability and stimulate neuronal differentiation confirmed that gene expression data could accurately distinguish biological activity of the E(2) analogs. The findings reported here confirm that cellular responses can be regulated by making key structural alterations to the core structure of endogenous ER ligands.

  13. Coprinus comatus Cap Inhibits Adipocyte Differentiation via Regulation of PPARγ and Akt Signaling Pathway

    PubMed Central

    Jang, Sun-Hee; Kang, Suk Nam; Jeon, Beong-Sam; Ko, Yeoung-Gyu; Kim, Hong-Duck; Won, Chung-Kil; Kim, Gon-Sup; Cho, Jae-Hyeon

    2014-01-01

    This study assessed the effects of Coprinus comatus cap (CCC) on adipogenesis in 3T3-L1 adipocytes and the effects of CCC on the development of diet-induced obesity in rats. Here, we showed that the CCC has an inhibitory effect on the adipocyte differentiation of 3T3-L1 cells, resulting in a significant decrease in lipid accumulation through the downregulation of several adipocyte specific-transcription factors, including CCAAT/enhancer binding protein β, C/EBPδ, and peroxisome proliferator-activated receptor gamma (PPARγ). Moreover, treatment with CCC during adipocyte differentiation induced a significant down-regulation of PPARγ and adipogenic target genes, including adipocyte protein 2, lipoprotein lipase, and adiponectin. Interestingly, the CCC treatment of the 3T3-L1 adipocytes suppressed the insulin-stimulated Akt and GSK3β phosphorylation, and these effects were stronger in the presence of an inhibitor of Akt phosphorylation, LY294002, suggesting that CCC inhibited adipocyte differentiation through the down-regulation of Akt signaling. In the animal study, CCC administration significantly reduced the body weight and adipose tissue weight of rats fed a high fat diet (HFD) and attenuated lipid accumulation in the adipose tissues of the HFD-induced obese rats. The size of the adipocyte in the epididymal fat of the CCC fed rats was significantly smaller than in the HFD rats. CCC treatment significantly reduced the total cholesterol and triglyceride levels in the serum of HFD rats. These results strongly indicated that the CCC-mediated decrease in body weight was due to a reduction in adipose tissue mass. The expression level of PPARγ and phospho-Akt was significantly lower in the CCC-treated HFD rats than that in the HFD obesity rats. These results suggested that CCC inhibited adipocyte differentiation by the down-regulation of major transcription factor involved in the adipogenesis pathway including PPARγ through the regulation of the Akt pathway in 3T3

  14. Changes in signaling pathways regulating neuroplasticity induced by neurokinin 1 receptor knockout.

    PubMed

    Musazzi, Laura; Perez, Jorge; Hunt, Stephen P; Racagni, Giorgio; Popoli, Maurizio

    2005-03-01

    Neurokinin 1 (NK-1) receptor knockout mice showed behavioral responses similar to animals chronically treated with antidepressants. The aim of this study was to analyse, in NK-1 receptor knockout, the molecular modifications of signaling pathways involved in the pathophysiology of depression and antidepressant mechanism. We found, in total cell cytosol from the prefrontal/frontal cortex, hippocampus and striatum, a marked up-regulation of Ca(2+)-independent enzymatic activity and Thr(286) autophosphorylation of Ca(2+)/calmodulin-dependent protein kinase (CaMK) II. Similar changes in CaMKII regulation were previously observed in rats chronically treated with antidepressants. In striatum, up-regulation of the activity and phosphorylation of CaMKII was also found in the homogenate and synaptosomes. No major changes were observed in the Ca(2+)-dependent kinase activity, with the exception of homogenate from the prefrontal/frontal cortex. We also analysed the expression and phosphorylation of presynaptic proteins, which modulate synaptic vesicle trafficking and exocytosis, and found a marked decrease in synapsin I total expression and basal phosphorylation of Ser(603) (the phosphorylation site for CaMKII) in the prefrontal/frontal cortex. Accordingly, the Ca(2+)/calmodulin-dependent posthoc endogenous phosphorylation of synapsin I in the same area was increased. The knockout of NK-1 receptor had no consequences on the expression or phosphorylation levels of the transcription factor cAMP-responsive element-binding protein and its regulating kinase CaMKIV. However, phosphorylation of ERK1/2-mitogen-activated protein kinases was reduced in the hippocampus and striatum, again resembling an effect previously observed in antidepressant-treated rats. These results show similarities between NK-1 knockouts and animals chronically treated with antidepressants and support the putative antidepressant activity of NK-1 receptor antagonists.

  15. Salicylic Acid Regulates Pollen Tip Growth through an NPR3/NPR4-Independent Pathway.

    PubMed

    Rong, Duoyan; Luo, Nan; Mollet, Jean Claude; Liu, Xuanming; Yang, Zhenbiao

    2016-11-07

    Tip growth is a common strategy for the rapid elongation of cells to forage the environment and/or to target to long-distance destinations. In the model tip growth system of Arabidopsis pollen tubes, several small-molecule hormones regulate their elongation, but how these rapidly diffusing molecules control extremely localized growth remains mysterious. Here we show that the interconvertible salicylic acid (SA) and methylated SA (MeSA), well characterized for their roles in plant defense, oppositely regulate Arabidopsis pollen tip growth with SA being inhibitory and MeSA stimulatory. The effect of SA and MeSA was independent of known NPR3/NPR4 SA receptor-mediated signaling pathways. SA inhibited clathrin-mediated endocytosis in pollen tubes associated with an increased accumulation of less stretchable demethylated pectin in the apical wall, whereas MeSA did the opposite. Furthermore, SA and MeSA alter the apical activation of ROP1 GTPase, a key regulator of tip growth in pollen tubes, in an opposite manner. Interestingly, both MeSA methylesterase and SA methyltransferase, which catalyze the interconversion between SA and MeSA, are localized at the apical region of pollen tubes, indicating of the tip-localized production of SA and MeSA and consistent with their effects on the apical cellular activities. These findings suggest that local generation of a highly diffusible signal can regulate polarized cell growth, providing a novel mechanism of cell polarity control apart from the one involving protein and mRNA polarization.

  16. TRIM14 regulates cell proliferation and invasion in osteosarcoma via promotion of the AKT signaling pathway

    PubMed Central

    Xu, Guoxing; Guo, Yongfei; Xu, Dabo; Wang, Yi; Shen, Yafeng; Wang, Feifei; Lv, Yuanyuan; Song, Fanglong; Jiang, Dawei; Zhang, Yinquan; Lou, Yi; Meng, Yake; Yang, Yongji; Kang, Yifan

    2017-01-01

    Recent studies have shown that some members of the tripartite motif-containing protein (TRIM) family serve as important regulators of tumorigenesis. However, the biological role of TRIM14 in osteosarcoma remains to be established. In this study, we showed that TRIM14 is upregulated in human osteosarcoma specimens and cell lines, and correlated with osteosarcoma progression and shorter patient survival times. Functional studies demonstrated that overexpression of TRIM14 enhances osteosarcoma cell proliferation, clone formation, cell cycle procession, migration and invasion in vitro and promotes tumor growth in vivo, and conversely, its silencing has the opposite effects. Furthermore, TRIM14 overexpression induced activation of the AKT pathway. Inhibition of AKT expression reversed the TRIM14-mediated promotory effects on cell growth and mobility, in addition to TRIM14-induced epithelial-to-mesenchymal transition (EMT) and cyclin D1 upregulation. Our findings collectively suggest that TRIM14 functions as an oncogene by upregulating the AKT signaling pathway in osteosarcoma cells, supporting its potential utility as a therapeutic target for this disease. PMID:28205534

  17. TRPM3 channels provide a regulated influx pathway for zinc in pancreatic beta cells.

    PubMed

    Wagner, Thomas F J; Drews, Anna; Loch, Sabine; Mohr, Florian; Philipp, Stephan E; Lambert, Sachar; Oberwinkler, Johannes

    2010-09-01

    Zinc is stored in insulin-containing dense core vesicles of pancreatic beta-cells where it forms crystals together with insulin and calcium ions. Zinc ions are therefore released together with insulin upon exocytosis of these vesicles. Consequently, pancreatic beta-cells need to take up large amounts of zinc from the extracellular space across their plasma membrane. The pathways for zinc uptake are only partially understood. TRPM3 channels are present in pancreatic beta-cells and can be activated by the endogenous steroid pregnenolone sulfate. We demonstrate here that recombinant TRPM3 channels are highly permeable for many divalent cations, in particular also for zinc ions. Importantly, TRPM3 channels endogenously expressed in pancreatic beta-cells are also highly permeable for zinc ions. Using FluoZin3 to image changes of the intracellular zinc concentration, we show that pancreatic beta-cells take up zinc through TRPM3 channels even when extracellular zinc concentrations are low and physiological levels of calcium and magnesium are present. Activation of TRPM3 channels also leads to depolarization of beta-cells and to additional zinc influx through voltage-gated calcium channels. Our data establish that TRPM3 channels constitute a regulated entry pathway for zinc ions in pancreatic beta-cells.

  18. Hypoxia Regulates mTORC1-Mediated Keratinocyte Motility and Migration via the AMPK Pathway

    PubMed Central

    Yan, Tiantian; Zhang, Junhui; Tang, Di; Zhang, Xingyue; Jiang, Xupin; Zhao, Liping; Zhang, Qiong; Zhang, Dongxia; Huang, Yuesheng

    2017-01-01

    Keratinocyte migration, the initial event and rate-limiting step in wound healing, plays a vital role in restoration of the intact skin barrier, also known as re-epithelialization. After acute tissue injury, hypoxic microenvironment gradually develops and acts as an early stimulus to initiate the healing process. Although we have previously found that hypoxia induces keratinocyte migration, the underlying mechanism remains unknown. Here, we first observed that hypoxia increased mTORC1 activity. Recombinant lentivirus vector and Rapamycin were used for silencing mTORC1 in HaCaT cells and primary mouse keratinocytes (MKs). Using cell migration assay and a Zeiss chamber equipped with imaging system, we also demonstrated that mTORC1 downregulation reversed hypoxia-induced keratinocyte motility and lateral migration. Importantly, hypoxia-activated mTORC1 was accompanied by the AMPK downregulation, and we found that the AMPK pathway activators Metformin (Met) and 5-Aminoimidazole-4-carboxamide 1-β-D-ribofuranoside (AICAR) decreased the mTORC1 activity, cell motility and lateral migration. Thus, our results suggest that hypoxia regulates mTORC1-mediated keratinocyte motility and migration via the AMPK pathway. PMID:28068384

  19. Signal pathways regulating hyaluronan secretion into static and cycled synovial joints of rabbits.

    PubMed

    Ingram, K R; Wann, A K T; Wingate, R M; Coleman, P J; McHale, N; Levick, J R

    2009-09-01

    Joint lubrication, synovial fluid conservation and many pathophysiological processes depend on hyaluronan (HA). Intra-articular HA injection and exercise, which stimulates articular HA production, ameliorate osteoarthritis. We therefore investigated the pathways regulating movement-stimulated articular HA secretion rate ( ) in vivo. Endogenous HA was removed from the knee joint cavity of anaesthetised rabbits by washout. Joints were then cycled passively or remained static for 5 h, with/without intra-articular agonist/inhibitor, after which newly secreted HA was harvested for analysis. Movement almost doubled . Similar or larger increases were elicited in static joints by the intra-articular Ca(2+) ionophore ionomycin, prostaglandin E(2), cAMP-raising agents, serine/threonine phosphatase inhibitor and activation of protein kinase C (PKC). PKC-stimulated secretion was inhibited by the PKC inhibitor bisindolylmaleimide I and inhibitors of the downstream kinases MEK-ERK (U0126, PD98059). These agents inhibited movement-stimulated secretion of HA (MSHA) only when the parallel p38 kinase path was simultaneously inhibited by SB203580 (ineffective alone). The phospholipase C inhibitor U73122 almost fully blocked MSHA (P = 0.001, n = 10), without affecting static . The ENaC channel blocker amiloride inhibited MSHA, whereas other inhibitors of stretch-activated channels (Gd(3+), ruthenium red, SKF96365) did not. It is proposed that MSHA may be mediated by PLC activation, leading to activation of parallel PKC-MEK-ERK and p38 kinase pathways.

  20. miR-323a-3p regulates lung fibrosis by targeting multiple profibrotic pathways

    PubMed Central

    Habiel, David M.; Hansbro, Phil M.; Kim, Richard Y.; Gharib, Sina A.; Edelman, Jeffery D.; Königshoff, Melanie; Parimon, Tanyalak; Huang, Ying; Allen, Jenieke; Jiang, Dianhua; Kurkciyan, Adrianne A.; Mizuno, Takako; Stripp, Barry R.; Noble, Paul W.; Hogaboam, Cory M.

    2016-01-01

    Maladaptive epithelial repair from chronic injury is a common feature in fibrotic diseases, which in turn activates a pathogenic fibroblast response that produces excessive matrix deposition. Dysregulated microRNAs (miRs) can regulate expression of multiple genes and fundamentally alter cellular phenotypes during fibrosis. Although several miRs have been shown to be associated with lung fibrosis, the mechanisms by which miRs modulate epithelial behavior in lung fibrosis are lacking. Here, we identified miR-323a-3p to be downregulated in the epithelium of lungs with bronchiolitis obliterans syndrome (BOS) after lung transplantation, idiopathic pulmonary fibrosis (IPF), and murine bleomycin-induced fibrosis. Antagomirs for miR-323a-3p augment, and mimics suppress, murine lung fibrosis after bleomycin injury, indicating that this miR may govern profibrotic signals. We demonstrate that miR-323a-3p attenuates TGF-α and TGF-β signaling by directly targeting key adaptors in these important fibrogenic pathways. Moreover, miR-323a-3p lowers caspase-3 expression, thereby limiting programmed cell death from inducers of apoptosis and ER stress. Finally, we find that epithelial expression of miR-323a-3p modulates inhibitory crosstalk with fibroblasts. These studies demonstrate that miR-323a-3p has a central role in lung fibrosis that spans across murine and human disease, and downregulated expression by the lung epithelium releases inhibition of various profibrotic pathways to promote fibroproliferation. PMID:27942594

  1. Frankincense and myrrh suppress inflammation via regulation of the metabolic profiling and the MAPK signaling pathway

    PubMed Central

    Su, Shulan; Duan, Jinao; Chen, Ting; Huang, Xiaochen; Shang, Erxin; Yu, Li; Wei, Kaifeng; Zhu, Yue; Guo, Jianming; Guo, Sheng; Liu, Pei; Qian, Dawei; Tang, Yuping

    2015-01-01

    Frankincense and myrrh are highly effective in treatment of inflammatary diseases, but lacking of the therapy mechanisms. We undertook this stuty to evaluate the effects on Adjuvant-induced Arthritis (AIA) rats and to explore the underlying mechanisms by analyzing the metabolic profiling and signaling pathway evaluated by expression of inflammatory cytokines, c-jun and c-fos and corresponding phosphorylation levels. The results stated the elevated expression levels of TNFα, PGE2, IL-2, NO, and MDA in serum and swelling paw of AIA rats were significantly decreased after treatment, which exerted more remarkable inhibitive effects of combined therapy. The metbolic profiling of plasma and urine were clearly improved and twenty-one potential biomarkers were identified. Moreover, the inhibited effects of five bioactive components on cytokine transcription in PHA stimulated-PBMC showed the MAPK pathway might account for this phenomenon with considerable reduction in phosphorylated forms of all the three MAPK (ERK1/2, p38 and JNK) and down regulation of c-jun and c-fos. PMID:26329643

  2. miR-58 family and TGF-β pathways regulate each other in Caenorhabditis elegans

    PubMed Central

    de Lucas, María Pilar; Sáez, Alberto G.; Lozano, Encarnación

    2015-01-01

    Despite the fact that microRNAs (miRNAs) modulate the expression of around 60% of protein-coding genes, it is often hard to elucidate their precise role and target genes. Studying miRNA families as opposed to single miRNAs alone increases our chances of observing not only mutant phenotypes but also changes in the expression of target genes. Here we ask whether the TGF-β signalling pathways, which control many animal processes, might be modulated by miRNAs in Caenorhabditis elegans. Using a mutant for four members of the mir-58 family, we show that both TGF-β Sma/Mab (controlling body size) and TGF-β Dauer (regulating dauer, a stress-resistant larval stage) are upregulated. Thus, mir-58 family directly inhibits the expression of dbl-1 (ligand), daf-1, daf-4 and sma-6 (receptors) of TGF-β pathways. Epistasis experiments reveal that whereas the small body phenotype of the mir-58 family mutant must invoke unknown targets independent from TGF-β Sma/Mab, its dauer defectiveness can be rescued by DAF-1 depletion. Additionally, we found a negative feedback loop between TGF-β Sma/Mab and mir-58 and the related mir-80. Our results suggest that the interaction between mir-58 family and TGF-β genes is key on decisions about animal growth and stress resistance in C. elegans and perhaps other organisms. PMID:26400166

  3. Differential Maturation of the Two Regulated Secretory Pathways in Human iPSC-Derived Neurons.

    PubMed

    Emperador Melero, Javier; Nadadhur, Aishwarya G; Schut, Desiree; Weering, Jan V; Heine, Vivi M; Toonen, Ruud F; Verhage, Matthijs

    2017-03-14

    Neurons communicate by regulated secretion of chemical signals from synaptic vesicles (SVs) and dense-core vesicles (DCVs). Here, we investigated the maturation of these two secretory pathways in micro-networks of human iPSC-derived neurons. These micro-networks abundantly expressed endogenous SV and DCV markers, including neuropeptides. DCV transport was microtubule dependent, preferentially anterograde in axons, and 2-fold faster in axons than in dendrites. SV and DCV secretion were strictly Ca(2+) and SNARE dependent. DCV secretion capacity matured until day in vitro (DIV) 36, with intense stimulation releasing 6% of the total DCV pool, and then plateaued. This efficiency is comparable with mature mouse neurons. In contrast, SV secretion capacity continued to increase until DIV50, with substantial further increase in secretion efficiency and decrease in silent synapses. These data show that the two secretory pathways can be studied in human neurons and that they mature differentially, with DCV secretion reaching maximum efficiency when that of SVs is still low.

  4. Selective overproduction of the proteasome inhibitor salinosporamide A via precursor pathway regulation

    PubMed Central

    Lechner, Anna; Eustáquio, Alessandra S.; Gulder, Tobias A. M.; Hafner, Mathias; Moore, Bradley S.

    2011-01-01

    The chlorinated natural product salinosporamide A is a potent 20S proteasome inhibitor currently in clinical trials as an anticancer agent. To deepen our understanding of salinosporamide biosynthesis, we investigated the function of a LuxR-type pathway-specific regulatory gene, salR2, and observed a selective effect on the production of salinosporamide A over its less active aliphatic analogs. SalR2 was shown to specifically activate genes involved in the biosynthesis of the halogenated precursor chloroethylmalonyl-CoA, which is a dedicated precursor of salinosporamide A. Specifically, SalR2 activates transcription of two divergent operons – one of which contains the unique S-adenosyl-L-methionine-dependent chlorinase encoding gene salL. By applying this knowledge towards rational engineering, we were able to selectively double salinosporamide A production. This study exemplifies the specialized regulation of a polyketide precursor pathway and its application to the selective overproduction of a specific natural product congener. PMID:22195555

  5. MT1-MMP proinvasive activity is regulated by a novel Rab8-dependent exocytic pathway.

    PubMed

    Bravo-Cordero, Jose J; Marrero-Diaz, Raquel; Megías, Diego; Genís, Laura; García-Grande, Aranzazu; García, Maria A; Arroyo, Alicia G; Montoya, María C

    2007-03-21

    MT1-matrix metalloproteinase (MT1-MMP) is one of the most critical factors in the invasion machinery of tumor cells. Subcellular localization to invasive structures is key for MT1-MMP proinvasive activity. However, the mechanism driving this polarized distribution remains obscure. We now report that polarized exocytosis of MT1-MMP occurs during MDA-MB-231 adenocarcinoma cell migration into collagen type I three-dimensional matrices. Polarized trafficking of MT1-MMP is triggered by beta1 integrin-mediated adhesion to collagen, and is required for protease localization at invasive structures. Localization of MT1-MMP within VSV-G/Rab8-positive vesicles, but not in Rab11/Tf/TfRc-positive compartment in invasive cells, suggests the involvement of the exocytic traffic pathway. Furthermore, constitutively active Rab8 mutants induce MT1-MMP exocytic traffic, collagen degradation and invasion, whereas Rab8- but not Rab11-knockdown inhibited these processes. Altogether, these data reveal a novel pathway of MT1-MMP redistribution to invasive structures, exocytic vesicle trafficking, which is crucial for its role in tumor cell invasiveness. Mechanistically, MT1-MMP delivery to invasive structures, and therefore its proinvasive activity, is regulated by Rab8 GTPase.

  6. NRF2 Regulates HER2 and HER3 Signaling Pathway to Modulate Sensitivity to Targeted Immunotherapies.

    PubMed

    Khalil, Hilal S; Langdon, Simon P; Kankia, Ibrahim H; Bown, James; Deeni, Yusuf Y

    2016-01-01

    NF-E2 related factor-2 (NRF2) is an essential transcription factor for multiple genes encoding antioxidants and detoxification enzymes. NRF2 is implicated in promoting cancer therapeutic resistance by its detoxification function and crosstalk with proproliferative pathways. However, the exact mechanism of this intricate connectivity between NRF2 and growth factor induced proliferative pathway remains elusive. Here, we have demonstrated that pharmacological activation of NRF2 by tert-butylhydroquinone (tBHQ) upregulates the HER family receptors, HER2 and HER3 expression, elevates pAKT levels, and enhances the proliferation of ovarian cancer cells. Preactivation of NRF2 also attenuates the combined growth inhibitory effects of HER2 targeting monoclonal antibodies, Pertuzumab and Trastuzumab. Further, tBHQ caused transcriptional induction of HER2 and HER3, while SiRNA-mediated knockdown of NRF2 prevented this and further caused transcriptional repression and enhanced cytotoxicity of the HER2 inhibitors. Hence, NRF2 regulates both HER2 and HER3 receptors to influence cellular responses to HER2 targeting monoclonal antibodies. This deciphered crosstalk mechanism reinforces the role of NRF2 in drug resistance and as a relevant anticancer target.

  7. A chemical proteomic atlas of brain serine hydrolases identifies cell type-specific pathways regulating neuroinflammation

    PubMed Central

    Viader, Andreu; Ogasawara, Daisuke; Joslyn, Christopher M; Sanchez-Alavez, Manuel; Mori, Simone; Nguyen, William; Conti, Bruno; Cravatt, Benjamin F

    2016-01-01

    Metabolic specialization among major brain cell types is central to nervous system function and determined in large part by the cellular distribution of enzymes. Serine hydrolases are a diverse enzyme class that plays fundamental roles in CNS metabolism and signaling. Here, we perform an activity-based proteomic analysis of primary mouse neurons, astrocytes, and microglia to furnish a global portrait of the cellular anatomy of serine hydrolases in the brain. We uncover compelling evidence for the cellular compartmentalization of key chemical transmission pathways, including the functional segregation of endocannabinoid (eCB) biosynthetic enzymes diacylglycerol lipase-alpha (DAGLα) and –beta (DAGLβ) to neurons and microglia, respectively. Disruption of DAGLβ perturbed eCB-eicosanoid crosstalk specifically in microglia and suppressed neuroinflammatory events in vivo independently of broader effects on eCB content. Mapping the cellular distribution of metabolic enzymes thus identifies pathways for regulating specialized inflammatory responses in the brain while avoiding global alterations in CNS function. DOI: http://dx.doi.org/10.7554/eLife.12345.001 PMID:26779719

  8. Two anthranilate synthase genes in Arabidopsis: defense-related regulation of the tryptophan pathway.

    PubMed Central

    Niyogi, K K; Fink, G R

    1992-01-01

    Arabidopsis thaliana has two genes, ASA1 and ASA2, encoding the alpha subunit of anthranilate synthase, the enzyme catalyzing the first reaction in the tryptophan biosynthetic pathway. As a branchpoint enzyme in aromatic amino acid biosynthesis, anthranilate synthase has an important regulatory role. The sequences of the plant genes are homologous to their microbial counterparts. Both predicted proteins have putative chloroplast transit peptides at their amino termini and conserved amino acids involved in feedback inhibition by tryptophan. ASA1 and ASA2 cDNAs complement anthranilate synthase alpha subunit mutations in the yeast Saccharomyces cerevisiae and in Escherichia coli, confirming that both genes encode functional anthranilate synthase proteins. The distributions of ASA1 and ASA2 mRNAs in various parts of Arabidopsis plants are overlapping but nonidentical, and ASA1 mRNA is approximately 10 times more abundant in whole plants. Whereas ASA2 is expressed at a constitutive basal level, ASA1 is induced by wounding and bacterial pathogen infiltration, suggesting a novel role for ASA1 in the production of tryptophan pathway metabolites as part of an Arabidopsis defense response. Regulation of key steps in aromatic amino acid biosynthesis in Arabidopsis appears to involve differential expression of duplicated genes. PMID:1392592

  9. Ndel1 suppresses ciliogenesis in proliferating cells by regulating the trichoplein-Aurora A pathway.

    PubMed

    Inaba, Hironori; Goto, Hidemasa; Kasahara, Kousuke; Kumamoto, Kanako; Yonemura, Shigenobu; Inoko, Akihito; Yamano, Shotaro; Wanibuchi, Hideki; He, Dongwei; Goshima, Naoki; Kiyono, Tohru; Hirotsune, Shinji; Inagaki, Masaki

    2016-02-15

    Primary cilia protrude from the surface of quiescent cells and disassemble at cell cycle reentry. We previously showed that ciliary reassembly is suppressed by trichoplein-mediated Aurora A activation pathway in growing cells. Here, we report that Ndel1, a well-known modulator of dynein activity, localizes at the subdistal appendage of the mother centriole, which nucleates a primary cilium. In the presence of serum, Ndel1 depletion reduces trichoplein at the mother centriole and induces unscheduled primary cilia formation, which is reverted by forced trichoplein expression or coknockdown of KCTD17 (an E3 ligase component protein for trichoplein). Serum starvation induced transient Ndel1 degradation, subsequent to the disappearance of trichoplein at the mother centriole. Forced expression of Ndel1 suppressed trichoplein degradation and axonemal microtubule extension during ciliogenesis, similar to trichoplein induction or KCTD17 knockdown. Most importantly, the proportion of ciliated and quiescent cells was increased in the kidney tubular epithelia of newborn Ndel1-hypomorphic mice. Thus, Ndel1 acts as a novel upstream regulator of the trichoplein-Aurora A pathway to inhibit primary cilia assembly.

  10. Engineering of the glycerol decomposition pathway and cofactor regulation in an industrial yeast improves ethanol production.

    PubMed

    Zhang, Liang; Tang, Yan; Guo, Zhongpeng; Shi, Guiyang

    2013-10-01

    Glycerol is a major by-product of industrial ethanol production and its formation consumes up to 4 % of the sugar substrate. This study modified the glycerol decomposition pathway of an industrial strain of Saccharomyces cerevisiae to optimize the consumption of substrate and yield of ethanol. This study is the first to couple glycerol degradation with ethanol formation, to the best of our knowledge. The recombinant strain overexpressing GCY1 and DAK1, encoding glycerol dehydrogenase and dihydroxyacetone kinase, respectively, in glycerol degradation pathway, exhibited a moderate increase in ethanol yield (2.9 %) and decrease in glycerol yield (24.9 %) compared to the wild type with the initial glucose concentration of 15 % under anaerobic conditions. However, when the mhpF gene, encoding acetylating NAD⁺-dependent acetaldehyde dehydrogenase from Escherichia coli, was co-expressed in the aforementioned recombinant strain, a further increase in ethanol yield by 5.5 % and decrease in glycerol yield by 48 % were observed for the resultant recombinant strain GDMS1 when acetic acid was added into the medium prior to inoculation compared to the wild type. The process outlined in this study which enhances glycerol consumption and cofactor regulation in an industrial yeast is a promising metabolic engineering strategy to increase ethanol production by reducing the formation of glycerol.

  11. Undaria pinnatifida soluble fiber regulates Angptl3-LPL pathway to lessen hyperlipidemia in mice.

    PubMed

    Xiao, Hong-Bo; Lu, Xiang-Yang; Zhang, Heng-Bo; Sun, Zhi-Liang; Fang, Jun

    2013-12-01

    Angiopoietin-like protein 3 (Angptl3)-lipoprotein lipase (LPL) pathway may be a useful pharmacologic target for hyperlipidemia. The present study was conducted to test the effect of soluble fiber extracted from Undaria pinnatifida (UP), on hyperlipidemia in apolipoprotein E-deficient (ApoE(-/-)) mice. Forty mice were divided into four groups (n = 10): control group (C57BL/6J mice), ApoE(-/-) mice group, and two groups of ApoE(-/-) mice treated with UP fiber (5 or 10 % per day). UP soluble fiber treatment significantly decreased plasma and hepatic total cholesterol, triglycerides levels, plasma low-density lipoprotein cholesterol, and malondialdehyde concentrations and increased plasma high-density lipoprotein cholesterol level and downregulated protein expression of Angptl3 concomitantly with upregulated protein expression of LPL. In addition, T0901317 caused elevated expression of hepatic Angptl3 protein, and the effect of T0901317 was also abrogated by UP soluble fiber in C57BL/6J mice. The present results suggest that the UP soluble fiber regulates Angptl3-LPL pathway to lessen hyperlipidemia in mice.

  12. Role of peroxynitrite in the redox regulation of cell signal transduction pathways

    PubMed Central

    Liaudet, Lucas; Vassalli, Giuseppe; Pacher, Pal

    2009-01-01

    Peroxynitrite is a potent oxidant and nitrating species formed from the reaction between the free radicals nitric oxide and superoxide. An excessive formation of peroxynitrite represents an important mechanism contributing to cell death and dysfunction in multiple cardiovascular pathologies, such as myocardial infarction, heart failure and atherosclerosis. Whereas initial works focused on direct oxidative biomolecular damage as the main route of peroxynitrite toxicity, more recent evidence, mainly obtained in vitro, indicates that peroxynitrite also behaves as a potent modulator of various cell signal transduction pathways. Due to its ability to nitrate tyrosine residues, peroxynitrite affects cellular processes dependent on tyrosine phosphorylation. Peroxynitrite also exerts complex effects on the activity of various kinases and phosphatases, resulting in the up- or downregulation of signalling cascades, in a concentration- and cell-dependent manner. Such roles of peroxynitrite in the redox regulation of key signalling pathways for cardiovascular homeostasis, including protein kinase B and C, the MAP kinases, Nuclear Factor Kappa B, as well as signalling dependent on insulin and the sympatho-adrenergic system are presented in detail in this review. PMID:19273391

  13. Bmi1 regulates mitochondrial function and the DNA damage response pathway.

    PubMed

    Liu, Jie; Cao, Liu; Chen, Jichun; Song, Shiwei; Lee, In Hye; Quijano, Celia; Liu, Hongjun; Keyvanfar, Keyvan; Chen, Haoqian; Cao, Long-Yue; Ahn, Bong-Hyun; Kumar, Neil G; Rovira, Ilsa I; Xu, Xiao-Ling; van Lohuizen, Maarten; Motoyama, Noboru; Deng, Chu-Xia; Finkel, Toren

    2009-05-21

    Mice deficient in the Polycomb repressor Bmi1 develop numerous abnormalities including a severe defect in stem cell self-renewal, alterations in thymocyte maturation and a shortened lifespan. Previous work has implicated de-repression of the Ink4a/Arf (also known as Cdkn2a) locus as mediating many of the aspects of the Bmi1(-/-) phenotype. Here we demonstrate that cells derived from Bmi1(-/-) mice also have impaired mitochondrial function, a marked increase in the intracellular levels of reactive oxygen species and subsequent engagement of the DNA damage response pathway. Furthermore, many of the deficiencies normally observed in Bmi1(-/-) mice improve after either pharmacological treatment with the antioxidant N-acetylcysteine or genetic disruption of the DNA damage response pathway by Chk2 (also known as Chek2) deletion. These results demonstrate that Bmi1 has an unexpected role in maintaining mitochondrial function and redox homeostasis and indicate that the Polycomb family of proteins can coordinately regulate cellular metabolism with stem and progenitor cell function.

  14. Berberine regulates AMP-activated protein kinase signaling pathways and inhibits colon tumorigenesis in mice.

    PubMed

    Li, Weidong; Hua, Baojin; Saud, Shakir M; Lin, Hongsheng; Hou, Wei; Matter, Matthias S; Jia, Libin; Colburn, Nancy H; Young, Matthew R

    2015-10-01

    Colorectal cancer, a leading cause of cancer death, has been linked to inflammation and obesity. Berberine, an isoquinoline alkaloid, possesses anti-inflammatory, anti-diabetes and anti-tumor properties. In the azoxymethane initiated and dextran sulfate sodium (AOM/DSS) promoted colorectal carcinogenesis mouse model, berberine treated mice showed a 60% reduction in tumor number (P = 0.009), a 48% reduction in tumors <2 mm, (P = 0.05); 94% reduction in tumors 2-4 mm, (P = 0.001), and 100% reduction in tumors >4 mm (P = 0.02) compared to vehicle treated mice. Berberine also decreased AOM/DSS induced Ki-67 and COX-2 expression. In vitro analysis showed that in addition to its anti-proliferation activity, berberine also induced apoptosis in colorectal cancer cell lines. Berberine activated AMP-activated protein kinase (AMPK), a major regulator of metabolic pathways, and inhibited mammalian target of rapamycin (mTOR), a downstream target of AMPK. Furthermore, 4E-binding protein-1 and p70 ribosomal S6 kinases, downstream targets of mTOR, were down regulated by berberine treatment. Berberine did not affect Liver kinase B1 (LKB1) activity or the mitogen-activated protein kinase pathway. Berberine inhibited Nuclear Factor kappa-B (NF-κB) activity, reduced the expression of cyclin D1 and survivin, induced phosphorylation of p53 and increased caspase-3 cleavage in vitro. Berberine inhibition of mTOR activity and p53 phosphorylation was found to be AMPK dependent, while inhibition NF-κB was AMPK independent. In vivo, berberine also activated AMPK, inhibited mTOR and p65 phosphorylation and activated caspase-3 cleavage. Our data suggests that berberine suppresses colon epithelial proliferation and tumorigenesis via AMPK dependent inhibition of mTOR activity and AMPK independent inhibition of NF-κB.

  15. The Akt1/IL-6/STAT3 pathway regulates growth of lung tumor initiating cells

    PubMed Central

    Malanga, Donatella; De Marco, Carmela; Guerriero, Ilaria; Colelli, Fabiana; Rinaldo, Nicola; Scrima, Marianna; Mirante, Teresa; De Vitis, Claudia; Zoppoli, Pietro; Ceccarelli, Michele; Riccardi, Miriam; Ravo, Maria; Weisz, Alessandro; Federico, Antonella; Franco, Renato; Rocco, Gaetano; Mancini, Rita; Rizzuto, Antonia; Gulletta, Elio; Ciliberto, Gennaro; Viglietto, Giuseppe

    2015-01-01

    Here we report that the PI3K/Akt1/IL-6/STAT3 signalling pathway regulates generation and stem cell-like properties of Non-Small Cell Lung Cancer (NSCLC) tumor initiating cells (TICs). Mutant Akt1, mutant PIK3CA or PTEN loss enhances formation of lung cancer spheroids (LCS), self-renewal, expression of stemness markers and tumorigenic potential of human immortalized bronchial cells (BEAS-2B) whereas Akt inhibition suppresses these activities in established (NCI-H460) and primary NSCLC cells. Matched microarray analysis of Akt1-interfered cells and LCSs identified IL-6 as a critical target of Akt signalling in NSCLC TICs. Accordingly, suppression of Akt in NSCLC cells decreases IL-6 levels, phosphorylation of IkK and IkB, NF-kB transcriptional activity, phosphorylation and transcriptional activity of STAT3 whereas active Akt1 up-regulates them. Exposure of LCSs isolated from NSCLC cells to blocking anti-IL-6 mAbs, shRNA to IL-6 receptor or to STAT3 markedly reduces the capability to generate LCSs, to self-renew and to form tumors, whereas administration of IL-6 to Akt-interfered cells restores the capability to generate LCSs. Finally, immunohistochemical studies in NSCLC patients demonstrated a positive correlative trend between activated Akt, IL-6 expression and STAT3 phosphorylation (n = 94; p < 0.05). In conclusion, our data indicate that aberrant Akt signalling contributes to maintaining stemness in lung cancer TICs through a NF-kB/IL-6/STAT3 pathway and provide novel potential therapeutic targets for eliminating these malignant cells in NSCLC. PMID:26486080

  16. ADI pathway and histidine decarboxylation are reciprocally regulated in Lactobacillus hilgardii ISE 5211: proteomic evidence.

    PubMed

    Lamberti, Cristina; Purrotti, Micol; Mazzoli, Roberto; Fattori, Paolo; Barello, Cristina; Coïsson, Jean Daniel; Giunta, Carlo; Pessione, Enrica

    2011-07-01

    Amine production by amino acid decarboxylation is a common feature that is used by lactic acid bacteria (LAB) to complement lactic fermentation, since it is coupled with a proton-extruding antiport system which leads to both metabolic energy production and the attenuation of intracellular acidity. Analogous roles are played in LAB by both malolactic fermentation (MLF) and the arginine deiminase (ADI) pathway. The present investigation was aimed at establishing reciprocal interactions between amino acid decarboxylation and the two above mentioned routes. The analyses were carried out on a Lactobacillus hilgardii strain (ISE 5211) that is able to decarboxylate histidine to histamine, which had previously been isolated from wine and whose complete genome is still unknown. The 2DE proteomic approach, followed by MALDI TOF-TOF and De Novo Sequencing, was used to study the protein expression levels. The experimental evidence has indicated that malate does not influence histidine decarboxylase (HDC) biosynthesis and that histidine does not affect the malolactic enzyme level. However, the expression of the ADI route enzymes, arginine deiminase and ornithine transcarbamylase, is down-regulated by histidine: this biosynthetic repression is more important (4-fold) in cultures that are not supplemented with arginine, but is also significant (2-fold) in an arginine supplemented medium that normally induces the ADI pathway. On the other hand, arginine partially represses HDC expression, but only when histidine and arginine are both present in the culture medium. This proteomic study has also pointed out a down-regulation exerted by histidine over sugar metabolism enzymes and a GroEL stress protein. These data, together with the reciprocal antagonism between arginine deimination and histidine decarboxylation, offer clue keys to the understanding of the accumulation of lactate, amine, ammonia and ethylcarbamate in wine, with consequent implications on different health risk

  17. Differential regulation of the Hippo pathway by adherens junctions and apical-basal cell polarity modules.

    PubMed

    Yang, Chih-Chao; Graves, Hillary K; Moya, Ivan M; Tao, Chunyao; Hamaratoglu, Fisun; Gladden, Andrew B; Halder, Georg

    2015-02-10

    Adherens junctions (AJs) and cell polarity complexes are key players in the establishment and maintenance of apical-basal cell polarity. Loss of AJs or basolateral polarity components promotes tumor formation and metastasis. Recent studies in vertebrate models show that loss of AJs or loss of the basolateral component Scribble (Scrib) cause deregulation of the Hippo tumor suppressor pathway and hyperactivation of its downstream effectors Yes-associated protein (YAP) and Transcriptional coactivator with PDZ-binding motif (TAZ). However, whether AJs and Scrib act through the same or independent mechanisms to regulate Hippo pathway activity is not known. Here, we dissect how disruption of AJs or loss of basolateral components affect the activity of the Drosophila YAP homolog Yorkie (Yki) during imaginal disc development. Surprisingly, disruption of AJs and loss of basolateral proteins produced very different effects on Yki activity. Yki activity was cell-autonomously decreased but non-cell-autonomously elevated in tissues where the AJ components E-cadherin (E-cad) or α-catenin (α-cat) were knocked down. In contrast, scrib knockdown caused a predominantly cell-autonomous activation of Yki. Moreover, disruption of AJs or basolateral proteins had different effects on cell polarity and tissue size. Simultaneous knockdown of α-cat and scrib induced both cell-autonomous and non-cell-autonomous Yki activity. In mammalian cells, knockdown of E-cad or α-cat caused nuclear accumulation and activation of YAP without overt effects on Scrib localization and vice versa. Therefore, our results indicate the existence of multiple, genetically separable inputs from AJs and cell polarity complexes into Yki/YAP regulation.

  18. Single GDP-dissociation Inhibitor Protein regulates endocytic and secretory pathways in Leishmania

    PubMed Central

    Shanmugam, Senthil kumar; Kumar, Kamal; Singh, Pawan Kishor; Rastogi, Ruchir; Mukhopadhyay, Amitabha

    2016-01-01

    The role of GDP dissociation inhibitor (GDI) protein in regulation of Rab cycle in Leishmania is not known. Here, we have cloned and characterized the functions of GDI homologue in vivo in Leishmania. Our results have shown that LdGDI:WT along with GDP removes the Rab5 from purified endosomes and inhibits the homotypic fusion between early endosomes. Whereas, LdGDI:R239A, a dominant negative mutant of GDI, under the same condition neither removes the Rab5 from endosome nor inhibits fusion. To determine the role of Ld-GDI in vivo, transgenic parasites overexpressing GFP-LdGDI:WT or GFP-LdGDI:R239A, are co-expressed with RFP-LdRab5:WT, RFP-LdRab7:WT or RFP-LdRab1:WT. Our results have shown that overexpression of GFP-LdGDI:WT extracts the RFP-LdRab5, RFP-LdRab7 or RFP-LdRab1 from their discrete endomembrane predominantly into cytosol. No change in the distribution of indicated Rabs is detected with overexpression of GFP-LdGDI:R239A. To determine the functional significance, we have used hemoglobin as an endocytic marker and gp63 as a marker for secretory pathway. We have found that overexpression of GFP-LdGDI:WT enhances the lysosomal targeting of internalized hemoglobin and the secretion of gp63 in the parasites possibly by triggering Rab cycle. This is the first demonstration of a single GDI ubiquitously regulating both endocytic and secretory pathways in Leishmania. PMID:27841328

  19. The Akt1/IL-6/STAT3 pathway regulates growth of lung tumor initiating cells.

    PubMed

    Malanga, Donatella; De Marco, Carmela; Guerriero, Ilaria; Colelli, Fabiana; Rinaldo, Nicola; Scrima, Marianna; Mirante, Teresa; De Vitis, Claudia; Zoppoli, Pietro; Ceccarelli, Michele; Riccardi, Miriam; Ravo, Maria; Weisz, Alessandro; Federico, Antonella; Franco, Renato; Rocco, Gaetano; Mancini, Rita; Rizzuto, Antonia; Gulletta, Elio; Ciliberto, Gennaro; Viglietto, Giuseppe

    2015-12-15

    Here we report that the PI3K/Akt1/IL-6/STAT3 signalling pathway regulates generation and stem cell-like properties of Non-Small Cell Lung Cancer (NSCLC) tumor initiating cells (TICs). Mutant Akt1, mutant PIK3CA or PTEN loss enhances formation of lung cancer spheroids (LCS), self-renewal, expression of stemness markers and tumorigenic potential of human immortalized bronchial cells (BEAS-2B) whereas Akt inhibition suppresses these activities in established (NCI-H460) and primary NSCLC cells. Matched microarray analysis of Akt1-interfered cells and LCSs identified IL-6 as a critical target of Akt signalling in NSCLC TICs. Accordingly, suppression of Akt in NSCLC cells decreases IL-6 levels, phosphorylation of IkK and IkB, NF-kB transcriptional activity, phosphorylation and transcriptional activity of STAT3 whereas active Akt1 up-regulates them. Exposure of LCSs isolated from NSCLC cells to blocking anti-IL-6 mAbs, shRNA to IL-6 receptor or to STAT3 markedly reduces the capability to generate LCSs, to self-renew and to form tumors, whereas administration of IL-6 to Akt-interfered cells restores the capability to generate LCSs. Finally, immunohistochemical studies in NSCLC patients demonstrated a positive correlative trend between activated Akt, IL-6 expression and STAT3 phosphorylation (n = 94; p < 0.05). In conclusion, our data indicate that aberrant Akt signalling contributes to maintaining stemness in lung cancer TICs through a NF-kB/IL-6/STAT3 pathway and provide novel potential therapeutic targets for eliminating these malignant cells in NSCLC.

  20. Extending the translational potential of targeting NO/cGMP-regulated pathways in the CVS

    PubMed Central

    Papapetropoulos, Andreas; Hobbs, Adrian J; Topouzis, Stavros

    2015-01-01

    The discovery of NO as both an endogenous signalling molecule and as a mediator of the cardiovascular effects of organic nitrates was acknowledged in 1998 by the Nobel Prize in Physiology/Medicine. The characterization of its downstream signalling, mediated through stimulation of soluble GC (sGC) and cGMP generation, initiated significant translational interest, but until recently this was almost exclusively embodied by the use of PDE5 inhibitors in erectile dysfunction. Since then, research progress in two areas has contributed to an impressive expansion of the therapeutic targeting of the NO-sGC-cGMP axis: first, an increased understanding of the molecular events operating within this complex pathway and second, a better insight into its dys-regulation and uncoupling in human disease. Already-approved PDE5 inhibitors and novel, first-in-class molecules, which up-regulate the activity of sGC independently of NO and/or of the enzyme's haem prosthetic group, are undergoing clinical evaluation to treat pulmonary hypertension and myocardial failure. These molecules, as well as combinations or second-generation compounds, are also being assessed in additional experimental disease models and in patients in a wide spectrum of novel indications, such as endotoxic shock, diabetic cardiomyopathy and Becker's muscular dystrophy. There is well-founded optimism that the modulation of the NO-sGC-cGMP pathway will sustain the development of an increasing number of successful clinical candidates for years to come. Linked Articles This article is part of a themed section on Pharmacology of the Gasotransmitters. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue–6 PMID:25302549

  1. Endogenous hydrogen sulfide regulates inflammatory response by activating the ERK pathway in polymicrobial sepsis.

    PubMed

    Zhang, Huili; Moochhala, Shabbir M; Bhatia, Madhav

    2008-09-15

    Hydrogen sulfide (H(2)S) up-regulates inflammatory response in several inflammatory diseases. However, to date, little is known about the molecular mechanism by which H(2)S provokes the inflammatory response in sepsis. Thus, the aim of this study was to investigate the signaling pathway underlying the proinflammatory role of H(2)S in cecal ligation and puncture (CLP)-induced sepsis. Male Swiss mice were subjected to CLP and treated with dl-propargylglycine (PAG; 50 mg/kg i.p., an inhibitor of H(2)S formation), NaHS (10 mg/kg, i.p., an H(2)S donor), or saline. PAG was administered 1 h before CLP, whereas NaHS was given at the time of CLP. CLP-induced sepsis resulted in a time-dependent increase in the synthesis of endogenous H(2)S. Maximum phosphorylation of ERK1/2 and degradation of IkappaBalpha in lung and liver were observed 4 h after CLP. Inhibition of H(2)S formation by PAG significantly reduced the phosphorylation of ERK1/2 in lung and liver 4 h after CLP, coupled with decreased degradation of IkappaBalpha and activation of NF-kappaB. In contrast, injection of NaHS significantly enhanced the activation of ERK1/2 in lung and liver, therefore leading to a further rise in tissue NF-kappaB activity. As a result, pretreatment with PAG significantly reduced the production of cytokines and chemokines in sepsis, whereas exogenous H(2)S greatly increased it. In addition, pretreatment with PD98059, an inhibitor of ERK kinase (MEK-1), significantly prevented NaHS from aggravating systemic inflammation in sepsis. In conclusion, the present study shows for the first time that H(2)S may regulate systemic inflammatory response in sepsis via ERK pathway.

  2. Parathyroid hormone-dependent signaling pathways regulating genes in bone cells

    NASA Technical Reports Server (NTRS)

    Swarthout, John T.; D'Alonzo, Richard C.; Selvamurugan, Nagarajan; Partridge, Nicola C.

    2002-01-01

    Parathyroid hormone (PTH) is an 84-amino-acid polypeptide hormone functioning as a major mediator of bone remodeling and as an essential regulator of calcium homeostasis. PTH and PTH-related protein (PTHrP) indirectly activate osteoclasts resulting in increased bone resorption. During this process, PTH changes the phenotype of the osteoblast from a cell involved in bone formation to one directing bone resorption. In addition to these catabolic effects, PTH has been demonstrated to be an anabolic factor in skeletal tissue and in vitro. As a result, PTH has potential medical application to the treatment of osteoporosis, since intermittent administration of PTH stimulates bone formation. Activation of osteoblasts by PTH results in expression of genes important for the degradation of the extracellular matrix, production of growth factors, and stimulation and recruitment of osteoclasts. The ability of PTH to drive changes in gene expression is dependent upon activation of transcription factors such as the activator protein-1 family, RUNX2, and cAMP response element binding protein (CREB). Much of the regulation of these processes by PTH is protein kinase A (PKA)-dependent. However, while PKA is linked to many of the changes in gene expression directed by PTH, PKA activation has been shown to inhibit mitogen-activated protein kinase (MAPK) and proliferation of osteoblasts. It is now known that stimulation of MAPK and proliferation by PTH at low concentrations is protein kinase C (PKC)-dependent in both osteoblastic and kidney cells. Furthermore, PTH has been demonstrated to regulate components of the cell cycle. However, whether this regulation requires PKC and/or extracellular signal-regulated kinases or whether PTH is able to stimulate other components of the cell cycle is unknown. It is possible that stimulation of this signaling pathway by PTH mediates a unique pattern of gene expression resulting in proliferation in osteoblastic and kidney cells; however, specific

  3. Rubicon and PLEKHM1 negatively regulate the endocytic/autophagic pathway via a novel Rab7-binding domain.

    PubMed

    Tabata, Keisuke; Matsunaga, Kohichi; Sakane, Ayuko; Sasaki, Takuya; Noda, Takeshi; Yoshimori, Tamotsu

    2010-12-01

    The endocytic and autophagic pathways are involved in the membrane trafficking of exogenous and endogenous materials to lysosomes. However, the mechanisms that regulate these pathways are largely unknown. We previously reported that Rubicon, a Beclin 1-binding protein, negatively regulates both the autophagic and endocytic pathways by unidentified mechanisms. In this study, we performed database searches to identify potential Rubicon homologues that share the common C-terminal domain, termed the RH domain. One of them, PLEKHM1, the causative gene of osteopetrosis, also suppresses endocytic transport but not autophagosome maturation. Rubicon and PLEKHM1 specifically and directly interact with Rab7 via their RH domain, and this interaction is critical for their function. Furthermore, we show that Rubicon but not PLEKHM1 uniquely regulates membrane trafficking via simultaneously binding both Rab7 and PI3-kinase.

  4. Regulating ehrlich and demethiolation pathways for alcohols production by the expression of ubiquitin-protein ligase gene HUWE1

    PubMed Central

    Zhang, Quan; Jia, Kai-Zhi; Xia, Shi-Tao; Xu, Yang-Hua; Liu, Rui-Sang; Li, Hong-Mei; Tang, Ya-Jie

    2016-01-01

    Ehrlich and demethiolation pathways as two competing branches converted amino acid into alcohols. Controlling both pathways offers considerable potential for industrial applications including alcohols overproduction, flavor-quality control and developing new flavors. While how to regulate ehrlich and demethiolation pathways is still not applicable. Taking the conversion of methionine into methionol and methanethiol for example, we constructed two suppression subtractive cDNA libraries of Clonostachys rosea by using suppression subtractive hybridization (SSH) technology for screening regulators controlling the conversion. E3 ubiquitin-protein ligase gene HUWE1 screened from forward SSH library was validated to be related with the biosynthesis of end products. Overexpressing HUWE1 in C. rosea and S. cerevisiae significantly increased the biosynthesis of methanethiol and its derivatives in demethiolation pathway, while suppressed the biosynthesis of methional and methionol in ehrlich pathway. These results attained the directional regulation of both pathways by overexpressing HUWE1. Thus, HUWE1 has potential to be a key target for controlling and enhancing alcohols production by metabolic engineering. PMID:26860895

  5. Protein Analysis of Sapienic Acid-Treated Porphyromonas gingivalis Suggests Differential Regulation of Multiple Metabolic Pathways

    PubMed Central

    Dawson, Deborah V.; Blanchette, Derek R.; Drake, David R.; Wertz, Philip W.; Brogden, Kim A.

    2015-01-01

    ABSTRACT Lipids endogenous to skin and mucosal surfaces exhibit potent antimicrobial activity against Porphyromonas gingivalis, an important colonizer of the oral cavity implicated in periodontitis. Our previous work demonstrated the antimicrobial activity of the fatty acid sapienic acid (C16:1Δ6) against P. gingivalis and found that sapienic acid treatment alters both protein and lipid composition from those in controls. In this study, we further examined whole-cell protein differences between sapienic acid-treated bacteria and untreated controls, and we utilized open-source functional association and annotation programs to explore potential mechanisms for the antimicrobial activity of sapienic acid. Our analyses indicated that sapienic acid treatment induces a unique stress response in P. gingivalis resulting in differential expression of proteins involved in a variety of metabolic pathways. This network of differentially regulated proteins was enriched in protein-protein interactions (P = 2.98 × 10−8), including six KEGG pathways (P value ranges, 2.30 × 10−5 to 0.05) and four Gene Ontology (GO) molecular functions (P value ranges, 0.02 to 0.04), with multiple suggestive enriched relationships in KEGG pathways and GO molecular functions. Upregulated metabolic pathways suggest increases in energy production, lipid metabolism, iron acquisition and processing, and respiration. Combined with a suggested preferential metabolism of serine, which is necessary for fatty acid biosynthesis, these data support our previous findings that the site of sapienic acid antimicrobial activity is likely at the bacterial membrane. IMPORTANCE P. gingivalis is an important opportunistic pathogen implicated in periodontitis. Affecting nearly 50% of the population, periodontitis is treatable, but the resulting damage is irreversible and eventually progresses to tooth loss. There is a great need for natural products that can be used to treat and/or prevent the overgrowth of

  6. Spatial landmarks regulate a Cdc42-dependent MAPK pathway to control differentiation and the response to positional compromise

    PubMed Central

    Basu, Sukanya; Vadaie, Nadia; Prabhakar, Aditi; Li, Boyang; Adhikari, Hema; Pitoniak, Andrew; Chow, Jacky; Chavel, Colin A.; Cullen, Paul J.

    2016-01-01

    A fundamental problem in cell biology is to understand how spatial information is recognized and integrated into morphogenetic responses. Budding yeast undergoes differentiation to filamentous growth, which involves changes in cell polarity through mechanisms that remain obscure. Here we define a regulatory input where spatial landmarks (bud-site–selection proteins) regulate the MAPK pathway that controls filamentous growth (fMAPK pathway). The bud-site GTPase Rsr1p regulated the fMAPK pathway through Cdc24p, the guanine nucleotide exchange factor for the polarity establishment GTPase Cdc42p. Positional landmarks that direct Rsr1p to bud sites conditionally regulated the fMAPK pathway, corresponding to their roles in regulating bud-site selection. Therefore, cell differentiation is achieved in part by the reorganization of polarity at bud sites. In line with this conclusion, dynamic changes in budding pattern during filamentous growth induced corresponding changes in fMAPK activity. Intrinsic compromise of bud-site selection also impacted fMAPK activity. Therefore, a surveillance mechanism monitors spatial position in response to extrinsic and intrinsic stress and modulates the response through a differentiation MAPK pathway. PMID:27001830

  7. Chemical genetics reveals negative regulation of abscisic acid signaling by a plant immune response pathway.

    PubMed

    Kim, Tae-Houn; Hauser, Felix; Ha, Tracy; Xue, Shaowu; Böhmer, Maik; Nishimura, Noriyuki; Munemasa, Shintaro; Hubbard, Katharine; Peine, Nora; Lee, Byeong-Ha; Lee, Stephen; Robert, Nadia; Parker, Jane E; Schroeder, Julian I

    2011-06-07

    Coordinated regulation of protection mechanisms against environmental abiotic stress and pathogen attack is essential for plant adaptation and survival. Initial abiotic stress can interfere with disease-resistance signaling [1-6]. Conversely, initial plant immune signaling may interrupt subsequent abscisic acid (ABA) signal transduction [7, 8]. However, the processes involved in this crosstalk between these signaling networks have not been determined. By screening a 9600-compound chemical library, we identified a small molecule [5-(3,4-dichlorophenyl)furan-2-yl]-piperidine-1-ylmethanethione (DFPM) that rapidly downregulates ABA-dependent gene expression and also inhibits ABA-induced stomatal closure. Transcriptome analyses show that DFPM also stimulates expression of plant defense-related genes. Major early regulators of pathogen-resistance responses, including EDS1, PAD4, RAR1, and SGT1b, are required for DFPM-and notably also for Pseudomonas-interference with ABA signal transduction, whereas salicylic acid, EDS16, and NPR1 are not necessary. Although DFPM does not interfere with early ABA perception by PYR/RCAR receptors or ABA activation of SnRK2 kinases, it disrupts cytosolic Ca(2+) signaling and downstream anion channel activation in a PAD4-dependent manner. Our findings provide evidence that activation of EDS1/PAD4-dependent plant immune responses rapidly disrupts ABA signal transduction and that this occurs at the level of Ca(2+) signaling, illuminating how the initial biotic stress pathway interferes with ABA signaling.

  8. Piperine regulates UCP1 through the AMPK pathway by generating intracellular lactate production in muscle cells

    PubMed Central

    Kim, Nami; Nam, Miso; Kang, Mi Sun; Lee, Jung Ok; Lee, Yong Woo; Hwang, Geum-Sook; Kim, Hyeon Soo

    2017-01-01

    This study characterizes the human metabolic response to piperine, a curcumin extract, and the details of its underlying molecular mechanism. Using 1H-NMR-based metabolome analysis, we showed the metabolic effect of piperine on skeletal muscle and found that piperine increased the level of intracellular lactate, an important metabolic intermediate that controls expression of several genes involved in mitochondrial activity. Piperine also induced the phosphorylation of AMP-activated protein kinase (AMPK) and its downstream target, acetyl-CoA carboxylase (ACC), while additionally stimulating glucose uptake in an AMPK dependent manner. Piperine also stimulates the p38 mitogen-activated protein kinase (p38 MAPK), an effect that was reversed by pretreatment with compound C, an AMPK inhibitor. Inhibition of p38 MAPK resulted in no piperine-induced glucose uptake. Increased level of lactate resulted in increased expression of mitochondrial uncoupling protein 1 (UCP1), which regulates energy expenditure, thermogenesis, and fat browning. Knock-down of AMPK blocked piperine-induced UCP1 up-regulation, demonstrating the required role of AMPK in this effect. Taken together, these results suggest that piperine leads to benign metabolic effects by activating the AMPK-p38 MAPK signaling pathway and UCP1 expression by activating intracellular lactate production in skeletal muscle. PMID:28117414

  9. Prostaglandin E2 negatively regulates AMP-activated protein kinase via protein kinase A signaling pathway.

    PubMed

    Funahashi, Koji; Cao, Xia; Yamauchi, Masako; Kozaki, Yasuko; Ishiguro, Naoki; Kambe, Fukushi

    2009-01-01

    We investigated possible involvement of prostaglandin (PG) E2 in regulation of AMP-activated protein kinase (AMPK). When osteoblastic MG63 cells were cultured in serum-deprived media, Thr-172 phosphorylation of AMPK alpha-subunit was markedly increased. Treatment of the cells with PGE2 significantly reduced the phosphorylation. Ser-79 phosphorylation of acetyl-CoA carboxylase, a direct target for AMPK, was also reduced by PGE2. On the other hand, PGE2 reciprocally increased Ser-485 phosphorylation of the alpha-subunit that could be associated with inhibition of AMPK activity. These effects of PGE2 were mimicked by PGE2 receptor EP2 and EP4 agonists and forskolin, but not by EP1 and EP3 agonists, and the effects were suppressed by an adenylate cyclase inhibitor SQ22536 and a protein kinase A inhibitor H89. Additionally, the PGE2 effects were duplicated in primary calvarial osteoblasts. Together, the present study demonstrates that PGE2 negatively regulates AMPK activity via activation of protein kinase A signaling pathway.

  10. Spatial Propagation and Localization of Blood Coagulation Are Regulated by Intrinsic and Protein C Pathways, Respectively

    PubMed Central

    Panteleev, Mikhail A.; Ovanesov, Mikhail V.; Kireev, Dmitrii A.; Shibeko, Aleksei M.; Sinauridze, Elena I.; Ananyeva, Natalya M.; Butylin, Andrey A.; Saenko, Evgueni L.; Ataullakhanov, Fazoil I.

    2006-01-01

    Blood coagulation in vivo is a spatially nonuniform, multistage process: coagulation factors from plasma bind to tissue factor (TF)-expressing cells, become activated, dissociate, and diffuse into plasma to form enzymatic complexes on the membranes of activated platelets. We studied spatial regulation of coagulation using two approaches: 1), an in vitro experimental model of clot formation in a thin layer of plasma activated by a monolayer of TF-expressing cells; and 2), a computer simulation model. Clotting in factor VIII- and factor XI-deficient plasmas was initiated normally, but further clot elongation was impaired in factor VIII- and, at later stages, in factor XI-deficient plasma. The data indicated that clot elongation was regulated by factor Xa formation by intrinsic tenase, whereas factor IXa was formed by extrinsic tenase on activating cells and diffused into plasma, thus sustaining clot growth. Far from the activating cells, additional factor IXa was produced by factor XIa. Exogenously added TF had no effect on the clot growth rate, suggesting that plasma TF does not contribute significantly to the clot propagation process in a reaction-diffusion system without flow. Addition of thrombomodulin at 3–100 nM caused dose-dependent termination of clot elongation with a final clot size of 2–0.2 mm. These results identify roles of specific coagulation pathways at different stages of spatial clot formation (initiation, elongation, and termination) and provide a possible basis for their therapeutic targeting. PMID:16326897

  11. The cytochrome P450 epoxygenase pathway regulates the hepatic inflammatory response in fatty liver disease.

    PubMed

    Schuck, Robert N; Zha, Weibin; Edin, Matthew L; Gruzdev, Artiom; Vendrov, Kimberly C; Miller, Tricia M; Xu, Zhenghong; Lih, Fred B; DeGraff, Laura M; Tomer, Kenneth B; Jones, H Michael; Makowski, Liza; Huang, Leaf; Poloyac, Samuel M; Zeldin, Darryl C; Lee, Craig R

    2014-01-01

    Fatty liver disease is an emerging public health problem without effective therapies, and chronic hepatic inflammation is a key pathologic mediator in its progression. Cytochrome P450 (CYP) epoxygenases metabolize arachidonic acid to biologically active epoxyeicosatrienoic acids (EETs), which have potent anti-inflammatory effects. Although promoting the effects of EETs elicits anti-inflammatory and protective effects in the cardiovascular system, the contribution of CYP-derived EETs to the regulation of fatty liver disease-associated inflammation and injury is unknown. Using the atherogenic diet model of non-alcoholic fatty liver disease/non-alcoholic steatohepatitis (NAFLD/NASH), our studies demonstrated that induction of fatty liver disease significantly and preferentially suppresses hepatic CYP epoxygenase expression and activity, and both hepatic and circulating levels of EETs in mice. Furthermore, mice with targeted disruption of Ephx2 (the gene encoding soluble epoxide hydrolase) exhibited restored hepatic and circulating EET levels and a significantly attenuated induction of hepatic inflammation and injury. Collectively, these data suggest that suppression of hepatic CYP-mediated EET biosynthesis is an important pathological consequence of fatty liver disease-associated inflammation, and that the CYP epoxygenase pathway is a central regulator of the hepatic inflammatory response in NAFLD/NASH. Future studies investigating the utility of therapeutic strategies that promote the effects of CYP-derived EETs in NAFLD/NASH are warranted.

  12. Distinct ECM mechanosensing pathways regulate microtubule dynamics to control endothelial cell branching morphogenesis

    PubMed Central

    Myers, Kenneth A.; Applegate, Kathryn T.

    2011-01-01

    During angiogenesis, cytoskeletal dynamics that mediate endothelial cell branching morphogenesis during vascular guidance are thought to be regulated by physical attributes of the extracellular matrix (ECM) in a process termed mechanosensing. Here, we tested the involvement of microtubules in linking mechanosensing to endothelial cell branching morphogenesis. We used a recently developed microtubule plus end–tracking program to show that specific parameters of microtubule assembly dynamics, growth speed and growth persistence, are globally and regionally modified by, and contribute to, ECM mechanosensing. We demonstrated that engagement of compliant two-dimensional or three-dimensional ECMs induces local differences in microtubule growth speed that require myosin II contractility. Finally, we found that microtubule growth persistence is modulated by myosin II–mediated compliance mechanosensing when cells are cultured on two-dimensional ECMs, whereas three-dimensional ECM engagement makes microtubule growth persistence insensitive to changes in ECM compliance. Thus, compliance and dimensionality ECM mechanosensing pathways independently regulate specific and distinct microtubule dynamics parameters in endothelial cells to guide branching morphogenesis in physically complex ECMs. PMID:21263030

  13. Exosome proteomics reveals transcriptional regulator proteins with potential to mediate downstream pathways.

    PubMed

    Ung, Timothy H; Madsen, Helen J; Hellwinkel, Justin E; Lencioni, Alex M; Graner, Michael W

    2014-11-01

    Exosomes are virus-sized, membrane-enclosed vesicles with origins in the cellular endosomal system, but are released extracellularly. As a population, these tiny vesicles carry relatively enormous amounts of information in their protein, lipid and nucleic acid content, and the vesicles can have profound impacts on recipient cells. This review employs publically-available data combined with gene ontology applications to propose a novel concept, that exosomes transport transcriptional and translational machinery that may have direct impacts on gene expression in recipient cells. Here, we examine the previously published proteomic contents of medulloblastoma-derived exosomes, focusing on transcriptional regulators; we found that there are numerous proteins that may have potential roles in transcriptional and translational regulation with putative influence on downstream, cancer-related pathways. We expanded this search to all of the proteins in the Vesiclepedia database; using gene ontology approaches, we see that these regulatory factors are implicated in many of the processes involved in cancer initiation and progression. This information suggests that some of the effects of exosomes on recipient cells may be due to the delivery of protein factors that can directly and fundamentally change the transcriptional landscape of the cells. Within a tumor environment, this has potential to tilt the advantage towards the cancer.

  14. Reconstituting regulation of the canonical Wnt pathway by engineering a minimal β-catenin destruction machine

    PubMed Central

    Pronobis, Mira I.; Deuitch, Natalie; Posham, Vinya; Mimori-Kiyosue, Yuko; Peifer, Mark

    2017-01-01

    Negatively regulating key signaling pathways is critical to development and altered in cancer. Wnt signaling is kept off by the destruction complex, which is assembled around the tumor suppressors APC and Axin and targets β-catenin for destruction. Axin and APC are large proteins with many domains and motifs that bind other partners. We hypothesized that if we identified the essential regions required for APC:Axin cooperative function and used these data to design a minimal β-catenin-destruction machine, we would gain new insights into the core mechanisms of destruction complex function. We identified five key domains/motifs in APC or Axin that are essential for their function in reconstituting Wnt regulation. Strikingly, however, certain APC and Axin mutants that are nonfunctional on their own can complement one another in reducing β-catenin, revealing that the APC:Axin complex is a highly robust machine. We used these insights to design a minimal β-catenin-destruction machine, revealing that a minimized chimeric protein covalently linking the five essential regions of APC and Axin reconstitutes destruction complex internal structure, size, and dynamics, restoring efficient β-catenin destruction in colorectal tumor cells. On the basis of our data, we propose a new model of the mechanistic function of the destruction complex as an integrated machine. PMID:27852897

  15. Osteoblastic regulation of B lymphopoiesis is mediated by Gs{alpha}-dependent signaling pathways.

    PubMed

    Wu, Joy Y; Purton, Louise E; Rodda, Stephen J; Chen, Min; Weinstein, Lee S; McMahon, Andrew P; Scadden, David T; Kronenberg, Henry M

    2008-11-04

    Osteoblasts play an increasingly recognized role in supporting hematopoietic development and recently have been implicated in the regulation of B lymphopoiesis. Here we demonstrate that the heterotrimeric G protein alpha subunit G(s)alpha is required in cells of the osteoblast lineage for normal postnatal B lymphocyte production. Deletion of G(s)alpha early in the osteoblast lineage results in a 59% decrease in the percentage of B cell precursors in the bone marrow. Analysis of peripheral blood from mutant mice revealed a 67% decrease in the number of circulating B lymphocytes by 10 days of age. Strikingly, other mature hematopoietic lineages are not decreased significantly. Mice lacking G(s)alpha in cells of the osteoblast lineage exhibit a reduction in pro-B and pre-B cells. Furthermore, interleukin (IL)-7 expression is attenuated in G(s)alpha-deficient osteoblasts, and exogenous IL-7 is able to restore B cell precursor populations in the bone marrow of mutant mice. Finally, the defect in B lymphopoiesis can be rescued by transplantation into a WT microenvironment. These findings confirm that osteoblasts are an important component of the B lymphocyte niche and demonstrate in vivo that G(s)alpha-dependent signaling pathways in cells of the osteoblast lineage extrinsically regulate bone marrow B lymphopoiesis, at least partially in an IL-7-dependent manner.

  16. 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

  17. Reconstituting regulation of the canonical Wnt pathway by engineering a minimal β-catenin destruction machine.

    PubMed

    Pronobis, Mira I; Deuitch, Natalie; Posham, Vinya; Mimori-Kiyosue, Yuko; Peifer, Mark

    2017-01-01

    Negatively regulating key signaling pathways is critical to development and altered in cancer. Wnt signaling is kept off by the destruction complex, which is assembled around the tumor suppressors APC and Axin and targets β-catenin for destruction. Axin and APC are large proteins with many domains and motifs that bind other partners. We hypothesized that if we identified the essential regions required for APC:Axin cooperative function and used these data to design a minimal β-catenin-destruction machine, we would gain new insights into the core mechanisms of destruction complex function. We identified five key domains/motifs in APC or Axin that are essential for their function in reconstituting Wnt regulation. Strikingly, however, certain APC and Axin mutants that are nonfunctional on their own can complement one another in reducing β-catenin, revealing that the APC:Axin complex is a highly robust machine. We used these insights to design a minimal β-catenin-destruction machine, revealing that a minimized chimeric protein covalently linking the five essential regions of APC and Axin reconstitutes destruction complex internal structure, size, and dynamics, restoring efficient β-catenin destruction in colorectal tumor cells. On the basis of our data, we propose a new model of the mechanistic function of the destruction complex as an integrated machine.

  18. Redox and trace metal regulation of ion channels in the pain pathway

    PubMed Central

    Evans, J. Grayson; Todorovic, Slobodan M.

    2015-01-01

    Given the clinical significance of pain disorders and the relative ineffectiveness of current therapeutics, it is important to identify alternative means of modulating nociception. The most obvious pharmacological targets are the ion channels that facilitate nervous transmission from pain sensors in the periphery to the processing regions within the brain and spinal cord. In order to design effective pharmacological tools for this purpose, however, it is first necessary to understand how these channels are regulated. A growing area of research involves the investigation of the role that trace metals and endogenous redox agents play in modulating the activity of a diverse group of ion channels within the pain pathway. In the present review, the most recent literature concerning trace metal and redox regulation of T-type calcium channels, NMDA (N-methyl-D-aspartate) receptors, GABAA (γ-aminobutyric acid A) receptors and TRP (transient receptor potential) channels are described to gain a comprehensive understanding of the current state of the field as well as to provide a basis for future thought and experimentation. PMID:26341484

  19. Rassf5 and Ndr kinases regulate neuronal polarity through Par3 phosphorylation in a novel pathway.

    PubMed

    Yang, Rui; Kong, Eryan; Jin, Jing; Hergovich, Alexander; Püschel, Andreas W

    2014-08-15

    The morphology and polarized growth of cells depend on pathways that control the asymmetric distribution of regulatory factors. The evolutionarily conserved Ndr kinases play important roles in cell polarity and morphogenesis in yeast and invertebrates but it is unclear whether they perform a similar function in mammalian cells. Here, we analyze the function of mammalian Ndr1 and Ndr2 (also known as STK38 or STK38L, respectively) in the establishment of polarity in neurons. We show that they act downstream of the tumor suppressor Rassf5 and upstream of the polarity protein Par3 (also known as PARD3). Rassf5 and Ndr1 or Ndr2 are required during the polarization of hippocampal neurons to prevent the formation of supernumerary axons. Mechanistically, the Ndr kinases act by phosphorylating Par3 at Ser383 to inhibit its interaction with dynein, thereby polarizing the distribution of Par3 and reinforcing axon specification. Our results identify a novel Rassf5-Ndr-Par3 signaling cascade that regulates the transport of Par3 during the establishment of neuronal polarity. Their role in neuronal polarity suggests that Ndr kinases perform a conserved function as regulators of cell polarity.

  20. The Drosophila secreted protein Argos regulates signal transduction in the Ras/MAPK pathway.

    PubMed

    Sawamoto, K; Okabe, M; Tanimura, T; Mikoshiba, K; Nishida, Y; Okano, H

    1996-08-25

    The Drosophila argos gene encodes a secreted protein with an EGF motif which acts as an inhibitor of cellular differentiation in multiple developmental processes. To investigate the cellular pathways regulated by Argos, we screened for mutations which could modify the phenotype caused by overexpression of argos. We show that the effects of argos overexpression on the eye and wing vein development are suppressed by gain-of-function mutations of the MAPKK/D-MEK gene (Dsor1/D-mek) and the MAPK/ERK-A gene (rolled) and were enhanced by loss-of-function mutations of Star. Loss-of-function mutations in components of the Ras/MAPK signaling cascade act as dominant suppressors of the phenotype caused by the argos null mutations. A loss-of-function argos mutation enhanced the overproduction of R7 neurons caused by gain-of-function alleles of Son of sevenless and Dsor1. Conversely, overexpression of argos inhibited formation of the extra R7 cells that was caused by high-level MAPK/ERK-A activity. A phenotype of the sev; argos double mutants revealed that sev is epistatic to argos. These results provide evidence that Argos negatively regulates signal transduction events in the Ras/MAPK cascade.

  1. A developmentally regulated translational control pathway establishes the meiotic chromosome segregation pattern

    PubMed Central

    Berchowitz, Luke E.; Gajadhar, Aaron S.; van Werven, Folkert J.; De Rosa, Alexandra A.; Samoylova, Mariya L.; Brar, Gloria A.; Xu, Yifeng; Xiao, Che; Futcher, Bruce; Weissman, Jonathan S.; White, Forest M.; Amon, Angelika

    2013-01-01

    Production of haploid gametes from diploid progenitor cells is mediated by a specialized cell division, meiosis, where two divisions, meiosis I and II, follow a single S phase. Errors in progression from meiosis I to meiosis II lead to aneuploid and polyploid gametes, but the regulatory mechanisms controlling this transition are poorly understood. Here, we demonstrate that the conserved kinase Ime2 regulates the timing and order of the meiotic divisions by controlling translation. Ime2 coordinates translational activation of a cluster of genes at the meiosis I–meiosis II transition, including the critical determinant of the meiotic chromosome segregation pattern CLB3. We further show that Ime2 mediates translational control through the meiosis-specific RNA-binding protein Rim4. Rim4 inhibits translation of CLB3 during meiosis I by interacting with the 5′ untranslated region (UTR) of CLB3. At the onset of meiosis II, Ime2 kinase activity rises and triggers a decrease in Rim4 protein levels, thereby alleviating translational repression. Our results elucidate a novel developmentally regulated translational control pathway that establishes the meiotic chromosome segregation pattern. PMID:24115771

  2. Effect of surface chemistry on the integrin induced pathway in regulating vascular endothelial cells migration.

    PubMed

    Shen, Yang; Gao, Min; Ma, Yunlong; Yu, Hongchi; Cui, Fu-zhai; Gregersen, Hans; Yu, Qingsong; Wang, Guixue; Liu, Xiaoheng

    2015-02-01

    The migration of vascular endothelial cells (ECs) is essential for reendothelialization after implantation of cardiovascular biomaterials. Reendothelialization is largely determined by surface properties of implants. In this study, surfaces modified with various chemical functional groups (CH3, NH2, COOH, OH) prepared by self-assembled monolayers (SAMs) were used as model system. Expressions and distributions of critical proteins in the integrin-induced signaling pathway were examined to explore the mechanisms of surface chemistry regulating EC migration. The results showed that SAMs modulated cell migration were in the order CH3>NH2>OH>COOH, determined by differences in the expressions of focal adhesion components and Rho GTPases. Multiple integrin subunits showed difference in a surface chemistry-dependent manner, which induced a stepwise activation of signaling cascades associated with EC migration. This work provides a broad overview of surface chemistry regulated endothelial cell migration and establishes association among the surface chemistry, cell migration behavior and associated integrin signaling events. Understanding the relationship between these factors will help us to understand the surface/interface behavior between biomaterials and cells, reveal molecular mechanism of cells sensing surface characterization, and guide surface modification of cardiovascular implanted materials.

  3. THE PROTEASOME REGULATES BACTERIAL CpG DNA-INDUCED SIGNALING PATHWAYS IN MURINE MACROPHAGES

    PubMed Central

    Gao, Jian Jun; Shen, Jing; Kolbert, Christopher; Raghavakaimal, Sreekumar; Papasian, Christopher J.; Qureshi, Asaf A.; Vogel, Stefanie N.; Morrison, David C.; Qureshi, Nilofer

    2010-01-01

    Our previous work has provided strong evidence that the proteasome is central to the vast majority of genes induced in mouse macrophages in response to lipopolysaccharide (LPS) stimulation. In the studies presented here, we evaluated the role of the macrophage proteasome in response to a second microbial product CpG DNA (unmethylated bacterial DNA). For these studies, we applied Affymetrix microarray analysis of RNA derived from murine macrophages stimulated with CpG DNA in the presence or absence of proteasome inhibitor, lactacystin. The results of these studies revealed that similar to LPS, a vast majority of those macrophage genes regulated by CpG DNA are also under the control of the proteasome at 4 h. In contrast to LPS stimulation, however, many of these genes were induced much later than 4 h, at 18 h, in response to CpG DNA. Lactacystin treatment of macrophages completely blocked the CpG DNA-induced gene expression of TNF-α and other genes involved in production of inflammatory mediators. These data strongly support the conclusion that, similar to LPS, the macrophage proteasome is a key regulator of CpG DNA-induced signaling pathways. PMID:20160661

  4. WRN regulates pathway choice between classical and alternative non-homologous end joining

    PubMed Central

    Shamanna, Raghavendra A.; Lu, Huiming; de Freitas, Jessica K.; Tian, Jane; Croteau, Deborah L.; Bohr, Vilhelm A.

    2016-01-01

    Werner syndrome (WS) is an accelerated ageing disorder with genomic instability caused by WRN protein deficiency. Many features seen in WS can be explained by the diverse functions of WRN in DNA metabolism. However, the origin of the large genomic deletions and telomere fusions are not yet understood. Here, we report that WRN regulates the pathway choice between classical (c)- and alternative (alt)-nonhomologous end joining (NHEJ) during DNA double-strand break (DSB) repair. It promotes c-NHEJ via helicase and exonuclease activities and inhibits alt-NHEJ using non-enzymatic functions. When WRN is recruited to the DSBs it suppresses the recruitment of MRE11 and CtIP, and protects the DSBs from 5′ end resection. Moreover, knockdown of Wrn, alone or in combination with Trf2 in mouse embryonic fibroblasts results in increased telomere fusions, which were ablated by Ctip knockdown. We show that WRN regulates alt-NHEJ and shields DSBs from MRE11/CtIP-mediated resection to prevent large deletions and telomere fusions. PMID:27922005

  5. DNMT1-PPARγ pathway in macrophages regulates chronic inflammation and atherosclerosis development in mice

    PubMed Central

    Yu, Jie; Qiu, Youzhu; Yang, Jie; Bian, Shizhu; Chen, Guozhu; Deng, Mengyang; Kang, Huali; Huang, Lan

    2016-01-01

    The DNA methyltransferase-mediated proinflammatory activation of macrophages is causally linked to the development of atherosclerosis (AS). However, the role of DNMT1, a DNA methylation maintenance enzyme, in macrophage polarization and AS development remains obscure. Here, we established transgenic mice with macrophage-specific overexpression of DNMT1 (TgDNMT1) or PPAR-γ (TgPPAR-γ) to investigate their effects on AS progression in ApoE-knockout mice fed an atherogenic diet. Primary macrophages were extracted to study the role of the DNMT1/PPAR-γ pathway in regulating inflammatory cytokine production. We demonstrated that TgDNMT1 significantly increased proinflammatory cytokine production in macrophages and plasma, and it accelerated the progression of AS in the atherogenic diet-treated ApoE-knockout mice. Further, we found that the DNA methylation status of the proximal PPAR-γ promoter was regulated by DNMT1 in macrophages. Notably, additional TgPPAR-γ or pharmacological activation of PPAR-γ effectively prevented TgDNMT1-induced proinflammatory cytokine production in macrophages and AS development in the mouse model. Finally, we demonstrated that elevated DNMT1 was correlated with decreased PPAR-γ, and increased proinflammatory cytokine production in the peripheral blood monocytes isolated from the patients with AS, compared to those of healthy donors. Our findings shed light on a novel strategy for the prevention and therapy of AS. PMID:27530451

  6. N-Myristoylation Regulates the SnRK1 Pathway in Arabidopsis[W

    PubMed Central

    Pierre, Michèle; Traverso, José A.; Boisson, Bertrand; Domenichini, Séverine; Bouchez, David; Giglione, Carmela; Meinnel, Thierry

    2007-01-01

    Cotranslational and posttranslational modifications are increasingly recognized as important in the regulation of numerous essential cellular functions. N-myristoylation is a lipid modification ensuring the proper function and intracellular trafficking of proteins involved in many signaling pathways. Arabidopsis thaliana, like human, has two tightly regulated N-myristoyltransferase (NMT) genes, NMT1 and NMT2. Characterization of knockout mutants showed that NMT1 was strictly required for plant viability, whereas NMT2 accelerated flowering. NMT1 impairment induced extremely severe defects in the shoot apical meristem during embryonic development, causing growth arrest after germination. A transgenic plant line with an inducible NMT1 gene demonstrated that NMT1 expression had further effects at later stages. NMT2 did not compensate for NMT1 in the nmt1-1 mutant, but NMT2 overexpression resulted in shoot and root meristem abnormalities. Various data from complementation experiments in the nmt1-1 background, using either yeast or human NMTs, demonstrated a functional link between the developmental arrest of nmt1-1 mutants and the myristoylation state of an extremely small set of protein targets. We show here that protein N-myristoylation is systematically associated with shoot meristem development and that SnRK1 (for SNF1-related kinase) is one of its essential primary targets. PMID:17827350

  7. Regulation and disregulation of mammalian nucleotide excision repair: A pathway to nongermline breast carcinogenesis

    SciTech Connect

    Latimer, Jean J.; Majekwana, Vongai J.; Pabon-Padin, Yashira R.; Pimpley, Manasi R.; Grant, Stephen G.

    2014-12-19

    Nucleotide excision repair (NER) is important as a modulator of disease, especially in constitutive deficiencies, such as the cancer predisposition syndrome Xeroderma pigmentosum. We have found profound variation of NER capacity among normal individuals, between cell-types and during carcinogenesis. NER is a repair system for many types of DNA damage, and therefore many types of genotoxic carcinogenic exposures, including ultraviolet light, products of organic combustion, metals, oxidative stress, etc. Since NER is intimately related to cellular metabolism, requiring components of both the DNA replicative and transcription machinery, it has a narrow range of functional viability. Thus, genes in the NER pathway are expressed at the low levels manifested by, for example, nuclear transcription factors. Since NER activity and gene expression vary by cell-type, it is inherently epigenetically regulated. Furthermore, this epigenetic regulation is disregulated during sporadic breast carcinogenesis. Loss of NER is one basis of genomic instability, a required element in cellular transformation, and one that potentially modulates response to therapy. In this article, we demonstrate differences in NER capacity in eight adult mouse tissues, and place this result into the context of our previous work on mouse extraembryonic tissues, normal human tissues and sporadic early stage human breast cancer.

  8. Regulation and disregulation of mammalian nucleotide excision repair: A pathway to nongermline breast carcinogenesis

    DOE PAGES

    Latimer, Jean J.; Majekwana, Vongai J.; Pabon-Padin, Yashira R.; ...

    2014-12-19

    Nucleotide excision repair (NER) is important as a modulator of disease, especially in constitutive deficiencies, such as the cancer predisposition syndrome Xeroderma pigmentosum. We have found profound variation of NER capacity among normal individuals, between cell-types and during carcinogenesis. NER is a repair system for many types of DNA damage, and therefore many types of genotoxic carcinogenic exposures, including ultraviolet light, products of organic combustion, metals, oxidative stress, etc. Since NER is intimately related to cellular metabolism, requiring components of both the DNA replicative and transcription machinery, it has a narrow range of functional viability. Thus, genes in the NERmore » pathway are expressed at the low levels manifested by, for example, nuclear transcription factors. Since NER activity and gene expression vary by cell-type, it is inherently epigenetically regulated. Furthermore, this epigenetic regulation is disregulated during sporadic breast carcinogenesis. Loss of NER is one basis of genomic instability, a required element in cellular transformation, and one that potentially modulates response to therapy. In this article, we demonstrate differences in NER capacity in eight adult mouse tissues, and place this result into the context of our previous work on mouse extraembryonic tissues, normal human tissues and sporadic early stage human breast cancer.« less

  9. Transcriptional regulation of the Drosophila orc2 gene by the DREF pathway.

    PubMed

    Okudaira, Koji; Ohno, Katsuhito; Yoshida, Hideki; Asano, Maki; Hirose, Fumiko; Yamaguchi, Masamitsu

    2005-12-30

    DNA replication-related element (DRE) and the DRE-binding factor (DREF) play an important role in regulating DNA replication-related genes such as PCNA and DNA polymerase alpha in Drosophila. We have previously reported that overexpression of DREF in developing eye imaginal discs induced ectopic DNA synthesis and apoptosis, which results in rough eyes. To identify genetic interactants with the DREF gene, we have carried out a screen for modifiers of the rough eye phenotype. One of the suppressor genes identified was the Drosophila orc2 gene. A search for known transcription factor recognition sites revealed that the orc2 gene contains three DREs, named DRE1 (+14 to +21), DRE2 (-205 to -198), and DRE3 (-709 to -702). Band mobility shift analysis using Kc cell nuclear extracts detected the specific complex formed between DREF and the DRE1 or DRE2. Specific binding of DREF to genomic region containing the DRE1 or DRE2 was further demonstrated by chromatin immunoprecipitation assays, suggesting that these are the genuine complexes formed in vivo. The luciferase assay in Kc cells indicated that the DRE sites in the orc2 promoter are involved in a transcriptional regulation of the orc2 gene. The results, taken together, demonstrate that the orc2 gene is under the control of DREF pathway.

  10. Developmental and stress regulation of gene expression for plastid and cytosolic isoprenoid pathways in pepper fruits.

    PubMed Central

    Hugueney, P; Bouvier, F; Badillo, A; Quennemet, J; d'Harlingue, A; Camara, B

    1996-01-01

    Plant cells synthesize a myriad of isoprenoid compounds in different subcellular compartments, which include the plastid, the mitochondria, and the endoplasmic reticulum cytosol. To start the study of the regulation of these parallel pathways, we used pepper (Capsicum annuum) fruit as a model. Using different isoprenoid biosynthetic gene probes from cloned cDNAs, we showed that only genes encoding the plastid enzymes (geranylgeranyl pyrophosphate synthase, phytoene synthase, phytoene desaturase, and capasanthin-capsorubin synthase) are specifically triggered during the normal period of development, at the ripening stage. This pattern of expression can be mimicked and precociously induced by a simple wounding stress. Concerning the cytosol-located enzymes, we observed that the expression of the gene encoding farnesyl pyrophosphate synthase is constitutive, whereas that of farnesyl pyrophosphate cyclase (5-epi-aristolochene synthase) is undetectable during the normal development of the fruit. The expression of these later genes are, however, only selectively triggered after elicitor treatment. The results provide evidence for developmental control of isoprenoid biosynthesis occurring in plastids and that cytoplasmic isoprenoid biosynthesis is regulated, in part, by environmental signals. PMID:8787029

  11. Phytochrome and retrograde signalling pathways converge to antagonistically regulate a light-induced transcriptional network.

    PubMed

    Martín, Guiomar; Leivar, Pablo; Ludevid, Dolores; Tepperman, James M; Quail, Peter H; Monte, Elena

    2016-05-06

    Plastid-to-nucleus retrograde signals emitted by dysfunctional chloroplasts impact photomorphogenic development, but the molecular link between retrograde- and photosensory-receptor signalling has remained unclear. Here, we show that the phytochrome and retrograde signalling (RS) pathways converge antagonistically to regulate the expression of the nuclear-encoded transcription factor GLK1, a key regulator of a light-induced transcriptional network central to photomorphogenesis. GLK1 gene transcription is directly repressed by PHYTOCHROME-INTERACTING FACTOR (PIF)-class bHLH transcription factors in darkness, but light-activated phytochrome reverses this activity, thereby inducing expression. Conversely, we show that retrograde signals repress this induction by a mechanism independent of PIF mediation. Collectively, our data indicate that light at moderate levels acts through the plant's nuclear-localized sensory-photoreceptor system to induce appropriate photomorphogenic development, but at excessive levels, sensed through the separate plastid-localized RS system, acts to suppress such development, thus providing a mechanism for protection against photo-oxidative damage by minimizing the tissue exposure to deleterious radiation.

  12. Phytochrome and retrograde signalling pathways converge to antagonistically regulate a light-induced transcriptional network

    PubMed Central

    Martín, Guiomar; Leivar, Pablo; Ludevid, Dolores; Tepperman, James M.; Quail, Peter H.; Monte, Elena

    2016-01-01

    Plastid-to-nucleus retrograde signals emitted by dysfunctional chloroplasts impact photomorphogenic development, but the molecular link between retrograde- and photosensory-receptor signalling has remained unclear. Here, we show that the phytochrome and retrograde signalling (RS) pathways converge antagonistically to regulate the expression of the nuclear-encoded transcription factor GLK1, a key regulator of a light-induced transcriptional network central to photomorphogenesis. GLK1 gene transcription is directly repressed by PHYTOCHROME-INTERACTING FACTOR (PIF)-class bHLH transcription factors in darkness, but light-activated phytochrome reverses this activity, thereby inducing expression. Conversely, we show that retrograde signals repress this induction by a mechanism independent of PIF mediation. Collectively, our data indicate that light at moderate levels acts through the plant's nuclear-localized sensory-photoreceptor system to induce appropriate photomorphogenic development, but at excessive levels, sensed through the separate plastid-localized RS system, acts to suppress such development, thus providing a mechanism for protection against photo-oxidative damage by minimizing the tissue exposure to deleterious radiation. PMID:27150909

  13. Wnt/β-Catenin and Sonic Hedgehog Pathways Interact in the Regulation of the Development of the Dorsal Mesenchymal Protrusion

    PubMed Central

    Briggs, Laura E.; Burns, Tara A.; Lockhart, Marie M.; Phelps, Aimee L.; Van den Hoff, Maurice J.B.; Wessels, Andy

    2016-01-01

    Background The dorsal mesenchymal protrusion (DMP) is a second heart field (SHF) derived tissue involved in cardiac septation. Molecular mechanisms controlling SHF/DMP development include the Bone Morphogenetic Protein and Wnt/β-catenin signaling pathways. Reduced expression of components in these pathways leads to inhibition of proliferation of the SHF/DMP precursor population and failure of the DMP to develop. While the Sonic Hedgehog (Shh) pathway has also been demonstrated to be critically important for SHF/DMP development and atrioventricular septation, its role in the regulation of SHF proliferation is contentious. Results Tissue-specific deletion of the Shh receptor Smoothened from the SHF resulted in compromised DMP formation and atrioventricular septal defects (AVSDs). Immunohistochemical analysis at critical stages of DMP development showed significant proliferation defect as well as reduction in levels of the Wnt/β-catenin pathway-intermediates β-catenin, Lef1, and Axin2. To determine whether the defects seen in the conditional Smoothened knock-out mouse could be attributed to reduced Wnt/β-catenin signaling, LiCl, a pharmacological activator of this Wnt/β-catenin pathway, was administered. This resulted in restoration of proliferation and partial rescue of the AVSD phenotype. Conclusions The data presented suggest that the Wnt/β-catenin pathway interact with the Shh pathway in the regulation of SHF/DMP-precursor proliferation and, hence, the development of the DMP. PMID:26297872

  14. Early Diagnosis of Clear Cell Kidney Cancer via VHL/HIF Pathway Regulated-Circulating microRNA

    DTIC Science & Technology

    2016-05-01

    Award Number: W81XWH-11-1-0715 TITLE: Early Diagnosis of Clear Cell Kidney Cancer via VHL/HIF Pathway-Regulated Circulating microRNA PRINCIPAL...TITLE AND SUBTITLE Sa. CONTRACT NUMBER Early Diagnosis of Clear Cell Kidney Cancer via VHL/HIF Pathway- Regulated Circulating microRNA Sb. GRANT NUMBER...panel of diagnostic miRNAs that are measurable in serum and will be able to identify kidney cancer in its earliest stages. We hypothesized that serum

  15. Early Diagnosis of Clear Cell Kidney Cancer via VHL/HIF Pathway-Regulated Circulating microRNA

    DTIC Science & Technology

    2014-09-01

    Award Number: W81XWH-11-1-0715 TITLE: Early Diagnosis of Clear Cell Kidney Cancer via VHL/HIF Pathway-Regulated Circulating microRNA PRINCIPAL...31Aug2014 4. TITLE AND SUBTITLE Sa. CONTRACT NUMBER Early Diagnosis of Clear Cell Kidney Cancer via VHL/HIF Pathway- Regulated Circulating microRNA Sb...identify a panel of diagnostic miRNAs that are measurable in serum and will be able to identify kidney cancer in its earliest stages. We hypothesized

  16. Regulation of cell migration via the EGFR signaling pathway in oral squamous cell carcinoma cells

    PubMed Central

    Ohnishi, Yuichi; Yasui, Hiroki; Kakudo, Kenji; Nozaki, Masami

    2017-01-01

    Cell migration potency is essential in cancer metastasis and is often regulated by extracellular stimuli. Oral squamous cell carcinoma cell lines include those that are sensitive, as well as resistant, to the effects of the epidermal growth factor receptor (EGFR) inhibitor cetuximab on cell migration. In the present study, the molecular differences in the EGFR response to cell migration between the SAS cetuximab-sensitive and HSC4 cetuximab-resistant cell lines was examined. Treatment with the EGFR inhibitors AG1478 and cetuximab reduced the migration potency of SAS cells, but not HSC4 cells. The migration of the two cell lines was inhibited under serum-free culture conditions, and the addition of EGF to the serum-free medium promoted the migration of SAS cells, but not HSC4 cells. In addition, SAS cell migration was reduced by the mitogen-activated protein kinase kinase and protein kinase B (Akt) inhibitors PD98059 and MK2206, whereas HSC4 cell migration was only inhibited by MK2206. EGF induced an increase in extracellular signal-regulated kinase phosphorylation levels in HSC4 cells, and stimulated Akt phosphorylation levels in SAS cells. Furthermore, the staining of actin filaments with phalloidin was significantly increased by the inhibition of EGFR in SAS cells, but was not observed as altered in HSC4 cells. Conversely, the addition of EGF to the culture medium decreased the accumulation of actin filaments in SAS cells. The results suggest that the EGF-EGFR signaling pathway has an important role in SAS cell migration via the modulation of actin dynamics, and that HSC4 cell migration is regulated by a serum component other than EGFR.

  17. Complex Regulation Pathways of AmpC-Mediated β-Lactam Resistance in Enterobacter cloacae Complex.

    PubMed

    Guérin, François; Isnard, Christophe; Cattoir, Vincent; Giard, Jean Christophe

    2015-12-01

    Enterobacter cloacae complex (ECC), an opportunistic pathogen causing numerous infections in hospitalized patients worldwide, is able to resist β-lactams mainly by producing the AmpC β-lactamase enzyme. AmpC expression is highly inducible in the presence of some β-lactams, but the underlying genetic regulation, which is intricately linked to peptidoglycan recycling, is still poorly understood. In this study, we constructed different mutant strains that were affected in genes encoding enzymes suspected to be involved in this pathway. As expected, the inactivation of ampC, ampR (which encodes the regulator protein of ampC), and ampG (encoding a permease) abolished β-lactam resistance. Reverse transcription-quantitative PCR (qRT-PCR) experiments combined with phenotypic studies showed that cefotaxime (at high concentrations) and cefoxitin induced the expression of ampC in different ways: one involving NagZ (a N-acetyl-β-D-glucosaminidase) and another independent of NagZ. Unlike the model established for Pseudomonas aeruginosa, inactivation of DacB (also known as PBP4) was not responsible for a constitutive ampC overexpression in ECC, whereas it caused AmpC-mediated high-level β-lactam resistance, suggesting a post-transcriptional regulation mechanism. Global transcriptomic analysis by transcriptome sequencing (RNA-seq) of a dacB deletion mutant confirmed these results. Lastly, analysis of 37 ECC clinical isolates showed that amino acid changes in the AmpD sequence were likely the most crucial event involved in the development of high-level β-lactam resistance in vivo as opposed to P. aeruginosa where dacB mutations have been commonly found. These findings bring new elements for a better understanding of β-lactam resistance in ECC, which is essential for the identification of novel potential drug targets.

  18. Complex Regulation Pathways of AmpC-Mediated β-Lactam Resistance in Enterobacter cloacae Complex

    PubMed Central

    Guérin, François; Isnard, Christophe; Giard, Jean Christophe

    2015-01-01

    Enterobacter cloacae complex (ECC), an opportunistic pathogen causing numerous infections in hospitalized patients worldwide, is able to resist β-lactams mainly by producing the AmpC β-lactamase enzyme. AmpC expression is highly inducible in the presence of some β-lactams, but the underlying genetic regulation, which is intricately linked to peptidoglycan recycling, is still poorly understood. In this study, we constructed different mutant strains that were affected in genes encoding enzymes suspected to be involved in this pathway. As expected, the inactivation of ampC, ampR (which encodes the regulator protein of ampC), and ampG (encoding a permease) abolished β-lactam resistance. Reverse transcription-quantitative PCR (qRT-PCR) experiments combined with phenotypic studies showed that cefotaxime (at high concentrations) and cefoxitin induced the expression of ampC in different ways: one involving NagZ (a N-acetyl-β-d-glucosaminidase) and another independent of NagZ. Unlike the model established for Pseudomonas aeruginosa, inactivation of DacB (also known as PBP4) was not responsible for a constitutive ampC overexpression in ECC, whereas it caused AmpC-mediated high-level β-lactam resistance, suggesting a post-transcriptional regulation mechanism. Global transcriptomic analysis by transcriptome sequencing (RNA-seq) of a dacB deletion mutant confirmed these results. Lastly, analysis of 37 ECC clinical isolates showed that amino acid changes in the AmpD sequence were likely the most crucial event involved in the development of high-level β-lactam resistance in vivo as opposed to P. aeruginosa where dacB mutations have been commonly found. These findings bring new elements for a better understanding of β-lactam resistance in ECC, which is essential for the identification of novel potential drug targets. PMID:26438498

  19. HDAC6 regulates GR signaling in serotonin pathways with critical impact on stress resilience

    PubMed Central

    Espallergues, Julie; Teegarden, Sarah L.; Veerakumar, Avin; Boulden, Janette; Challis, Collin; Jochems, Jeanine; Chan, Michael; Petersen, Tess; Deneris, Evan; Matthias, Patrick; Hahn, Chang-Gyu; Lucki, Irwin; Beck, Sheryl G.; Berton, Olivier

    2012-01-01

    Genetic variations in certain components of the glucocorticoid receptor (GR) chaperone complex have been associated with the development of stress-related affective disorders and individual variability in therapeutic responses to antidepressants. Mechanisms that link GR chaperoning and stress susceptibility are not well understood. Here, we show that the effects of glucocorticoid hormones on socioaffective behaviors are critically regulated via reversible acetylation of Hsp90, a key component of the GR chaperone complex. We provide pharmacological and genetic evidence indicating that the cytoplasmic lysine deacetylase HDAC6 controls Hsp90 acetylation in the brain, and thereby modulates Hsp90-GR protein-protein interactions, as well as hormone- and stress-induced GR translocation, with a critical impact on GR downstream signaling and behavior. Pet1-Cre driven deletion of HDAC6 in serotonin neurons, the densest HDAC6-expressing cell group in the mouse brain, dramatically reduced acute anxiogenic effects of the glucocorticoid hormone corticosterone in the open field, elevated plus maze, and social interaction tests. Serotonin-selective depletion of HDAC6 also blocked the expression of social avoidance in mice exposed to chronic social defeat and concurrently prevented the electrophysiological and morphological changes induced, in serotonin neurons, by this murine model of traumatic stress. Together, these results identify HDAC6 inhibition as a potential new strategy for pro-resilience and antidepressant interventions through regulation of the Hsp90-GR heterocomplex and focal prevention of GR signaling in serotonin pathways. Our data thus uncover an alternate mechanism by which pan-HDAC inhibitors may regulate stress-related behaviors independently of their action on histones. PMID:22457490

  20. Aromatic amino acid biosynthesis in the yeast Saccharomyces cerevisiae: a model system for the regulation of a eukaryotic biosynthetic pathway.

    PubMed Central

    Braus, G H

    1991-01-01

    This review focuses on the gene-enzyme relationships and the regulation of different levels of the aromatic amino acid biosynthetic pathway in a simple eukaryotic system, the unicellular yeast Saccharomyces cerevisiae. Most reactions of this branched pathway are common to all organisms which are able to synthesize tryptophan, phenylalanine, and tyrosine. The current knowledge about the two main control mechanisms of the yeast aromatic amino acid biosynthesis is reviewed. (i) At the transcriptional level, most structural genes are regulated by the transcriptional activator GCN4, the regulator of the general amino acid control network, which couples transcriptional derepression to amino acid starvation of numerous structural genes in multiple amino acid biosynthetic pathways. (ii) At the enzyme level, the carbon flow is controlled mainly by modulating the enzyme activities at the first step of the pathway and at the branch points by feedback action of the three aromatic amino acid end products. Implications of these findings for the relationship of S. cerevisiae to prokaryotic as well as to higher eukaryotic organisms and for general regulatory mechanisms occurring in a living cell such as initiation of transcription, enzyme regulation, and the regulation of a metabolic branch point are discussed. PMID:1943992

  1. Regulation of PGE2 signaling pathways and TNF-alpha signaling pathways on the function of bone marrow-derived dendritic cells and the effects of CP-25.

    PubMed

    Li, Ying; Sheng, Kangliang; Chen, Jingyu; Wu, Yujing; Zhang, Feng; Chang, Yan; Wu, Huaxun; Fu, Jingjing; Zhang, Lingling; Wei, Wei

    2015-12-15

    This study was to investigate PGE2 and TNF-alpha signaling pathway involving in the maturation and activation of bone marrow dendritic cells (DCs) and the effect of CP-25. Bone marrow DCs were isolated and stimulated by PGE2 and TNF-alpha respectively. The markers of maturation and activation expressed on DCs, such as CD40, CD80, CD83, CD86, MHC-II, and the ability of antigen uptake of DCs were analyzed by flow cytometry. The proliferation of T cells co-cultured with DCs, the signaling pathways of PGE2-EP4-cAMP and TNF-alpha-TRADD-TRAF2-NF-κB in DCs were analyzed. The results showed that both PGE2 and TNF-alpha up-regulated the expressions of CD40, CD80, CD83, CD86, and MHC-II, decreased the antigen uptake of DCs, and DCs stimulated by PGE2 or TNF-alpha could increase T cell proliferation. CP-25 (10(-5), 10(-6), and 10(-7)mol/l) decreased significantly the expressions of CD40, CD80, CD83, CD86 and MHC-II, increased the antigen uptake of DCs, and suppressed T cell proliferation induced by DCs. PGE2 increased the expressions of EP4, NF-κB and down-regulated cAMP level of DCs. TNF-alpha could also up-regulate TNFR1, TRADD, TRAF2, and NF-κB expression of DCs. CP-25 (10(-5), 10(-6), and 10(-7)mol/l) decreased the expressions of EP4 and NF-κB, increased cAMP level in DCs stimulated by PGE2. CP-25 (10(-5), 10(-6), and 10(-7)mol/l) also could down-regulate significantly TNFR1, TRADD, TRAF2, and NF-κB expression in DCs stimulated by TNF-alpha. These results demonstrate that PGE2 and TNF-alpha could enhance DCs functions by mediating PGE2-EP4-cAMP pathway, TNF-alpha-TNFR1-TRADD-TRAF2-NF-κB pathway respectively. CP-25 might inhibit the function of DCs through regulating PGE2-EP4-cAMP and TNF-alpha-TNFR1-TRADD-TRAF2-NF-κB pathways.

  2. An Evaluation of Active Learning Causal Discovery Methods for Reverse-Engineering Local Causal Pathways of Gene Regulation.

    PubMed

    Ma, Sisi; Kemmeren, Patrick; Aliferis, Constantin F; Statnikov, Alexander

    2016-03-04

    Reverse-engineering of causal pathways that implicate diseases and vital cellular functions is a fundamental problem in biomedicine. Discovery of the local causal pathway of a target variable (that consists of its direct causes and direct effects) is essential for effective intervention and can facilitate accurate diagnosis and prognosis. Recent research has provided several active learning methods that can leverage passively observed high-throughput data to draft causal pathways and then refine the inferred relations with a limited number of experiments. The current study provides a comprehensive evaluation of the performance of active learning methods for local causal pathway discovery in real biological data. Specifically, 54 active learning methods/variants from 3 families of algorithms were applied for local causal pathways reconstruction of gene regulation for 5 transcription factors in S. cerevisiae. Four aspects of the methods' performance were assessed, including adjacency discovery quality, edge orientation accuracy, complete pathway discovery quality, and experimental cost. The results of this study show that some methods provide significant performance benefits over others and therefore should be routinely used for local causal pathway discovery tasks. This study also demonstrates the feasibility of local causal pathway reconstruction in real biological systems with significant quality and low experimental cost.

  3. Testosterone regulates smooth muscle contractile pathways in the rat prostate: emphasis on PDE5 signaling

    PubMed Central

    Zhang, Xinhua; Zang, Ning; Wei, Yu; Yin, Jin; Teng, Ruobing; Seftel, Allen

    2012-01-01

    Testosterone (T) plays a permissive role in the development of benign prostatic hyperplasia (BPH), and phosphodiesterase 5 inhibitors (PDE5is) have been found to be effective for BPH and lower urinary tract symptoms (LUTS) in clinical trials. This study investigated the effect of T on smooth muscle (SM) contractile and regulatory signaling pathways, including PDE5 expression and functional activity in prostate in male rats (sham-operated, surgically castrated, and castrated with T supplementation). In vitro organ bath studies, real-time RT-PCR, Western blot analysis, and immunohistochemistry were performed. Castration heavily attenuated contractility, including sensitivity to phenylephrine with SM myosin immunostaining revealing a disrupted SM cell arrangement in the stroma. PDE5 was immunolocalized exclusively in the prostate stroma, and orchiectomy signficantly reduced PDE5 immunopositivity, mRNA, and protein expression, along with nNOS and ROKβ mRNA, whereas it increased eNOS plus α1a and α1b adrenoreceptor expression in castrated animals. The PDE5i zaprinast significantly increased prostate strip relaxation to the nitric oxide donor sodium nitroprusside (SNP) in control but not castrated rats. But SNP alone was more effective on castrated rats, comparable with sham treated with SNP plus zaprinast. T supplementation prevented or restored all above changes, including SNP and zaprinast in vitro responsiveness. In conclusion, our data show that T positively regulates PDE5 expression and functional activities in prostate, and T ablation not only suppresses prostate size but also reduces prostatic SM contractility, with several potential SM contraction/relaxation pathways implicated. Zaprinast findings strongly suggest a major role for PDE5/cGMP in this signaling cascade. PDE5 inhibition may represent a novel mechanism for treatment of BPH. PMID:22028410

  4. Notch pathway regulates female germ cell meiosis progression and early oogenesis events in fetal mouse.

    PubMed

    Feng, Yan-Min; Liang, Gui-Jin; Pan, Bo; Qin, Xun-Si; Zhang, Xi-Feng; Chen, Chun-Lei; Li, Lan; Cheng, Shun-Feng; De Felici, Massimo; Shen, Wei

    2014-01-01

    A critical process of early oogenesis is the entry of mitotic oogonia into meiosis, a cell cycle switch regulated by a complex gene regulatory network. Although Notch pathway is involved in numerous important aspects of oogenesis in invertebrate species, whether it plays roles in early oogenesis events in mammals is unknown. Therefore, the rationale of the present study was to investigate the roles of Notch signaling in crucial processes of early oogenesis, such as meiosis entry and early oocyte growth. Notch receptors and ligands were localized in mouse embryonic female gonads and 2 Notch inhibitors, namely DAPT and L-685,458, were used to attenuate its signaling in an in vitro culture system of ovarian tissues from 12.5 days post coitum (dpc) fetus. The results demonstrated that the expression of Stra8, a master gene for germ cell meiosis, and its stimulation by retinoic acid (RA) were reduced after suppression of Notch signaling, and the other meiotic genes, Dazl, Dmc1, and Rec8, were abolished or markedly decreased. Furthermore, RNAi of Notch1 also markedly inhibited the expression of Stra8 and SCP3 in cultured female germ cells. The increased methylation status of CpG islands within the Stra8 promoter of the oocytes was observed in the presence of DAPT, indicating that Notch signaling is probably necessary for maintaining the epigenetic state of this gene in a way suitable for RA stimulation. Furthermore, in the presence of Notch inhibitors, progression of oocytes through meiosis I was markedly delayed. At later culture periods, the rate of oocyte growth was decreased, which impaired subsequent primordial follicle assembly in cultured ovarian tissues. Taken together, these results suggested new roles of the Notch signaling pathway in female germ cell meiosis progression and early oogenesis events in mammals.

  5. WNT regulation of embryonic development likely involves pathways independent of nuclear CTNNB1.

    PubMed

    Tribulo, Paula; Moss, James I; Ozawa, Manabu; Jiang, Zongliang; Tian, Xiuchun Cindy; Hansen, Peter J

    2017-04-01

    The bovine was used to examine the potential for WNT signaling to affect the preimplantation embryo. Expression of seven key genes involved in canonical WNT signaling declined to a nadir at the morula or blastocyst stage. Expression of 80 genes associated with WNT signaling in the morula and inner cell mass (ICM) and trophectoderm (TE) of the blastocyst was also evaluated. Many genes associated with WNT signaling were characterized by low transcript abundance. Seven genes were different between ICM and TE, and all of them were overexpressed in TE as compared to ICM, including WNT6, FZD1, FZD7, LRP6, PORCN, APC and SFRP1 Immunoreactive CTNNB1 was localized primarily to the plasma membrane at all stages examined from the 2-cell to blastocyst stages of development. Strikingly, neither CTNNB1 nor non-phospho (i.e., active) CTNNB1 was observed in the nucleus of blastomeres at any stage of development even after the addition of WNT activators to culture. In contrast, CTNNB1 associated with the plasma membrane was increased by activators of WNT signaling. The planar cell polarity pathway (PCP) could be activated in the embryo as indicated by an experiment demonstrating an increase in phospho-JNK in the nucleus of blastocysts treated with the non-canonical WNT11. Furthermore, WNT11 improved development to the blastocyst stage. In conclusion, canonical WNT signaling is attenuated in the preimplantation bovine embryo but WNT can activate the PCP component JNK. Thus, regulation of embryonic development by WNT is likely to involve activation of pathways independent of nuclear actions of CTNNB1.

  6. miR-139-5p is a regulator of metastatic pathways in breast cancer

    PubMed Central

    Krishnan, Keerthana; Steptoe, Anita L.; Martin, Hilary C.; Pattabiraman, Diwakar R.; Nones, Katia; Waddell, Nic; Mariasegaram, Mythily; Simpson, Peter T.; Lakhani, Sunil R.; Vlassov, Alexander; Grimmond, Sean M.; Cloonan, Nicole

    2013-01-01

    Metastasis is a complex, multistep process involved in the progression of cancer from a localized primary tissue to distant sites, often characteristic of the more aggressive forms of this disease. Despite being studied in great detail in recent years, the mechanisms that govern this process remain poorly understood. In this study, we identify a novel role for miR-139-5p in the inhibition of breast cancer progression. We highlight its clinical relevance by reviewing miR-139-5p expression across a wide variety of breast cancer subtypes using in-house generated and online data sets to show that it is most frequently lost in invasive tumors. A biotin pull-down approach was then used to identify the mRNA targets of miR-139-5p in the breast cancer cell line MCF7. Functional enrichment analysis of the pulled-down targets showed significant enrichment of genes in pathways previously implicated in breast cancer metastasis (P < 0.05). Further bioinformatic analysis revealed a predicted disruption to the TGFβ, Wnt, Rho, and MAPK/PI3K signaling cascades, implying a potential role for miR-139-5p in regulating the ability of cells to invade and migrate. To corroborate this finding, using the MDA-MB-231 breast cancer cell line, we show that overexpression of miR-139-5p results in suppression of these cellular phenotypes. Furthermore, we validate the interaction between miR-139-5p and predicted targets involved in these pathways. Collectively, these results suggest a significant functional role for miR-139-5p in breast cancer cell motility and invasion and its potential to be used as a prognostic marker for the aggressive forms of breast cancer. PMID:24158791

  7. Breast cancer phenotypes regulated by tissue factor-factor VII pathway: Possible therapeutic targets

    PubMed Central

    Koizume, Shiro; Miyagi, Yohei

    2014-01-01

    Breast cancer is a leading cause of cancer death in women, worldwide. Fortunately, breast cancer is relatively chemosensitive, with recent advances leading to the development of effective therapeutic strategies, significantly increasing disease cure rate. However, disease recurrence and treatment of cases lacking therapeutic molecular targets, such as epidermal growth factor receptor 2 and hormone receptors, referred to as triple-negative breast cancers, still pose major hurdles in the treatment of breast cancer. Thus, novel therapeutic approaches to treat aggressive breast cancers are essential. Blood coagulation factor VII (fVII) is produced in the liver and secreted into the blood stream. Tissue factor (TF), the cellular receptor for fVII, is an integral membrane protein that plays key roles in the extrinsic coagulation cascade. TF is overexpressed in breast cancer tissues. The TF-fVII complex may be formed in the absence of injury, because fVII potentially exists in the tissue fluid within cancer tissues. The active form of this complex (TF-fVIIa) may stimulate the expression of numerous malignant phenotypes in breast cancer cells. Thus, the TF-fVII pathway is a potentially attractive target for breast cancer treatment. To date, a number of studies investigating the mechanisms by which TF-fVII signaling contributes to breast cancer progression, have been conducted. In this review, we summarize the mechanisms controlling TF and fVII synthesis and regulation in breast cancer cells. Our current understanding of the TF-fVII pathway as a mediator of breast cancer progression will be also described. Finally, we will discuss how this knowledge can be applied to the design of future therapeutic strategies. PMID:25493229

  8. Induction of mitochondrial alternative oxidase in response to a cell signal pathway down-regulating the cytochrome pathway prevents programmed cell death.

    PubMed

    Vanlerberghe, Greg C; Robson, Christine A; Yip, Justine Y H

    2002-08-01

    Treatment of tobacco (Nicotiana tabacum L. cv Petit Havana SR1) cells with cysteine (Cys) triggers a signal pathway culminating in a large loss of mitochondrial cytochrome (cyt) pathway capacity. This down-regulation of the cyt path likely requires events outside the mitochondrion and is effectively blocked by cantharidin or endothall, indicating that protein dephosphorylation is one critical process involved. Generation of reactive oxygen species, cytosolic protein synthesis, and Ca(2+) flux from organelles also appear to be involved. Accompanying the loss of cyt path is a large induction of alternative oxidase (AOX) protein and capacity. Induction of AOX allows the cells to maintain high rates of respiration, indicating that the lesion triggered by Cys is in the cyt path downstream of ubiquinone. Consistent with this, transgenic (AS8) cells unable to induce AOX (due to the presence of an antisense transgene) lose all respiratory capacity upon Cys treatment. This initiates in AS8 a programmed cell death pathway, as evidenced by the accumulation of oligonucleosomal fragments of DNA as the culture dies. Alternatively, wild-type cells remain viable and eventually recover their cyt path. Induction of AOX in response to a chemical inhibition of the cyt path (by antimycin A) is also dependent upon protein dephosphorylation and the generation of reactive oxygen species. Common events required for both down-regulation of the cyt path and induction of AOX may represent a mechanism to coordinate the biogenesis of these two electron transport paths. Such coordinate regulation may be necessary, not only to satisfy metabolic demands, but also to modulate the initiation of a programmed cell death pathway responsive to mitochondrial respiratory status.

  9. mir-30d Regulates multiple genes in the autophagy pathway and impairs autophagy process in human cancer cells

    SciTech Connect

    Yang, Xiaojun; Zhong, Xiaomin; Tanyi, Janos L.; Shen, Jianfeng; Xu, Congjian; Gao, Peng; Zheng, Tim M.; DeMichele, Angela; Zhang, Lin

    2013-02-15

    Highlights: ► Gene set enrichment analysis indicated mir-30d might regulate the autophagy pathway. ► mir-30d represses the expression of BECN1, BNIP3L, ATG12, ATG5 and ATG2. ► BECN1, BNIP3L, ATG12, ATG5 and ATG2 are direct targets of mir-30d. ► mir-30d inhibits autophagosome formation and LC3B-I conversion to LC3B-II. ► mir-30d regulates the autophagy process. -- Abstract: In human epithelial cancers, the microRNA (miRNA) mir-30d is amplified with high frequency and serves as a critical oncomir by regulating metastasis, apoptosis, proliferation, and differentiation. Autophagy, a degradation pathway for long-lived protein and organelles, regulates the survival and death of many cell types. Increasing evidence suggests that autophagy plays an important function in epithelial tumor initiation and progression. Using a combined bioinformatics approach, gene set enrichment analysis, and miRNA target prediction, we found that mir-30d might regulate multiple genes in the autophagy pathway including BECN1, BNIP3L, ATG12, ATG5, and ATG2. Our further functional experiments demonstrated that the expression of these core proteins in the autophagy pathway was directly suppressed by mir-30d in cancer cells. Finally, we showed that mir-30d regulated the autophagy process by inhibiting autophagosome formation and LC3B-I conversion to LC3B-II. Taken together, our results provide evidence that the oncomir mir-30d impairs the autophagy process by targeting multiple genes in the autophagy pathway. This result will contribute to understanding the molecular mechanism of mir-30d in tumorigenesis and developing novel cancer therapy strategy.

  10. Modulatory roles of microRNAs in the regulation of different signalling pathways in large bowel cancer stem cells.

    PubMed

    Mamoori, Afraa; Gopalan, Vinod; Smith, Robert Anthony; Lam, Alfred King-Yin

    2016-03-01

    There are emerging data to suggest that microRNAs (miRNAs) have significant roles in regulating the function of normal cells and cancer stem cells (CSCs). This review aims to analyse the roles of miRNAs in the regulation of colon CSCs through their interaction with various signalling pathways. Studies showed a large number of miRNAs that are reported to be deregulated in colon CSCs. However, few of the studies available were able to outline the function of miRNAs in colon CSCs and uncover their signalling pathways. From those miRNAs, which are better described, miR-21 followed by miR-34, miR-200 and miR-215 are the most reported miRNAs to have roles in colon CSC regulation. In particular, miRNAs have been reported to regulate the stemness features of colon CSCs mainly via Wnt/B-catenin and Notch signalling pathways. Additionally, miRNAs have been reported to act on processes involving CSCs through cell cycle regulation genes and epithelial-mesenchymal transition. The relative paucity of data available on the significance of miRNAs in CSCs means that new studies will be of great importance to determine their roles and to identify the signalling pathways through which they operate. Such studies may in future guide further research to target these genes for more effective cancer treatment. miRNAs were shown to regulate the function of cancer stem cells in large bowel cancer by targeting a few key signalling pathways in cells.

  11. The Cap1–claudin-4 regulatory pathway is important for renal chloride reabsorption and blood pressure regulation

    PubMed Central

    Gong, Yongfeng; Yu, Miao; Yang, Jing; Gonzales, Ernie; Perez, Ronaldo; Hou, Mingli; Tripathi, Piyush; Hering-Smith, Kathleen S.; Hamm, L. Lee; Hou, Jianghui

    2014-01-01

    The paracellular pathway through the tight junction provides an important route for transepithelial chloride reabsorption in the kidney, which regulates extracellular salt content and blood pressure. Defects in paracellular chloride reabsorption may in theory cause deregulation of blood pressure. However, there is no evidence to prove this theory or to demonstrate the in vivo role of the paracellular pathway in renal chloride handling. Here, using a tissue-specific KO approach, we have revealed a chloride transport pathway in the kidney that requires the tight junction molecule claudin-4. The collecting duct-specific claudin-4 KO animals developed hypotension, hypochloremia, and metabolic alkalosis due to profound renal wasting of chloride. The claudin-4–mediated chloride conductance can be regulated endogenously by a protease—channel-activating protease 1 (cap1). Mechanistically, cap1 regulates claudin-4 intercellular interaction and membrane stability. A putative cap1 cleavage site has been identified in the second extracellular loop of claudin-4, mutation of which abolished its regulation by cap1. The cap1 effects on paracellular chloride permeation can be extended to other proteases such as trypsin, suggesting a general mechanism may also exist for proteases to regulate the tight junction permeabilities. Together, we have discovered a theory that paracellular chloride permeability is physiologically regulated and essential to renal salt homeostasis and blood pressure control. PMID:25157135

  12. Regulation of the degradative pathway enzymes coded for by the TOL plasmid (pWWO) from Pseudomonas putida mt-2.

    PubMed Central

    Worsey, M J; Franklin, F C; Williams, P A

    1978-01-01

    Pseudomonas putida mt-2 carries a plasmid (TOL, pWWO) which codes for a single set of enzymes responsible for the catabolism of toluene and m- and p-xylene to central metabolites by way of benzoate and m- and p-toluate, respectively, and subsequently by a meta cleavage pathway. Characterization of strains with mutations in structural genes of this pathway demonstrates that the inducers of the enzymes responsible for further degradation of m-toluate include m-xylene, m-methylbenzyl alcohol, and m-toluate, whereas the inducers of the enzymes responsible for oxidation of m-xylene to m-toluate include m-xylene and m-methylbenzyl alcohol but not m-toluate. A regulatory mutant is described in which m-xylene and m-methylbenzyl alcohol no longer induce any of the pathway enzymes, but m-toluate is still able to induce the enzymes responsible for its own degradation. Among revertants of this mutant are some strains in which all the enzymes are expressed constitutively and are not further induced by m-xylene. A model is proposed for the regulation of the pathway in which the enzymes are in two regulatory blocks, which are under the control of two regulator gene products. The model is essentially the same as proposed earlier for the regulation of the isofunctional pathway on the TOL20 plasmid from P. putida MT20. PMID:659369

  13. Regulation of the Wnt/β-Catenin Signaling Pathway by Human Papillomavirus E6 and E7 Oncoproteins

    PubMed Central

    Muñoz Bello, Jesus Omar; Olmedo Nieva, Leslie; Contreras Paredes, Adriana; Fuentes Gonzalez, Alma Mariana; Rocha Zavaleta, Leticia; Lizano, Marcela

    2015-01-01

    Cell signaling pathways are the mechanisms by which cells transduce external stimuli, which control the transcription of genes, to regulate diverse biological effects. In cancer, distinct signaling pathways, such as the Wnt/β-catenin pathway, have been implicated in the deregulation of critical molecular processes that affect cell proliferation and differentiation. For example, changes in β-catenin localization have been identified in Human Papillomavirus (HPV)-related cancers as the lesion progresses. Specifically, β-catenin relocates from the membrane/cytoplasm to the nucleus, suggesting that this transcription regulator participates in cervical carcinogenesis. The E6 and E7 oncoproteins are responsible for the transforming activity of HPV, and some studies have implicated these viral oncoproteins in the regulation of the Wnt/β-catenin pathway. Nevertheless, new interactions of HPV oncoproteins with cellular proteins are emerging, and the study of the biological effects of such interactions will help to understand HPV-related carcinogenesis. This review addresses the accumulated evidence of the involvement of the HPV E6 and E7 oncoproteins in the activation of the Wnt/β-catenin pathway. PMID:26295406

  14. Metabolomic Identification of a Novel Pathway of Blood Pressure Regulation Involving Hexadecanedioate

    PubMed Central

    Menni, Cristina; Graham, Delyth; Kastenmüller, Gabi; Alharbi, Nora H.J.; Alsanosi, Safaa Md; McBride, Martin; Mangino, Massimo; Titcombe, Philip; Shin, So-Youn; Psatha, Maria; Geisendorfer, Thomas; Huber, Anja; Peters, Annette; Wang-Sattler, Rui; Xu, Tao; Brosnan, Mary Julia; Trimmer, Jeff; Reichel, Christian; Mohney, Robert P.; Soranzo, Nicole; Edwards, Mark H.; Cooper, Cyrus; Church, Alistair C.; Suhre, Karsten; Gieger, Christian; Dominiczak, Anna F.; Spector, Tim D.; Valdes, Ana M.

    2015-01-01

    High blood pressure is a major contributor to the global burden of disease and discovering novel causal pathways of blood pressure regulation has been challenging. We tested blood pressure associations with 280 fasting blood metabolites in 3980 TwinsUK females. Survival analysis for all-cause mortality was performed on significant independent metabolites (P<8.9×10−5). Replication was conducted in 2 independent cohorts KORA (n=1494) and Hertfordshire (n=1515). Three independent animal experiments were performed to establish causality: (1) blood pressure change after increasing circulating metabolite levels in Wistar–Kyoto rats; (2) circulating metabolite change after salt-induced blood pressure elevation in spontaneously hypertensive stroke-prone rats; and (3) mesenteric artery response to noradrenaline and carbachol in metabolite treated and control rats. Of the15 metabolites that showed an independent significant association with blood pressure, only hexadecanedioate, a dicarboxylic acid, showed concordant association with blood pressure (systolic BP: β [95% confidence interval], 1.31 [0.83–1.78], P=6.81×10−8; diastolic BP: 0.81 [0.5–1.11], P=2.96×10−7) and mortality (hazard ratio [95% confidence interval], 1.49 [1.08–2.05]; P=0.02) in TwinsUK. The blood pressure association was replicated in KORA and Hertfordshire. In the animal experiments, we showed that oral hexadecanedioate increased both circulating hexadecanedioate and blood pressure in Wistar–Kyoto rats, whereas blood pressure elevation with oral sodium chloride in hypertensive rats did not affect hexadecanedioate levels. Vascular reactivity to noradrenaline was significantly increased in mesenteric resistance arteries from hexadecanedioate-treated rats compared with controls, indicated by the shift to the left of the concentration–response curve (P=0.013). Relaxation to carbachol did not show any difference. Our findings indicate that hexadecanedioate is causally associated with

  15. Regulation of the PI3K pathway through a p85α monomer–homodimer equilibrium | Office of Cancer Genomics

    Cancer.gov

    The canonical action of the p85α regulatory subunit of phosphatidylinositol 3-kinase (PI3K) is to associate with the p110α catalytic subunit to allow stimuli-dependent activation of the PI3K pathway. We elucidate a p110α-independent role of homodimerized p85α in the positive regulation of PTEN stability and activity.

  16. The master switch gene Sex-lethal promotes female development by negatively regulating the N signaling pathway

    PubMed Central

    Penn, Jill K M; Schedl, Paul

    2011-01-01

    Summary Notch (N) signaling is used for cell fate determination in many different developmental contexts. Here we show that the master control gene for sex determination in Drosophila melanogaster, Sex-lethal (Sxl), negatively regulates the N signaling pathway in females. In genetic assays, reducing Sxl activity suppresses the phenotypic effects of N mutations while increasing Sxl activity enhances the effects. Sxl appears to negatively regulate the pathway by reducing N protein accumulation and higher levels of N are found in Sxl−clones than in adjacent wild type cells. The inhibition of N expression does not depend on the known downstream targets of Sxl; however we find that Sxl protein can bind to N mRNAs. Finally our results indicate that downregulation of the N pathway by Sxl contributes to sex specific differences in morphology and suggest that it may also play an important role in follicle cell specification during oogenesis. PMID:17276344

  17. Induction of TNF-A and IL-1 in Human Tuberculosis (CIC3)

    DTIC Science & Technology

    1992-01-27

    AD-A259 308 MIPR NO: 90MM0581 TITLE: INDUCTION OF TNF-A AND IL-1 IN HUMAN TUBERCULOSIS (CIC3) PRINCIPAL INVESTIGATOk: Greg T. Anders CONTRACTING...DATES COVERED 1 27 January 19921 Final Report 4. TITLE AND SUBTITLE S. FUNDING NUMBERS Induction of TNF-A and IL-i in Human Tuberculosis MIPR No. (CIC3...SU•4ECT TERMS IS. NUMBER OF PAGES Interleukin-1; Tumor; Tuberculosis ; RAII; Volunteers 16. PRICE COO( 17. SECURITY CLASSIFICATION I$. SECURITY

  18. Transcriptomes of purified gastric ECL and parietal cells: identification of a novel pathway regulating acid secretion.

    PubMed

    Lambrecht, Nils W G; Yakubov, Iskandar; Zer, Cindy; Sachs, George

    2006-03-13

    The gastric entero-chromaffin-like (ECL) cell plays a key regulatory role in peripheral regulation of acid secretion due to the release of histamine that stimulates acid secretion by the parietal cell. Studies in intact animals, gastric glands, and isolated cells after short-term culture have shown expression of stimulatory CCK2 and PAC1 and inhibitory SST2 and Gal1 receptors as well as histidine decarboxylase. However, the pattern of its gene expression as a neuroendocrine cell has not been explored. Comparison of gene expression by 95% pure ECL cells obtained by density gradient, elutriation, and fluorescence-assisted cell sorting with isolates of the intact fundic gastric epithelium (i.e., "subtractive hybridization") identified a variety of additional expressed gene families characteristic of this neuroendocrine cell. These include genes 1) involved in neuropeptide synthesis and secretory vesicle exocytosis, 2) involved in control of inflammation, 3) implicated in healing of the epithelium, 4) encoding inhibitory Gi protein-coupled receptors, 5) playing a role in neuroendocrine regulation of food intake, and 6) encoding proteins likely involved in maintenance of circadian rhythm, in addition to the ECL cell-specific genes histidine decarboxylase and monoamine transporter. Particularly, the inhibitory apelin receptor gene, APJ, was highly expressed in the ECL cell preparation. Because parietal cells express apelin, immunohistochemical and functional studies showed that there is an inhibitory feed back loop between the parietal and ECL cell during gastrin stimulation, providing evidence for a novel pathway of downregulation of acid secretion due to interaction between these two cell types.

  19. Presenilin 1 regulates epidermal growth factor receptor turnover and signaling in the endosomal-lysosomal pathway.

    PubMed

    Repetto, Emanuela; Yoon, Il-Sang; Zheng, Hui; Kang, David E

    2007-10-26

    Mutations in the gene encoding presenilin 1 (PS1) cause the most aggressive form of early-onset familial Alzheimer disease. In addition to its well established role in Abeta production and Notch proteolysis, PS1 has been shown to mediate other physiological activities, such as regulation of the Wnt/beta-catenin signaling pathway, modulation of phosphatidylinositol 3-kinase/Akt and MEK/ERK signaling, and trafficking of select membrane proteins and/or intracellular vesicles. In this study, we present evidence that PS1 is a critical regulator of a key signaling receptor tyrosine kinase, epidermal growth factor receptor (EGFR). Specifically, EGFR levels were robustly increased in fibroblasts deficient in both PS1 and PS2 (PS(-/-)) due to delayed turnover of EGFR protein. Stable transfection of wild-type PS1 but not PS2 corrected EGFR to levels comparable to PS(+/+) cells, while FAD PS1 mutations showed partial loss of activity. The C-terminal fragment of PS1 was sufficient to fully reduce EGFR levels. In addition, the rapid ligand-induced degradation of EGFR was markedly delayed in PS(-/-) cells, resulting in prolonged signal activation. Despite the defective turnover of EGFR, ligand-induced autophosphorylation, ubiquitination, and endocytosis of EGFR were not affected by the lack of PS1. Instead, the trafficking of EGFR from early endosomes to lysosomes was severely delayed by PS1 deficiency. Elevation of EGFR was also seen in brains of adult mice conditionally ablated in PS1 and in skin tumors associated with the loss of PS1. These findings demonstrate a critical role of PS1 in the trafficking and turnover of EGFR and suggest potential pathogenic effects of elevated EGFR as well as perturbed endosomal-lysosomal trafficking in cell cycle control and Alzheimer disease.

  20. Regulation of Manganese Antioxidants by Nutrient Sensing Pathways in Saccharomyces cerevisiae

    PubMed Central

    Reddi, Amit R.; Culotta, Valeria C.

    2011-01-01

    In aerobic organisms, protection from oxidative damage involves the combined action of enzymatic and nonproteinaceous cellular factors that collectively remove harmful reactive oxygen species. One class of nonproteinaceous antioxidants includes small molecule complexes of manganese (Mn) that can scavenge superoxide anion radicals and provide a backup for superoxide dismutase enzymes. Such Mn antioxidants have been identified in diverse organisms; however, nothing regarding their physiology in the context of cellular adaptation to stress was known. Using a molecular genetic approach in Bakers’ yeast, Saccharomyces cerevisiae, we report that the Mn antioxidants can fall under control of the same pathways used for nutrient sensing and stress responses. Specifically, a serine/threonine PAS-kinase, Rim15p, that is known to integrate phosphate, nitrogen, and carbon sensing, can also control Mn antioxidant activity in yeast. Rim15p is negatively regulated by the phosphate-sensing kinase complex Pho80p/Pho85p and by the nitrogen-sensing Akt/S6 kinase homolog, Sch9p. We observed that loss of either of these upstream kinase sensors dramatically inhibited the potency of Mn as an antioxidant. Downstream of Rim15p are transcription factors Gis1p and the redundant Msn2/Msn4p pair that typically respond to nutrient and stress signals. Both transcription factors were found to modulate the potency of the Mn antioxidant but in opposing fashions: loss of Gis1p was seen to enhance Mn antioxidant activity whereas loss of Msn2/4p greatly suppressed it. Our observed roles for nutrient and stress response kinases and transcription factors in regulating the Mn antioxidant underscore its physiological importance in aerobic fitness. PMID:21926297

  1. Glutathione peroxidase genes in Arabidopsis are ubiquitous and regulated by abiotic stresses through diverse signaling pathways.

    PubMed

    Rodriguez Milla, Miguel A; Maurer, Alberto; Rodriguez Huete, Alicia; Gustafson, J Perry

    2003-12-01

    Glutathione peroxidases (GPXs) are a group of enzymes that protect cells against oxidative damage generated by reactive oxygen species (ROS). The presence of GPXs in plants has been reported by several groups, but the roles of individual members of this family in a single plant species have not been studied. A family of seven related proteins named AtGPX1- AtGPX7 in Arabidopsis was identified, and the genomic organization of this family was reported. The putative subcellular localizations of the encoded proteins are the cytosol, chloroplast, mitochondria, and endoplasmic reticulum. Expressed sequence tags (ESTs) for all the genes except AtGPX7 were identified. Expression analysis of AtGPX genes in Arabidopsis tissues was performed, and different patterns were detected. Interestingly, several genes were up-regulated coordinately in response to abiotic stresses. AtGPX6, like human phospholipid hydroperoxide GPX (PHGPX), possibly encodes mitochondrial and cytosolic isoforms by alternative initiation. In addition, this gene showed the strongest responses under most abiotic stresses tested. AtGPX6::GUS analysis in transgenic Arabidopsis showed that AtGPX6 is highly expressed throughout development in most tissues, thus supporting an important role for this gene in protection against oxidative damage. The different effects of salicylic acid (SA), jasmonic acid (JA), abscisic acid (ABA), and auxin on the expression of the genes indicate that the AtGPX family is regulated by multiple signaling pathways. Analysis of the upstream region of the AtGPX genes revealed the presence of multiple conserved motifs, and some of them resembled antioxidant-responsive elements found in plant and human promoters. The potential regulatory role of specific sequences is discussed.

  2. Nuclear interactor of ARF and Mdm2 regulates multiple pathways to activate p53

    PubMed Central

    Reed, Sara M; Hagen, Jussara; Tompkins, Van S; Thies, Katie; Quelle, Frederick W; Quelle, Dawn E

    2014-01-01

    The p53 tumor suppressor is controlled by an interactive network of factors that stimulate or inhibit its transcriptional activity. Within that network, Mdm2 functions as the major antagonist of p53 by promoting its ubiquitylation and degradation. Conversely, Tip60 activates p53 through direct association on target promoters as well as acetylation of p53 at lysine 120 (K120). This study examines the functional relationship between Mdm2 and Tip60 with a novel p53 regulator, NIAM (nuclear interactor of ARF and Mdm2). Previous work showed NIAM can suppress proliferation and activate p53 independently of ARF, indicating that other factors mediate those activities. Here, we demonstrate that NIAM is a chromatin-associated protein that binds Tip60. NIAM can promote p53 K120 acetylation, although that modification is not required for NIAM to inhibit proliferation or induce p53 transactivation of the p21 promoter. Notably, Tip60 silencing showed it contributes to but is not sufficient for NIAM-mediated p53 activation, suggesting other mechanisms are involved. Indeed, growth-inhibitory forms of NIAM also bind to Mdm2, and increased NIAM expression levels disrupt p53–Mdm2 association, inhibit p53 polyubiquitylation, and prevent Mdm2-mediated inhibition of p53 transcriptional activity. Importantly, loss of NIAM significantly impairs p53 activation. Together, these results show that NIAM activates p53 through multiple mechanisms involving Tip60 association and Mdm2 inhibition. Thus, NIAM regulates 2 critical pathways that control p53 function and are altered in human cancers, implying an important role for NIAM in tumorigenesis. PMID:24621507

  3. The Role of IRE-XBP1 Pathway in Regulation of Retinal Pigment Epithelium Tight Junctions

    PubMed Central

    Ma, Jacey H.; Wang, Joshua J.; Li, Junhua; Pfeffer, Bruce A.; Zhong, Yiming; Zhang, Sarah X.

    2016-01-01

    Purpose The retinal pigment epithelium (RPE) tight junctions play a pivotal role in maintaining the homeostatic environment of the neural retina. Herein, we investigated the role of X-box binding protein 1 (XBP1), an endoplasmic reticulum (ER) stress-responsive transcription factor, in regulation of RPE tight junctions. Methods Human RPE cell line (ARPE-19) and primary primate RPE cells were used for in vitro experiments and RPE-specific XBP1 knockout (KO) mice were used for in vivo study. Endoplasmic reticulum stress was induced by a sublethal dose of thapsigargin or tunicamycin. XBP1 activation was manipulated by IRE inhibitor 4μ8C, which suppresses XBP1 mRNA splicing. The integrity of tight junctions and the involvement of calcium-dependent RhoA/Rho kinase pathway were examined. Results Induction of ER stress by thapsigargin, but not tunicamycin, disrupted RPE tight junctions in ARPE-