A one-dimensional model of PCP signaling: polarized cell behavior in the notochord of the ascidian Ciona

PubMed Central

Kourakis, Matthew J.; Reeves, Wendy; Newman-Smith, Erin; Maury, Benoit; Abdul-Wajid, Sarah; Smith, William C.

2014-01-01

Despite its importance in development and physiology the planar cell polarity (PCP) pathway remains one of the most enigmatic signaling mechanisms. The notochord of the ascidian Ciona provides a unique model for investigating the PCP pathway. Interestingly, the notochord appears to be the only embryonic structure in Ciona activating the PCP pathway. Moreover, the Ciona notochord as a single-file array of forty polarized cells is a uniquely tractable system for the study of polarization dynamics and the transmission of the PCP pathway. Here, we test models for propagation of a polarizing signal, interrogating temporal, spatial and signaling requirements. A simple cell-cell relay cascading through the entire length of the notochord is not supported; instead a more complex mechanism is revealed, with interactions influencing polarity between neighboring cells, but not distant ones. Mechanisms coordinating notochord-wide polarity remain elusive, but appear to entrain general (i.e., global) polarity even while local interactions remain important. However, this global polarizer does not appear to act as a localized, spatially-restricted determinant. Coordination of polarity along the long axis of the notochord requires the PCP pathway, a role we demonstrate is temporally distinct from this pathway’s earlier role in convergent extension and intercalation. We also reveal polarity in the notochord to be dynamic: a cell’s polarity state can be changed and then restored, underscoring the Ciona notochord’s amenability for in vivo studies of PCP. PMID:25173874

MyD88 signaling in T cells directs IgA-mediated control of the microbiota to promote health

PubMed Central

Kubinak, Jason L.; Petersen, Charisse; Stephens, W. Zac; Soto, Ray; Bake, Erin; O’Connell, Ryan M.; Round, June L.

2015-01-01

SUMMARY Altered commensal communities are associated with human disease. IgA mediates intestinal homeostasis and regulates microbiota composition. Intestinal IgA is produced at high levels as a result of T follicular helper cell (TFH) and B cell interactions in germinal centers. However, the pathways directing host IgA responses towards the microbiota remain unknown. Here, we report that signaling through the innate adaptor MyD88 in gut T cells coordinates germinal center responses, including TFH and IgA+ B cell development. TFH development is deficient in germfree mice and can be restored by feeding TLR2 agonists that activate T cell intrinsic MyD88 signaling. Loss of this pathway diminishes high affinity IgA targeting of the microbiota and fails to control the bacterial community, leading to worsened disease. Our findings identify that T cells converge innate and adaptive immune signals to coordinate IgA against the microbiota, constraining microbial community membership to promote symbiosis. PMID:25620548

Infected cell protein 0 functional domains and their coordination in herpes simplex virus replication

PubMed Central

Gu, Haidong

2016-01-01

Herpes simplex virus 1 (HSV-1) is a ubiquitous human pathogen that establishes latent infection in ganglia neurons. Its unique life cycle requires a balanced “conquer and compromise” strategy to deal with the host anti-viral defenses. One of HSV-1 α (immediate early) gene products, infected cell protein 0 (ICP0), is a multifunctional protein that interacts with and modulates a wide range of cellular defensive pathways. These pathways may locate in different cell compartments, which then migrate or exchange factors upon stimulation, for the purpose of a concerted and effective defense. ICP0 is able to simultaneously attack multiple host pathways by either degrading key restrictive factors or modifying repressive complexes. This is a viral protein that contains an E3 ubiquitin ligase, translocates among different cell compartments and interacts with major defensive complexes. The multiple functional domains of ICP0 can work independently and at the same time coordinate with each other. Dissecting the functional domains of ICP0 and delineating the coordination of these domains will help us understand HSV-1 pathogenicity as well as host defense mechanisms. This article focuses on describing individual ICP0 domains, their biochemical properties and their implication in HSV-1 infection. By putting individual domain functions back into the picture of host anti-viral defense network, this review seeks to elaborate the complex interactions between HSV-1 and its host. PMID:26870669

RAP-1 and the RAL-1/exocyst pathway coordinate hypodermal cell organization in Caenorhabditis elegans

PubMed Central

Frische, Ester W; Pellis-van Berkel, Wendy; van Haaften, Gijs; Cuppen, Edwin; Plasterk, Ronald H A; Tijsterman, Marcel; Bos, Johannes L; Zwartkruis, Fried J T

2007-01-01

The small Ras-like GTPase Rap1 has been identified as a regulator of integrin activation and cadherin-mediated cell–cell contacts. Surprisingly, null mutants of RAP-1 in Caenorhabditis elegans are viable and fertile. In a synthetic lethal RNAi screen with C. elegans rap-1 mutants, the Ras-like GTPase ral-1 emerged as one of seven genes specifically required for viability. Depletion of exoc-8 and sec-5, encoding two putative RAL-1 effectors and members of the exocyst complex, also caused lethality of rap-1 mutants, but did not affect wild-type worms. The RAP-1 and the RAL-1/exocyst pathway appear to coordinate hypodermal cell movement and elongation during embryonic development. They mediate their effect in part through targeting the α-catenin homologue HMP-1 to the lateral membrane. Genetic interactions show that the RAP-1 and RAL-1/exocyst pathway also act in parallel during larval stages. Together these data provide in vivo evidence for the exocyst complex as a downstream RAL-1 effector in cell migration. PMID:17989692

mTOR Pathways in Cancer and Autophagy.

PubMed

Paquette, Mathieu; El-Houjeiri, Leeanna; Pause, Arnim

2018-01-12

TOR (target of rapamycin), an evolutionarily-conserved serine/threonine kinase, acts as a central regulator of cell growth, proliferation and survival in response to nutritional status, growth factor, and stress signals. It plays a crucial role in coordinating the balance between cell growth and cell death, depending on cellular conditions and needs. As such, TOR has been identified as a key modulator of autophagy for more than a decade, and several deregulations of this pathway have been implicated in a variety of pathological disorders, including cancer. At the molecular level, autophagy regulates several survival or death signaling pathways that may decide the fate of cancer cells; however, the relationship between autophagy pathways and cancer are still nascent. In this review, we discuss the recent cellular signaling pathways regulated by TOR, their interconnections to autophagy, and the clinical implications of TOR inhibitors in cancer.

Imaging burst kinetics and spatial coordination during serial killing by single natural killer cells

PubMed Central

Choi, Paul J.; Mitchison, Timothy J.

2013-01-01

Cytotoxic lymphocytes eliminate virus-infected and cancerous cells by immune recognition and killing through the perforin-granzyme pathway. Traditional killing assays measure average target cell lysis at fixed times and high effector:target ratios. Such assays obscure kinetic details that might reveal novel physiology. We engineered target cells to report on granzyme activity, used very low effector:target ratios to observe potential serial killing, and performed low magnification time-lapse imaging to reveal time-dependent statistics of natural killer (NK) killing at the single-cell level. Most kills occurred during serial killing, and a single NK cell killed up to 10 targets over a 6-h assay. The first kill was slower than subsequent kills, especially on poor targets, or when NK signaling pathways were partially inhibited. Spatial analysis showed that sequential kills were usually adjacent. We propose that NK cells integrate signals from the previous and current target, possibly by simultaneous contact. The resulting burst kinetics and spatial coordination may control the activity of NK cells in tissues. PMID:23576740

Stage-specific integration of maternal and embryonic peroxisome proliferator-activated receptor delta signaling is critical to pregnancy success.

PubMed

Wang, Haibin; Xie, Huirong; Sun, Xiaofei; Tranguch, Susanne; Zhang, Hao; Jia, Xiangxu; Wang, Dingzhi; Das, Sanjoy K; Desvergne, Béatrice; Wahli, Walter; DuBois, Raymond N; Dey, Sudhansu K

2007-12-28

Successful pregnancy depends on well coordinated developmental events involving both maternal and embryonic components. Although a host of signaling pathways participate in implantation, decidualization, and placentation, whether there is a common molecular link that coordinates these processes remains unknown. By exploiting genetic, molecular, pharmacological, and physiological approaches, we show here that the nuclear transcription factor peroxisome proliferator-activated receptor (PPAR) delta plays a central role at various stages of pregnancy, whereas maternal PPARdelta is critical to implantation and decidualization, and embryonic PPARdelta is vital for placentation. Using trophoblast stem cells, we further elucidate that a reciprocal relationship between PPARdelta-AKT and leukemia inhibitory factor-STAT3 signaling pathways serves as a cell lineage sensor to direct trophoblast cell fates during placentation. This novel finding of stage-specific integration of maternal and embryonic PPARdelta signaling provides evidence that PPARdelta is a molecular link that coordinates implantation, decidualization, and placentation crucial to pregnancy success. This study is clinically relevant because deferral of on time implantation leads to spontaneous pregnancy loss, and defective trophoblast invasion is one cause of preeclampsia in humans.

Coordination of NF-kappaB and NFAT antagonism by the forkhead transcription factor Foxd1.

PubMed

Lin, Ling; Peng, Stanford L

2006-04-15

Forkhead transcription factors play critical roles in the maintenance of immune homeostasis. In this study, we demonstrate that this regulation most likely involves intricate interactions between the forkhead family members and inflammatory transcription factors: the forkhead member Foxd1 coordinates the regulation of the activity of two key inflammatory transcription factors, NF-AT and NF-kappaB, with Foxd1 deficiency resulting in multiorgan, systemic inflammation, exaggerated Th cell-derived cytokine production, and T cell proliferation in autologous MLRs. Foxd1-deficient T cells possess increased activity of both NF-AT and NF-kappaB: the former correlates with the ability of Foxd1 to regulate casein kinase 1, an NF-AT inhibitory kinase; the latter with the ability of Foxd1 to regulate Foxj1, which regulates the NF-kappaB inhibitory subunit IkappaB beta. Thus, Foxd1 modulates inflammatory reactions and prevents autoimmunity by directly regulating anti-inflammatory regulators of the NF-AT pathway, and by coordinating the suppression of the NF-kappaB pathway via Foxj1. These findings indicate the presence of a general network of forkhead proteins that enforce T cell quiescence.

Cytotoxicity of Cyclometalated Platinum Complexes Based on Tridentate NCN and CNN-coordinating ligands: Remarkable Coordination Dependence

PubMed Central

Vezzu, Dileep A. k.; Lu, Qun; Chen, Yan-Hua; Huo, Shouquan

2014-01-01

A series of cyclometalated platinum complexes with diverse coordination patterns and geometries were screened for their anticancer activity. It was discovered that the NʌCʌN-coordinated platinum complex based on 1,3-di(pyridyl)benzene displayed much higher cytotoxicity against human lung cancer cells NCI-H522, HCC827, and NCI-H1299, and human prostate cancer cell RV1 than cisplatin. In a sharp contrast, the CʌNʌN-coordinated platinum complex based on 6-phenyl-2,2′-bipyridine was ineffective on these cancer cells. This remarkable difference in cytotoxicity displayed by NʌCʌN- and CʌNʌN-coordinated platinum complexes was related to the trans effect of the carbon donor in the cyclometalated platinum complexes, which played a crucial role in facilitating the dissociation of the chloride ligand to create an active binding site. The DNA binding was studied for the NʌCʌN-coordinated platinum complex using electrophoresis and emission titration. The cellular uptake observed by fluorescent microscope showed the complex is largely concentrated in the cytoplasm. The possible pathways for the cell apoptosis was studied by western blot analysis and the activation of PARP via caspase 7 was observed. PMID:24531534

Mechanical feedback coordinates cell wall expansion and assembly in yeast mating morphogenesis

PubMed Central

2018-01-01

The shaping of individual cells requires a tight coordination of cell mechanics and growth. However, it is unclear how information about the mechanical state of the wall is relayed to the molecular processes building it, thereby enabling the coordination of cell wall expansion and assembly during morphogenesis. Combining theoretical and experimental approaches, we show that a mechanical feedback coordinating cell wall assembly and expansion is essential to sustain mating projection growth in budding yeast (Saccharomyces cerevisiae). Our theoretical results indicate that the mechanical feedback provided by the Cell Wall Integrity pathway, with cell wall stress sensors Wsc1 and Mid2 increasingly activating membrane-localized cell wall synthases Fks1/2 upon faster cell wall expansion, stabilizes mating projection growth without affecting cell shape. Experimental perturbation of the osmotic pressure and cell wall mechanics, as well as compromising the mechanical feedback through genetic deletion of the stress sensors, leads to cellular phenotypes that support the theoretical predictions. Our results indicate that while the existence of mechanical feedback is essential to stabilize mating projection growth, the shape and size of the cell are insensitive to the feedback. PMID:29346368

The Hippo-YAP signaling pathway and contact inhibition of growth

PubMed Central

Gumbiner, Barry M.; Kim, Nam-Gyun

2014-01-01

ABSTRACT The Hippo-YAP pathway mediates the control of cell proliferation by contact inhibition as well as other attributes of the physical state of cells in tissues. Several mechanisms sense the spatial and physical organization of cells, and function through distinct upstream modules to stimulate Hippo-YAP signaling: adherens junction or cadherin–catenin complexes, epithelial polarity and tight junction complexes, the FAT-Dachsous morphogen pathway, as well as cell shape, actomyosin or mechanotransduction. Soluble extracellular factors also regulate Hippo pathway signaling, often inhibiting its activity. Indeed, the Hippo pathway mediates a reciprocal relationship between contact inhibition and mitogenic signaling. As a result, cells at the edges of a colony, a wound in a tissue or a tumor are more sensitive to ambient levels of growth factors and more likely to proliferate, migrate or differentiate through a YAP and/or TAZ-dependent process. Thus, the Hippo-YAP pathway senses and responds to the physical organization of cells in tissues and coordinates these physical cues with classic growth-factor-mediated signaling pathways. This Commentary is focused on the biological significance of Hippo-YAP signaling and how upstream regulatory modules of the pathway interact to produce biological outcomes. PMID:24532814

Organotypic three-dimensional culture model of mesenchymal and epithelial cells to examine tissue fusion events.

EPA Science Inventory

Tissue fusion during early mammalian development requires coordination of multiple cell types, the extracellular matrix, and complex signaling pathways. Fusion events during processes including heart development, neural tube closure, and palatal fusion are dependent on signaling ...

Drosophila TIEG Is a Modulator of Different Signalling Pathways Involved in Wing Patterning and Cell Proliferation

PubMed Central

Rodriguez, Isabel

2011-01-01

Acquisition of a final shape and size during organ development requires a regulated program of growth and patterning controlled by a complex genetic network of signalling molecules that must be coordinated to provide positional information to each cell within the corresponding organ or tissue. The mechanism by which all these signals are coordinated to yield a final response is not well understood. Here, I have characterized the Drosophila ortholog of the human TGF-β Inducible Early Gene 1 (dTIEG). TIEG are zinc-finger proteins that belong to the Krüppel-like factor (KLF) family and were initially identified in human osteoblasts and pancreatic tumor cells for the ability to enhance TGF-β response. Using the developing wing of Drosophila as “in vivo” model, the dTIEG function has been studied in the control of cell proliferation and patterning. These results show that dTIEG can modulate Dpp signalling. Furthermore, dTIEG also regulates the activity of JAK/STAT pathway suggesting a conserved role of TIEG proteins as positive regulators of TGF-β signalling and as mediators of the crosstalk between signalling pathways acting in a same cellular context. PMID:21494610

Pathway connectivity and signaling coordination in the yeast stress-activated signaling network

PubMed Central

Chasman, Deborah; Ho, Yi-Hsuan; Berry, David B; Nemec, Corey M; MacGilvray, Matthew E; Hose, James; Merrill, Anna E; Lee, M Violet; Will, Jessica L; Coon, Joshua J; Ansari, Aseem Z; Craven, Mark; Gasch, Audrey P

2014-01-01

Stressed cells coordinate a multi-faceted response spanning many levels of physiology. Yet knowledge of the complete stress-activated regulatory network as well as design principles for signal integration remains incomplete. We developed an experimental and computational approach to integrate available protein interaction data with gene fitness contributions, mutant transcriptome profiles, and phospho-proteome changes in cells responding to salt stress, to infer the salt-responsive signaling network in yeast. The inferred subnetwork presented many novel predictions by implicating new regulators, uncovering unrecognized crosstalk between known pathways, and pointing to previously unknown ‘hubs’ of signal integration. We exploited these predictions to show that Cdc14 phosphatase is a central hub in the network and that modification of RNA polymerase II coordinates induction of stress-defense genes with reduction of growth-related transcripts. We find that the orthologous human network is enriched for cancer-causing genes, underscoring the importance of the subnetwork's predictions in understanding stress biology. PMID:25411400

A complex regulatory network coordinating cell cycles during C. elegans development is revealed by a genome-wide RNAi screen.

PubMed

Roy, Sarah H; Tobin, David V; Memar, Nadin; Beltz, Eleanor; Holmen, Jenna; Clayton, Joseph E; Chiu, Daniel J; Young, Laura D; Green, Travis H; Lubin, Isabella; Liu, Yuying; Conradt, Barbara; Saito, R Mako

2014-02-28

The development and homeostasis of multicellular animals requires precise coordination of cell division and differentiation. We performed a genome-wide RNA interference screen in Caenorhabditis elegans to reveal the components of a regulatory network that promotes developmentally programmed cell-cycle quiescence. The 107 identified genes are predicted to constitute regulatory networks that are conserved among higher animals because almost half of the genes are represented by clear human orthologs. Using a series of mutant backgrounds to assess their genetic activities, the RNA interference clones displaying similar properties were clustered to establish potential regulatory relationships within the network. This approach uncovered four distinct genetic pathways controlling cell-cycle entry during intestinal organogenesis. The enhanced phenotypes observed for animals carrying compound mutations attest to the collaboration between distinct mechanisms to ensure strict developmental regulation of cell cycles. Moreover, we characterized ubc-25, a gene encoding an E2 ubiquitin-conjugating enzyme whose human ortholog, UBE2Q2, is deregulated in several cancers. Our genetic analyses suggested that ubc-25 acts in a linear pathway with cul-1/Cul1, in parallel to pathways employing cki-1/p27 and lin-35/pRb to promote cell-cycle quiescence. Further investigation of the potential regulatory mechanism demonstrated that ubc-25 activity negatively regulates CYE-1/cyclin E protein abundance in vivo. Together, our results show that the ubc-25-mediated pathway acts within a complex network that integrates the actions of multiple molecular mechanisms to control cell cycles during development. Copyright © 2014 Roy et al.

Functional Coordination of WAVE and WASP in C. elegans Neuroblast Migration.

PubMed

Zhu, Zhiwen; Chai, Yongping; Jiang, Yuxiang; Li, Wenjing; Hu, Huifang; Li, Wei; Wu, Jia-Wei; Wang, Zhi-Xin; Huang, Shanjin; Ou, Guangshuo

2016-10-24

Directional cell migration is critical for metazoan development. We define two molecular pathways that activate the Arp2/3 complex during neuroblast migration in Caenorhabditis elegans. The transmembrane protein MIG-13/Lrp12 is linked to the Arp2/3 nucleation-promoting factors WAVE or WASP through direct interactions with ABL-1 or SEM-5/Grb2, respectively. WAVE mutations partially impaired F-actin organization and decelerated cell migration, and WASP mutations did not inhibit cell migration but enhanced migration defects in WAVE-deficient cells. Purified SEM-5 and MIG-2 synergistically stimulated the F-actin branching activity of WASP-Arp2/3 in vitro. In GFP knockin animals, WAVE and WASP were largely organized into separate clusters at the leading edge, and the amount of WASP was less than WAVE but could be elevated by WAVE mutations. Our results indicate that the MIG-13-WAVE pathway provides the major force for directional cell motility, whereas MIG-13-WASP partially compensates for its loss, underscoring their coordinated activities in facilitating robust cell migration. Copyright © 2016 Elsevier Inc. All rights reserved.

The Hippo Pathway Targets Rae1 to Regulate Mitosis and Organ Size and to Feed Back to Regulate Upstream Components Merlin, Hippo, and Warts.

PubMed

Jahanshahi, Maryam; Hsiao, Kuangfu; Jenny, Andreas; Pfleger, Cathie M

2016-08-01

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.

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

Insulin receptor-mediated signaling via phospholipase C-γ regulates growth and differentiation in Drosophila.

PubMed

Murillo-Maldonado, Juan M; Zeineddine, Fouad Bou; Stock, Rachel; Thackeray, Justin; Riesgo-Escovar, Juan R

2011-01-01

Coordination between growth and patterning/differentiation is critical if appropriate final organ structure and size is to be achieved. Understanding how these two processes are regulated is therefore a fundamental and as yet incompletely answered question. Here we show through genetic analysis that the phospholipase C-γ (PLC-γ) encoded by small wing (sl) acts as such a link between growth and patterning/differentiation by modulating some MAPK outputs once activated by the insulin pathway; particularly, sl promotes growth and suppresses ectopic differentiation in the developing eye and wing, allowing cells to attain a normal size and differentiate properly. sl mutants have previously been shown to have a combination of both growth and patterning/differentiation phenotypes: small wings, ectopic wing veins, and extra R7 photoreceptor cells. We show here that PLC-γ activated by the insulin pathway participates broadly and positively during cell growth modulating EGF pathway activity, whereas in cell differentiation PLC-γ activated by the insulin receptor negatively regulates the EGF pathway. These roles require different SH2 domains of PLC-γ, and act via classic PLC-γ signaling and EGF ligand processing. By means of PLC-γ, the insulin receptor therefore modulates differentiation as well as growth. Overall, our results provide evidence that PLC-γ acts during development at a time when growth ends and differentiation begins, and is important for proper coordination of these two processes.

Coupling of solute transport and cell expansion in pea stems

NASA Technical Reports Server (NTRS)

Schmalstig, J. G.; Cosgrove, D. J.

1990-01-01

As cells expand and are displaced through the elongation zone of the epicotyl of etiolated pea (Pisum sativum L. var Alaska) seedlings, there is little net dilution of the cell sap, implying a coordination between cell expansion and solute uptake from the phloem. Using [14C] sucrose as a phloem tracer (applied to the hypogeous cotyledons), the pattern of label accumulation along the stem closely matched the growth rate pattern: high accumulation in the growing zone, little accumulation in nongrowing regions. Several results suggest that a major portion of phloem contents enters elongating cells through the symplast. We propose that the coordination between phloem transport and cell expansion is accomplished via regulatory pathways affecting both plasmodesmata conductivity and cell expansion.

Unraveling Interfaces between Energy Metabolism and Cell Cycle in Plants.

PubMed

Siqueira, João Antonio; Hardoim, Pablo; Ferreira, Paulo C G; Nunes-Nesi, Adriano; Hemerly, Adriana S

2018-06-19

Oscillation in energy levels is widely variable in dividing and differentiated cells. To synchronize cell proliferation and energy fluctuations, cell cycle-related proteins have been implicated in the regulation of mitochondrial energy-generating pathways in yeasts and animals. Plants have chloroplasts and mitochondria, coordinating the cell energy flow. Recent findings suggest an integrated regulation of these organelles and the nuclear cell cycle. Furthermore, reports indicate a set of interactions between the cell cycle and energy metabolism, coordinating the turnover of proteins in plants. Here, we discuss how cell cycle-related proteins directly interact with energy metabolism-related proteins to modulate energy homeostasis and cell cycle progression. We provide interfaces between cell cycle and energy metabolism-related proteins that could be explored to maximize plant yield. Copyright © 2018 Elsevier Ltd. All rights reserved.

NADPH Oxidase 1 Modulates WNT and NOTCH1 Signaling To Control the Fate of Proliferative Progenitor Cells in the Colon▿

PubMed Central

Coant, Nicolas; Ben Mkaddem, Sanae; Pedruzzi, Eric; Guichard, Cécile; Tréton, Xavier; Ducroc, Robert; Freund, Jean-Noel; Cazals-Hatem, Dominique; Bouhnik, Yoram; Woerther, Paul-Louis; Skurnik, David; Grodet, Alain; Fay, Michèle; Biard, Denis; Lesuffleur, Thécla; Deffert, Christine; Moreau, Richard; Groyer, André; Krause, Karl-Heinz; Daniel, Fanny; Ogier-Denis, Eric

2010-01-01

The homeostatic self-renewal of the colonic epithelium requires coordinated regulation of the canonical Wnt/β-catenin and Notch signaling pathways to control proliferation and lineage commitment of multipotent stem cells. However, the molecular mechanisms by which the Wnt/β-catenin and Notch1 pathways interplay in controlling cell proliferation and fate in the colon are poorly understood. Here we show that NADPH oxidase 1 (NOX1), a reactive oxygen species (ROS)-producing oxidase that is highly expressed in colonic epithelial cells, is a pivotal determinant of cell proliferation and fate that integrates Wnt/β-catenin and Notch1 signals. NOX1-deficient mice reveal a massive conversion of progenitor cells into postmitotic goblet cells at the cost of colonocytes due to the concerted repression of phosphatidylinositol 3-kinase (PI3K)/AKT/Wnt/β-catenin and Notch1 signaling. This conversion correlates with the following: (i) the redox-dependent activation of the dual phosphatase PTEN, causing the inactivation of the Wnt pathway effector β-catenin, and (ii) the downregulation of Notch1 signaling that provokes derepression of mouse atonal homolog 1 (Math1) expression. We conclude that NOX1 controls the balance between goblet and absorptive cell types in the colon by coordinately modulating PI3K/AKT/Wnt/β-catenin and Notch1 signaling. This finding provides the molecular basis for the role of NOX1 in cell proliferation and postmitotic differentiation. PMID:20351171

PAK1 is a breast cancer oncogene that coordinately activates MAPK and MET signaling

PubMed Central

Shrestha, Yashaswi; Schafer, Eric J.; Boehm, Jesse S.; Thomas, Sapana R.; He, Frank; Du, Jinyan; Wang, Shumei; Barretina, Jordi; Weir, Barbara A.; Zhao, Jean J.; Polyak, Kornelia; Golub, Todd R.; Beroukhim, Rameen; Hahn, William C.

2011-01-01

Activating mutations in the RAS family or BRAF frequently occur in many types of human cancers but are rarely detected in breast tumors. However, activation of the RAS-RAF-MEK-ERK Mitogen-Activated Protein Kinase (MAPK) pathway is commonly observed in human breast cancers, suggesting that other genetic alterations lead to activation of this signaling pathway. To identify breast cancer oncogenes that activate the MAPK pathway, we screened a library of human kinases for their ability to induce anchorage-independent growth in a derivative of immortalized human mammary epithelial cells (HMLE). We identified PAK1 as a kinase that permitted HMLE cells to form anchorage-independent colonies. PAK1 is amplified in several human cancer types, including 33% of breast tumor samples and cancer cell lines. The kinase activity of PAK1 is necessary for PAK1-induced transformation. Moreover, we show that PAK1 simultaneously activates MAPK and MET signaling; the latter via inhibition of Merlin. Disruption of these activities inhibits PAK1-driven anchorage-independent growth. These observations establish PAK1 amplification as an alternative mechanism for MAPK activation in human breast cancer and credential PAK1 as a breast cancer oncogene that coordinately regulates multiple signaling pathways, the cooperation of which leads to malignant transformation. PMID:22105362

PAK1 is a breast cancer oncogene that coordinately activates MAPK and MET signaling.

PubMed

Shrestha, Y; Schafer, E J; Boehm, J S; Thomas, S R; He, F; Du, J; Wang, S; Barretina, J; Weir, B A; Zhao, J J; Polyak, K; Golub, T R; Beroukhim, R; Hahn, W C

2012-07-19

Activating mutations in the RAS family or BRAF frequently occur in many types of human cancers but are rarely detected in breast tumors. However, activation of the RAS-RAF-MEK-ERK MAPK pathway is commonly observed in human breast cancers, suggesting that other genetic alterations lead to activation of this signaling pathway. To identify breast cancer oncogenes that activate the MAPK pathway, we screened a library of human kinases for their ability to induce anchorage-independent growth in a derivative of immortalized human mammary epithelial cells (HMLE). We identified p21-activated kinase 1 (PAK1) as a kinase that permitted HMLE cells to form anchorage-independent colonies. PAK1 is amplified in several human cancer types, including 30--33% of breast tumor samples and cancer cell lines. The kinase activity of PAK1 is necessary for PAK1-induced transformation. Moreover, we show that PAK1 simultaneously activates MAPK and MET signaling; the latter via inhibition of merlin. Disruption of these activities inhibits PAK1-driven anchorage-independent growth. These observations establish PAK1 amplification as an alternative mechanism for MAPK activation in human breast cancer and credential PAK1 as a breast cancer oncogene that coordinately regulates multiple signaling pathways, the cooperation of which leads to malignant transformation.

Cytotoxicity of cyclometalated platinum complexes based on tridentate NCN and CNN-coordinating ligands: remarkable coordination dependence.

PubMed

Vezzu, Dileep A K; Lu, Qun; Chen, Yan-Hua; Huo, Shouquan

2014-05-01

A series of cyclometalated platinum complexes with diverse coordination patterns and geometries were screened for their anticancer activity. It was discovered that the N^C^N-coordinated platinum complex based on 1,3-di(pyridyl)benzene displayed much higher cytotoxicity against human lung cancer cells NCI-H522, HCC827, and NCI-H1299, and human prostate cancer cell RV1 than cisplatin. In a sharp contrast, the C^N^N-coordinated platinum complex based on 6-phenyl-2,2'-bipyridine was ineffective on these cancer cells. This remarkable difference in cytotoxicity displayed by N^C^N- and C^N^N-coordinated platinum complexes was related to the trans effect of the carbon donor in the cyclometalated platinum complexes, which played a crucial role in facilitating the dissociation of the chloride ligand to create an active binding site. The DNA binding was studied for the N^C^N-coordinated platinum complex using electrophoresis and emission titration. The cellular uptake observed by fluorescent microscope showed that the complex is largely concentrated in the cytoplasm. The possible pathways for the cell apoptosis were studied by western blot analysis and the activation of PARP via caspase 7 was observed. Copyright © 2014 Elsevier Inc. All rights reserved.

A Kinome RNAi Screen in Drosophila Identifies Novel Genes Interacting with Lgl, aPKC, and Crb Cell Polarity Genes in Epithelial Tissues.

PubMed

Parsons, Linda M; Grzeschik, Nicola A; Amaratunga, Kasun; Burke, Peter; Quinn, Leonie M; Richardson, Helena E

2017-08-07

In both Drosophila melanogaster and mammalian systems, epithelial structure and underlying cell polarity are essential for proper tissue morphogenesis and organ growth. Cell polarity interfaces with multiple cellular processes that are regulated by the phosphorylation status of large protein networks. To gain insight into the molecular mechanisms that coordinate cell polarity with tissue growth, we screened a boutique collection of RNAi stocks targeting the kinome for their capacity to modify Drosophila "cell polarity" eye and wing phenotypes. Initially, we identified kinase or phosphatase genes whose depletion modified adult eye phenotypes associated with the manipulation of cell polarity complexes (via overexpression of Crb or aPKC). We next conducted a secondary screen to test whether these cell polarity modifiers altered tissue overgrowth associated with depletion of Lgl in the wing. These screens identified Hippo, Jun kinase (JNK), and Notch signaling pathways, previously linked to cell polarity regulation of tissue growth. Furthermore, novel pathways not previously connected to cell polarity regulation of tissue growth were identified, including Wingless (Wg/Wnt), Ras, and lipid/Phospho-inositol-3-kinase (PI3K) signaling pathways. Additionally, we demonstrated that the "nutrient sensing" kinases Salt Inducible Kinase 2 and 3 ( SIK2 and 3 ) are potent modifiers of cell polarity phenotypes and regulators of tissue growth. Overall, our screen has revealed novel cell polarity-interacting kinases and phosphatases that affect tissue growth, providing a platform for investigating molecular mechanisms coordinating cell polarity and tissue growth during development. Copyright © 2017 Parsons et al.

Regulators of homologous recombination repair as novel targets for cancer treatment

PubMed Central

Krajewska, Małgorzata; Fehrmann, Rudolf S. N.; de Vries, Elisabeth G. E.; van Vugt, Marcel A. T. M.

2015-01-01

To cope with DNA damage, cells possess a complex signaling network called the ‘DNA damage response’, which coordinates cell cycle control with DNA repair. The importance of this network is underscored by the cancer predisposition that frequently goes along with hereditary mutations in DNA repair genes. One especially important DNA repair pathway in this respect is homologous recombination (HR) repair. Defects in HR repair are observed in various cancers, including hereditary breast, and ovarian cancer. Intriguingly, tumor cells with defective HR repair show increased sensitivity to chemotherapeutic reagents, including platinum-containing agents. These observations suggest that HR-proficient tumor cells might be sensitized to chemotherapeutics if HR repair could be therapeutically inactivated. HR repair is an extensively regulated process, which depends strongly on the activity of various other pathways, including cell cycle pathways, protein-control pathways, and growth factor-activated receptor signaling pathways. In this review, we discuss how the mechanistic wiring of HR is controlled by cell-intrinsic or extracellular pathways. Furthermore, we have performed a meta-analysis on available genome-wide RNA interference studies to identify additional pathways that control HR repair. Finally, we discuss how these HR-regulatory pathways may provide therapeutic targets in the context of radio/chemosensitization. PMID:25852742

Dnmt1-dependent Chk1 pathway suppression is protective against neuron division.

PubMed

Oshikawa, Mio; Okada, Kei; Tabata, Hidenori; Nagata, Koh-Ichi; Ajioka, Itsuki

2017-09-15

Neuronal differentiation and cell-cycle exit are tightly coordinated, even in pathological situations. When pathological neurons re-enter the cell cycle and progress through the S phase, they undergo cell death instead of division. However, the mechanisms underlying mitotic resistance are mostly unknown. Here, we have found that acute inactivation of retinoblastoma (Rb) family proteins (Rb, p107 and p130) in mouse postmitotic neurons leads to cell death after S-phase progression. Checkpoint kinase 1 (Chk1) pathway activation during the S phase prevented the cell death, and allowed the division of cortical neurons that had undergone acute Rb family inactivation, oxygen-glucose deprivation (OGD) or in vivo hypoxia-ischemia. During neurogenesis, cortical neurons became protected from S-phase Chk1 pathway activation by the DNA methyltransferase Dnmt1, and underwent cell death after S-phase progression. Our results indicate that Chk1 pathway activation overrides mitotic safeguards and uncouples neuronal differentiation from mitotic resistance. © 2017. Published by The Company of Biologists Ltd.

Hippo pathway coactivators Yap and Taz are required to coordinate mammalian liver regeneration

PubMed Central

Lu, Li; Finegold, Milton J; Johnson, Randy L

2018-01-01

The mammalian liver has a remarkable capacity for repair following injury. Removal of up to two-third of liver mass results in a series of events that include extracellular matrix remodeling, coordinated hepatic cell cycle re-entry, restoration of liver mass and tissue remodeling to return the damaged liver to its normal state. Although there has been considerable advancement of our knowledge concerning the regenerative capacity of the mammalian liver, many outstanding questions remaining, such as: how does the regenerating liver stop proliferating when appropriate mass is restored and how do these mechanisms relate to normal regulation of organ size during development? Hippo pathway has been proposed to be central in mediating both events: organ size control during development and following regeneration. In this report, we examined the role of Yap and Taz, key components of the Hippo pathway in liver organ size regulation, both in the context of development and homeostasis. Our studies reveal that contrary to the current paradigms that Yap/Taz are not required for developmental regulation of liver size but are required for proper liver regeneration. In livers depleted of Yap and Taz, liver mass is elevated in neonates and adults. However, Yap/Taz-depleted livers exhibit profound defects in liver regeneration, including an inability to restore liver mass and to properly coordinate cell cycle entry. Taken together, our results highlight requirements for the Hippo pathway during liver regeneration and indicate that there are additional pathways that cooperate with Hippo signaling to control liver size during development and in the adult. PMID:29303509

Induction and repair of DNA double strand breaks: the increasing spectrum of non-homologous end joining pathways.

PubMed

Mladenov, Emil; Iliakis, George

2011-06-03

A defining characteristic of damage induced in the DNA by ionizing radiation (IR) is its clustered character that leads to the formation of complex lesions challenging the cellular repair mechanisms. The most widely investigated such complex lesion is the DNA double strand break (DSB). DSBs undermine chromatin stability and challenge the repair machinery because an intact template strand is lacking to assist restoration of integrity and sequence in the DNA molecule. Therefore, cells have evolved a sophisticated machinery to detect DSBs and coordinate a response on the basis of inputs from various sources. A central function of cellular responses to DSBs is the coordination of DSB repair. Two conceptually different mechanisms can in principle remove DSBs from the genome of cells of higher eukaryotes. Homologous recombination repair (HRR) uses as template a homologous DNA molecule and is therefore error-free; it functions preferentially in the S and G2 phases. Non-homologous end joining (NHEJ), on the other hand, simply restores DNA integrity by joining the two ends, is error prone as sequence is only fortuitously preserved and active throughout the cell cycle. The basis of DSB repair pathway choice remains unknown, but cells of higher eukaryotes appear programmed to utilize preferentially NHEJ. Recent work suggests that when the canonical DNA-PK dependent pathway of NHEJ (D-NHEJ), becomes compromised an alternative NHEJ pathway and not HRR substitutes in a quasi-backup function (B-NHEJ). Here, we outline aspects of DSB induction by IR and review the mechanisms of their processing in cells of higher eukaryotes. We place particular emphasis on backup pathways of NHEJ and summarize their increasing significance in various cellular processes, as well as their potential contribution to carcinogenesis. 2011 Elsevier B.V. All rights reserved.

Intrinsic, Functional, and Structural Properties of β-Thymosins and β-Thymosin/WH2 Domains in the Regulation and Coordination of Actin Self-Assembly Dynamics and Cytoskeleton Remodeling.

PubMed

Renault, L

2016-01-01

β-Thymosins are a family of heat-stable multifunctional polypeptides that are expressed as small proteins of about 5kDa (~45 amino acids) almost exclusively in multicellular animals. They were first isolated from the thymus. As full-length or truncated polypeptides, they appear to stimulate a broad range of extracellular activities in various signaling pathways, including tissue repair and regeneration, inflammation, cell migration, and immune defense. However, their cell surface receptors and structural mechanisms of regulations in these multiple pathways remain still poorly understood. Besides their extracellular activities, they belong to a larger family of small, intrinsically disordered actin-binding domains called WH2/β-thymosin domains that have been identified in more than 1800 multidomain proteins found in different taxonomic domains of life and involved in various actin-based motile processes including cell morphogenesis, motility, adhesions, tissue development, intracellular trafficking, or pathogen infections. This review briefly surveys the main recent findings to understand how these small, intrinsically disordered but functional domains can interact with many unrelated partners and can thus integrate and coordinate various intracellular activities in actin self-assembly dynamics and cell signaling pathways linked to their cytoskeleton remodeling. © 2016 Elsevier Inc. All rights reserved.

Osmotic Stress Signaling and Osmoadaptation in Yeasts

PubMed Central

Hohmann, Stefan

2002-01-01

The ability to adapt to altered availability of free water is a fundamental property of living cells. The principles underlying osmoadaptation are well conserved. The yeast Saccharomyces cerevisiae is an excellent model system with which to study the molecular biology and physiology of osmoadaptation. Upon a shift to high osmolarity, yeast cells rapidly stimulate a mitogen-activated protein (MAP) kinase cascade, the high-osmolarity glycerol (HOG) pathway, which orchestrates part of the transcriptional response. The dynamic operation of the HOG pathway has been well studied, and similar osmosensing pathways exist in other eukaryotes. Protein kinase A, which seems to mediate a response to diverse stress conditions, is also involved in the transcriptional response program. Expression changes after a shift to high osmolarity aim at adjusting metabolism and the production of cellular protectants. Accumulation of the osmolyte glycerol, which is also controlled by altering transmembrane glycerol transport, is of central importance. Upon a shift from high to low osmolarity, yeast cells stimulate a different MAP kinase cascade, the cell integrity pathway. The transcriptional program upon hypo-osmotic shock seems to aim at adjusting cell surface properties. Rapid export of glycerol is an important event in adaptation to low osmolarity. Osmoadaptation, adjustment of cell surface properties, and the control of cell morphogenesis, growth, and proliferation are highly coordinated processes. The Skn7p response regulator may be involved in coordinating these events. An integrated understanding of osmoadaptation requires not only knowledge of the function of many uncharacterized genes but also further insight into the time line of events, their interdependence, their dynamics, and their spatial organization as well as the importance of subtle effects. PMID:12040128

Plant TOR signaling components

PubMed Central

John, Florian; Roffler, Stefan; Wicker, Thomas; Ringli, Christoph

2011-01-01

Cell growth is a process that needs to be tightly regulated. Cells must be able to sense environmental factors like nutrient abundance, the energy level or stress signals and coordinate growth accordingly. The Target Of Rapamycin (TOR) pathway is a major controller of growth-related processes in all eukaryotes. If environmental conditions are favorable, the TOR pathway promotes cell and organ growth and restrains catabolic processes like autophagy. Rapamycin is a specific inhibitor of the TOR kinase and acts as a potent inhibitor of TOR signaling. As a consequence, interfering with TOR signaling has a strong impact on plant development. This review summarizes the progress in the understanding of the biological significance and the functional analysis of the TOR pathway in plants. PMID:22057328

Coordination of receptor signaling in multiple hematopoietic cell lineages by the adaptor protein SLP-76.

PubMed

Jordan, Martha S; Koretzky, Gary A

2010-04-01

The adaptor protein SLP-76 is expressed in multiple hematopoietic lineages including T cells, platelets, and neutrophils. SLP-76 mediated signaling is dependent on its multiple protein interaction domains, as it creates a scaffold on which key signaling complexes are built. SLP-76 is critical for supporting signaling downstream of both immunoreceptors and integrins. The signaling molecules used both upstream and downstream of SLP-76 are similar among these receptors and across cell types; however, important differences exist. Appreciating how SLP-76 coordinates signal transduction across different cell and receptor types provides insights into the complex interplay of pathways critical for activation of cells of the immune system that are essential for host defense.

Linking the Primary Cilium to Cell Migration in Tissue Repair and Brain Development

PubMed Central

Veland, Iben Rønn; Lindbæk, Louise; Christensen, Søren Tvorup

2014-01-01

Primary cilia are unique sensory organelles that coordinate cellular signaling networks in vertebrates. Inevitably, defects in the formation or function of primary cilia lead to imbalanced regulation of cellular processes that causes multisystemic disorders and diseases, commonly known as ciliopathies. Mounting evidence has demonstrated that primary cilia coordinate multiple activities that are required for cell migration, which, when they are aberrantly regulated, lead to defects in organogenesis and tissue repair, as well as metastasis of tumors. Here, we present an overview on how primary cilia may contribute to the regulation of the cellular signaling pathways that control cyclic processes in directional cell migration. PMID:26955067

Premetazoan origin of the Hippo signaling pathway

PubMed Central

Sebé-Pedrós, Arnau; Zheng, Yonggang; Ruiz-Trillo, Iñaki; Pan, Duojia

2012-01-01

Summary Non-aggregative multicellularity requires strict control of cell number. The Hippo signaling pathway coordinates cell proliferation and apoptosis and is a central regulator of organ size in animals. Recent studies have shown the presence of key members of the Hippo pathway in non-bilaterian animals, but failed to identify this pathway outside Metazoa. Through comparative analyses of recently sequenced holozoan genomes, we show that Hippo pathway components, such as the kinases Hippo and Warts, the co-activator Yorkie and the transcription factor Scalloped, were already present in the unicellular ancestors of animals. Remarkably, functional analysis of Hippo components of the amoeboid holozoan Capsaspora owczarzaki, performed in Drosophila, demonstrate that the growth-regulatory activity of the Hippo pathway is conserved in this unicellular lineage. Our findings show that the Hippo pathway evolved well before the origin of Metazoa and highlight the importance of Hippo signaling as a key developmental mechanism pre-dating the origin of Metazoa. PMID:22832104

The pathway to femaleness: current knowledge on embryonic development of the ovary

PubMed Central

Yao, Humphrey Hung-Chang

2014-01-01

Increasing evidence indicates that organogenesis of the ovary is not a passive process arising by default in the absence of the testis pathway. A coordinated interaction is actually in force between somatic cells and female germ cells in embryonic ovaries, thus creating a unique microenvironment that facilitates the formation of follicles. Identification of the functional roles of several novel regulatory elements such as Figα, Foxl2, follistatin, and Wnt4 reveals the complexity of early ovarian organization. Challenges await us to establish the molecular connections of these molecules as well as to discover new candidates in the pathway of early ovarian development. PMID:15664455

Interactions of C4 subtype metabolic activities and transport in maize are revealed through the characterization of DCT2 mutants

USDA-ARS?s Scientific Manuscript database

C4 photosynthesis is an elaborate set of metabolic pathways that utilize specialized anatomical and biochemical adaptations to concentrate CO2 around RuBisCO. The activities of the C4 pathways are coordinated between two specialized leaf cell types, mesophyll (M) and bundle sheath (BS), and rely hea...

Accumulation of specific sterol precursors targets a MAP kinase cascade mediating cell-cell recognition and fusion.

PubMed

Weichert, Martin; Lichius, Alexander; Priegnitz, Bert-Ewald; Brandt, Ulrike; Gottschalk, Johannes; Nawrath, Thorben; Groenhagen, Ulrike; Read, Nick D; Schulz, Stefan; Fleißner, André

2016-10-18

Sterols are vital components of eukaryotic cell membranes. Defects in sterol biosynthesis, which result in the accumulation of precursor molecules, are commonly associated with cellular disorders and disease. However, the effects of these sterol precursors on the metabolism, signaling, and behavior of cells are only poorly understood. In this study, we show that the accumulation of only ergosterol precursors with a conjugated double bond in their aliphatic side chain specifically disrupts cell-cell communication and fusion in the fungus Neurospora crassa Genetically identical germinating spores of this fungus undergo cell-cell fusion, thereby forming a highly interconnected supracellular network during colony initiation. Before fusion, the cells use an unusual signaling mechanism that involves the coordinated and alternating switching between signal sending and receiving states of the two fusion partners. Accumulation of only ergosterol precursors with a conjugated double bond in their aliphatic side chain disrupts this coordinated cell-cell communication and suppresses cell fusion. These specific sterol precursors target a single ERK-like mitogen-activated protein (MAP) kinase (MAK-1)-signaling cascade, whereas a second MAP kinase pathway (MAK-2), which is also involved in cell fusion, is unaffected. These observations indicate that a minor specific change in sterol structure can exert a strong detrimental effect on a key signaling pathway of the cell, resulting in the absence of cell fusion.

Planar cell polarity controls directional Notch signaling in the Drosophila leg

PubMed Central

Capilla, Amalia; Johnson, Ruth; Daniels, Maki; Benavente, María; Bray, Sarah J.; Galindo, Máximo Ibo

2012-01-01

The generation of functional structures during development requires tight spatial regulation of signaling pathways. Thus, in Drosophila legs, in which Notch pathway activity is required to specify joints, only cells distal to ligand-producing cells are capable of responding. Here, we show that the asymmetric distribution of planar cell polarity (PCP) proteins correlates with this spatial restriction of Notch activation. Frizzled and Dishevelled are enriched at distal sides of each cell and hence localize at the interface with ligand-expressing cells in the non-responding cells. Elimination of PCP gene function in cells proximal to ligand-expressing cells is sufficient to alleviate the repression, resulting in ectopic Notch activity and ectopic joint formation. Mutations that compromise a direct interaction between Dishevelled and Notch reduce the efficacy of repression. Likewise, increased Rab5 levels or dominant-negative Deltex can suppress the ectopic joints. Together, these results suggest that PCP coordinates the spatial activity of the Notch pathway by regulating endocytic trafficking of the receptor. PMID:22736244

The APC/C Coordinates Retinal Differentiation with G1 Arrest through the Nek2-Dependent Modulation of Wingless Signaling.

PubMed

Martins, Torcato; Meghini, Francesco; Florio, Francesca; Kimata, Yuu

2017-01-09

The cell cycle is coordinated with differentiation during animal development. Here we report a cell-cycle-independent developmental role for a master cell-cycle regulator, the anaphase-promoting complex or cyclosome (APC/C), in the regulation of cell fate through modulation of Wingless (Wg) signaling. The APC/C controls both cell-cycle progression and postmitotic processes through ubiquitin-dependent proteolysis. Through an RNAi screen in the developing Drosophila eye, we found that partial APC/C inactivation severely inhibits retinal differentiation independently of cell-cycle defects. The differentiation inhibition coincides with hyperactivation of Wg signaling caused by the accumulation of a Wg modulator, Drosophila Nek2 (dNek2). The APC/C degrades dNek2 upon synchronous G1 arrest prior to differentiation, which allows retinal differentiation through local suppression of Wg signaling. We also provide evidence that decapentaplegic signaling may posttranslationally regulate this APC/C function. Thus, the APC/C coordinates cell-fate determination with the cell cycle through the modulation of developmental signaling pathways. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

The Regulation of Filamentous Growth in Yeast

PubMed Central

Cullen, Paul J.; Sprague, George F.

2012-01-01

Filamentous growth is a nutrient-regulated growth response that occurs in many fungal species. In pathogens, filamentous growth is critical for host–cell attachment, invasion into tissues, and virulence. The budding yeast Saccharomyces cerevisiae undergoes filamentous growth, which provides a genetically tractable system to study the molecular basis of the response. Filamentous growth is regulated by evolutionarily conserved signaling pathways. One of these pathways is a mitogen activated protein kinase (MAPK) pathway. A remarkable feature of the filamentous growth MAPK pathway is that it is composed of factors that also function in other pathways. An intriguing challenge therefore has been to understand how pathways that share components establish and maintain their identity. Other canonical signaling pathways—rat sarcoma/protein kinase A (RAS/PKA), sucrose nonfermentable (SNF), and target of rapamycin (TOR)—also regulate filamentous growth, which raises the question of how signals from multiple pathways become integrated into a coordinated response. Together, these pathways regulate cell differentiation to the filamentous type, which is characterized by changes in cell adhesion, cell polarity, and cell shape. How these changes are accomplished is also discussed. High-throughput genomics approaches have recently uncovered new connections to filamentous growth regulation. These connections suggest that filamentous growth is a more complex and globally regulated behavior than is currently appreciated, which may help to pave the way for future investigations into this eukaryotic cell differentiation behavior. PMID:22219507

Loss of Gα12/13 exacerbates apical area dependence of actomyosin contractility

PubMed Central

Xie, Shicong; Mason, Frank M.; Martin, Adam C.

2016-01-01

During development, coordinated cell shape changes alter tissue shape. In the Drosophila ventral furrow and other epithelia, apical constriction of hundreds of epithelial cells folds the tissue. Genes in the Gα12/13 pathway coordinate collective apical constriction, but the mechanism of coordination is poorly understood. Coupling live-cell imaging with a computational approach to identify contractile events, we discovered that differences in constriction behavior are biased by initial cell shape. Disrupting Gα12/13 exacerbates this relationship. Larger apical area is associated with delayed initiation of contractile pulses, lower apical E-cadherin and F-actin levels, and aberrantly mobile Rho-kinase structures. Our results suggest that loss of Gα12/13 disrupts apical actin cortex organization and pulse initiation in a size-dependent manner. We propose that Gα12/13 robustly organizes the apical cortex despite variation in apical area to ensure the timely initiation of contractile pulses in a tissue with heterogeneity in starting cell shape. PMID:27489340

Arabidopsis AIP1-2 restricted by WER-mediated patterning modulates planar polarity

PubMed Central

Kiefer, Christian S.; Claes, Andrea R.; Nzayisenga, Jean-Claude; Pietra, Stefano; Stanislas, Thomas; Hüser, Anke; Ikeda, Yoshihisa; Grebe, Markus

2015-01-01

The coordination of cell polarity within the plane of the tissue layer (planar polarity) is crucial for the development of diverse multicellular organisms. Small Rac/Rho-family GTPases and the actin cytoskeleton contribute to planar polarity formation at sites of polarity establishment in animals and plants. Yet, upstream pathways coordinating planar polarity differ strikingly between kingdoms. In the root of Arabidopsis thaliana, a concentration gradient of the phytohormone auxin coordinates polar recruitment of Rho-of-plant (ROP) to sites of polar epidermal hair initiation. However, little is known about cytoskeletal components and interactions that contribute to this planar polarity or about their relation to the patterning machinery. Here, we show that ACTIN7 (ACT7) represents a main actin isoform required for planar polarity of root hair positioning, interacting with the negative modulator ACTIN-INTERACTING PROTEIN1-2 (AIP1-2). ACT7, AIP1-2 and their genetic interaction are required for coordinated planar polarity of ROP downstream of ethylene signalling. Strikingly, AIP1-2 displays hair cell file-enriched expression, restricted by WEREWOLF (WER)-dependent patterning and modified by ethylene and auxin action. Hence, our findings reveal AIP1-2, expressed under control of the WER-dependent patterning machinery and the ethylene signalling pathway, as a modulator of actin-mediated planar polarity. PMID:25428588

Structural basis for leucine sensing by the Sestrin2-mTORC1 pathway

DOE PAGES

Saxton, Robert A.; Knockenhauer, Kevin E.; Wolfson, Rachel L.; ...

2015-11-19

Eukaryotic cells coordinate growth with the availability of nutrients through mTOR complex 1 (mTORC1), a master growth regulator. Leucine is of particular importance and activates mTORC1 via the Rag GTPases and their regulators GATOR1 and GATOR2. Sestrin2 interacts with GATOR2 and is a leucine sensor. We present the 2.7-Å crystal structure of Sestrin2 in complex with leucine. Leucine binds through a single pocket that coordinates its charged functional groups and confers specificity for the hydrophobic side chain. A loop encloses leucine and forms a lid-latch mechanism required for binding. A structure-guided mutation in Sestrin2 that decreases its affinity for leucinemore » leads to a concomitant increase in the leucine concentration required for mTORC1 activation in cells. Lastly, these results provide a structural mechanism of amino acid sensing by the mTORC1 pathway.« less

Structural basis for leucine sensing by the Sestrin2-mTORC1 pathway

PubMed Central

Saxton, Robert A.; Knockenhauer, Kevin E.; Wolfson, Rachel L.; Chantranupong, Lynne; Pacold, Michael E.; Wang, Tim; Schwartz, Thomas U.; Sabatini, David M.

2015-01-01

Eukaryotic cells coordinate growth with the availability of nutrients through mTOR complex 1 (mTORC1), a master growth regulator. Leucine is of particular importance and activates mTORC1 via the Rag GTPases and their regulators GATOR1 and GATOR2. Sestrin2 interacts with GATOR2 and is a leucine sensor. We present the 2.7-Å crystal structure of Sestrin2 in complex with leucine. Leucine binds through a single pocket that coordinates its charged functional groups and confers specificity for the hydrophobic side chain. A loop encloses leucine and forms a lid-latch mechanism required for binding. A structure-guided mutation in Sestrin2 that decreases its affinity for leucine leads to a concomitant increase in the leucine concentration required for mTORC1 activation in cells. These results provide a structural mechanism of amino acid sensing by the mTORC1 pathway. PMID:26586190

Enhancer of polycomb coordinates multiple signaling pathways to promote both cyst and germline stem cell differentiation in the Drosophila adult testis

PubMed Central

Feng, Lijuan; Shi, Zhen; Chen, Xin

2017-01-01

Stem cells reside in a particular microenvironment known as a niche. The interaction between extrinsic cues originating from the niche and intrinsic factors in stem cells determines their identity and activity. Maintenance of stem cell identity and stem cell self-renewal are known to be controlled by chromatin factors. Herein, we use the Drosophila adult testis which has two adult stem cell lineages, the germline stem cell (GSC) lineage and the cyst stem cell (CySC) lineage, to study how chromatin factors regulate stem cell differentiation. We find that the chromatin factor Enhancer of Polycomb [E(Pc)] acts in the CySC lineage to negatively control transcription of genes associated with multiple signaling pathways, including JAK-STAT and EGF, to promote cellular differentiation in the CySC lineage. E(Pc) also has a non-cell-autonomous role in regulating GSC lineage differentiation. When E(Pc) is specifically inactivated in the CySC lineage, defects occur in both germ cell differentiation and maintenance of germline identity. Furthermore, compromising Tip60 histone acetyltransferase activity in the CySC lineage recapitulates loss-of-function phenotypes of E(Pc), suggesting that Tip60 and E(Pc) act together, consistent with published biochemical data. In summary, our results demonstrate that E(Pc) plays a central role in coordinating differentiation between the two adult stem cell lineages in Drosophila testes. PMID:28196077

Accumulation of specific sterol precursors targets a MAP kinase cascade mediating cell–cell recognition and fusion

PubMed Central

Weichert, Martin; Lichius, Alexander; Priegnitz, Bert-Ewald; Brandt, Ulrike; Gottschalk, Johannes; Nawrath, Thorben; Groenhagen, Ulrike; Read, Nick D.; Schulz, Stefan; Fleißner, André

2016-01-01

Sterols are vital components of eukaryotic cell membranes. Defects in sterol biosynthesis, which result in the accumulation of precursor molecules, are commonly associated with cellular disorders and disease. However, the effects of these sterol precursors on the metabolism, signaling, and behavior of cells are only poorly understood. In this study, we show that the accumulation of only ergosterol precursors with a conjugated double bond in their aliphatic side chain specifically disrupts cell–cell communication and fusion in the fungus Neurospora crassa. Genetically identical germinating spores of this fungus undergo cell–cell fusion, thereby forming a highly interconnected supracellular network during colony initiation. Before fusion, the cells use an unusual signaling mechanism that involves the coordinated and alternating switching between signal sending and receiving states of the two fusion partners. Accumulation of only ergosterol precursors with a conjugated double bond in their aliphatic side chain disrupts this coordinated cell–cell communication and suppresses cell fusion. These specific sterol precursors target a single ERK-like mitogen-activated protein (MAP) kinase (MAK-1)-signaling cascade, whereas a second MAP kinase pathway (MAK-2), which is also involved in cell fusion, is unaffected. These observations indicate that a minor specific change in sterol structure can exert a strong detrimental effect on a key signaling pathway of the cell, resulting in the absence of cell fusion. PMID:27708165

Intracellular coordination of potyviral RNA functions in infection

PubMed Central

Mäkinen, Kristiina; Hafrén, Anders

2014-01-01

Establishment of an infection cycle requires mechanisms to allocate the genomes of (+)-stranded RNA viruses in a balanced ratio to translation, replication, encapsidation, and movement, as well as mechanisms to prevent translocation of viral RNA (vRNA) to cellular RNA degradation pathways. The ratio of vRNA allocated to various functions is likely balanced by the availability of regulatory proteins or competition of the interaction sites within regulatory ribonucleoprotein complexes. Due to the transient nature of viral processes and the interdependency between vRNA pathways, it is technically demanding to work out the exact molecular mechanisms underlying vRNA regulation. A substantial number of viral and host proteins have been identified that facilitate the steps that lead to the assembly of a functional potyviral RNA replication complex and their fusion with chloroplasts. Simultaneously with on-going viral replication, part of the replicated potyviral RNA enters movement pathways. Although not much is known about the processes of potyviral RNA release from viral replication complexes, the molecular interactions involved in these processes determine the fate of the replicated vRNA. Some viral and host cell proteins have been described that direct replicated potyviral RNA to translation to enable potyviral gene expression and productive infection. The antiviral defense of the cell causes vRNA degradation by RNA silencing. We hypothesize that also plant pathways involved in mRNA decay may have a role in the coordination of potyviral RNA expression. In this review, we discuss the roles of different potyviral and host proteins in the coordination of various potyviral RNA functions. PMID:24723931

Intracellular coordination of potyviral RNA functions in infection.

PubMed

Mäkinen, Kristiina; Hafrén, Anders

2014-01-01

Establishment of an infection cycle requires mechanisms to allocate the genomes of (+)-stranded RNA viruses in a balanced ratio to translation, replication, encapsidation, and movement, as well as mechanisms to prevent translocation of viral RNA (vRNA) to cellular RNA degradation pathways. The ratio of vRNA allocated to various functions is likely balanced by the availability of regulatory proteins or competition of the interaction sites within regulatory ribonucleoprotein complexes. Due to the transient nature of viral processes and the interdependency between vRNA pathways, it is technically demanding to work out the exact molecular mechanisms underlying vRNA regulation. A substantial number of viral and host proteins have been identified that facilitate the steps that lead to the assembly of a functional potyviral RNA replication complex and their fusion with chloroplasts. Simultaneously with on-going viral replication, part of the replicated potyviral RNA enters movement pathways. Although not much is known about the processes of potyviral RNA release from viral replication complexes, the molecular interactions involved in these processes determine the fate of the replicated vRNA. Some viral and host cell proteins have been described that direct replicated potyviral RNA to translation to enable potyviral gene expression and productive infection. The antiviral defense of the cell causes vRNA degradation by RNA silencing. We hypothesize that also plant pathways involved in mRNA decay may have a role in the coordination of potyviral RNA expression. In this review, we discuss the roles of different potyviral and host proteins in the coordination of various potyviral RNA functions.

Cross-Talk Between Mitochondrial Fusion and the Hippo Pathway in Controlling Cell Proliferation During Drosophila Development.

PubMed

Deng, Qiannan; Guo, Ting; Zhou, Xiu; Xi, Yongmei; Yang, Xiaohang; Ge, Wanzhong

2016-08-01

Cell proliferation and tissue growth depend on the coordinated regulation of multiple signaling molecules and pathways during animal development. Previous studies have linked mitochondrial function and the Hippo signaling pathway in growth control. However, the underlying molecular mechanisms are not fully understood. Here we identify a Drosophila mitochondrial inner membrane protein ChChd3 as a novel regulator for tissue growth. Loss of ChChd3 leads to tissue undergrowth and cell proliferation defects. ChChd3 is required for mitochondrial fusion and removal of ChChd3 increases mitochondrial fragmentation. ChChd3 is another mitochondrial target of the Hippo pathway, although it is only partially required for Hippo pathway-mediated overgrowth. Interestingly, lack of ChChd3 leads to inactivation of Hippo activity under normal development, which is also dependent on the transcriptional coactivator Yorkie (Yki). Furthermore, loss of ChChd3 induces oxidative stress and activates the JNK pathway. In addition, depletion of other mitochondrial fusion components, Opa1 or Marf, inactivates the Hippo pathway as well. Taken together, we propose that there is a cross-talk between mitochondrial fusion and the Hippo pathway, which is essential in controlling cell proliferation and tissue homeostasis in Drosophila. Copyright © 2016 by the Genetics Society of America.

In vivo gene manipulation reveals the impact of stress-responsive MAPK pathways on tumor progression

PubMed Central

Kamiyama, Miki; Naguro, Isao; Ichijo, Hidenori

2015-01-01

It has been widely accepted that tumor cells and normal stromal cells in the host environment coordinately modulate tumor progression. Mitogen-activated protein kinase pathways are the representative stress-responsive cascades that exert proper cellular responses to divergent environmental stimuli. Genetically engineered mouse models and chemically induced tumorigenesis models have revealed that components of the MAPK pathway not only regulate the behavior of tumor cells themselves but also that of surrounding normal stromal cells in the host environment during cancer pathogenesis. The individual functions of MAPK pathway components in tumor initiation and progression vary depending on the stimuli and the stromal cell types involved in tumor progression, in addition to the molecular isoforms of the components and the origins of the tumor. Recent studies have indicated that MAPK pathway components synergize with environmental factors (e.g. tobacco smoke and diet) to affect tumor initiation and progression. Moreover, some components play distinct roles in the course of tumor progression, such as before and after the establishment of tumors. Hence, a comprehensive understanding of the multifaceted functions of MAPK pathway components in tumor initiation and progression is essential for the improvement of cancer therapy. In this review, we focus on the reports that utilized knockout, conditional knockout, and transgenic mice of MAPK pathway components to investigate the effects of MAPK pathway components on tumor initiation and progression in the host environment. PMID:25880821

Signaling pathways effecting crosstalk between cartilage and adjacent tissues: Seminars in cell and developmental biology: The biology and pathology of cartilage.

PubMed

Maes, Christa

2017-02-01

Endochondral ossification, the mechanism responsible for the development of the long bones, is dependent on an extremely stringent coordination between the processes of chondrocyte maturation in the growth plate, vascular expansion in the surrounding tissues, and osteoblast differentiation and osteogenesis in the perichondrium and the developing bone center. The synchronization of these processes occurring in adjacent tissues is regulated through vigorous crosstalk between chondrocytes, endothelial cells and osteoblast lineage cells. Our knowledge about the molecular constituents of these bidirectional communications is undoubtedly incomplete, but certainly some signaling pathways effective in cartilage have been recognized to play key roles in steering vascularization and osteogenesis in the perichondrial tissues. These include hypoxia-driven signaling pathways, governed by the hypoxia-inducible factors (HIFs) and vascular endothelial growth factor (VEGF), which are absolutely essential for the survival and functioning of chondrocytes in the avascular growth plate, at least in part by regulating the oxygenation of developing cartilage through the stimulation of angiogenesis in the surrounding tissues. A second coordinating signal emanating from cartilage and regulating developmental processes in the adjacent perichondrium is Indian Hedgehog (IHH). IHH, produced by pre-hypertrophic and early hypertrophic chondrocytes in the growth plate, induces the differentiation of adjacent perichondrial progenitor cells into osteoblasts, thereby harmonizing the site and time of bone formation with the developmental progression of chondrogenesis. Both signaling pathways represent vital mediators of the tightly organized conversion of avascular cartilage into vascularized and mineralized bone during endochondral ossification. Copyright © 2016. Published by Elsevier Ltd.

Targeting Developmental Pathways: The Achilles Heel of Cancer?

PubMed

Dempke, Wolfram C M; Fenchel, Klaus; Uciechowski, Peter; Chevassut, Timothy

2017-01-01

Developmental pathways (e.g., Notch, Hippo, Hedgehog, Wnt, and TGF-β/BMP/FGF) are networks of genes that act co-ordinately to establish the body plan, and disruptions of genes in one pathway can have effects in related pathways and may result in serious dysmorphogenesis or cancer. Interestingly, all developmental pathways are highly conserved cell signalling systems present in almost all multicellular organisms. In addition, they have a crucial role in cell proliferation, apoptosis, differentiation, and finally in organ development. Of note, almost all of these pathways promote oncogenesis through synergistic associations with the Hippo signalling pathway, and several lines of evidence have also indicated that these pathways (e.g., Wnt/β-catenin) may be implicated in checkpoint inhibitor resistance (e.g., CTLA-4, PD-1, and PD-L1). Since Notch inhibition in vivo results in partial loss of its stemness features such as self-renewal, chemoresistance, invasive and migratory potential, and tumorigenesis, these highly conserved developmental pathways are regarded as being critical for regulation of self-renewal in both embryonic and adult stem cells and hence are likely to be implicated in the maintenance of cancer stem cells. Many small molecules are currently in preclinical and early clinical development, and only two compounds are approved for treatment of advanced or metastatic basal cell carcinoma (vismodegib and sonidegib). Furthermore, therapeutic targeting of cancer stem cells using drugs that disrupt activated developmental pathways may also represent an attractive strategy that is potentially relevant to many types of malignancy, notably blood cancers, where the evidence for leukaemia stem cells is well established. Future work will hopefully pave the way for the development of new strategies for targeting these pervasive oncogenic pathways. © 2017 S. Karger AG, Basel.

Sphingosine 1-Phosphate (S1P) Receptors 1 and 2 Coordinately Induce Mesenchymal Cell Migration through S1P Activation of Complementary Kinase Pathways*

PubMed Central

Quint, Patrick; Ruan, Ming; Pederson, Larry; Kassem, Moustapha; Westendorf, Jennifer J.; Khosla, Sundeep; Oursler, Merry Jo

2013-01-01

Normal bone turnover requires tight coupling of bone resorption and bone formation to preserve bone quantity and structure. With aging and during several pathological conditions, this coupling breaks down, leading to either net bone loss or excess bone formation. To preserve or restore normal bone metabolism, it is crucial to determine the mechanisms by which osteoclasts and osteoblast precursors interact and contribute to coupling. We showed that osteoclasts produce the chemokine sphingosine 1-phosphate (S1P), which stimulates osteoblast migration. Thus, osteoclast-derived S1P may recruit osteoblasts to sites of bone resorption as an initial step in replacing lost bone. In this study we investigated the mechanisms by which S1P stimulates mesenchymal (skeletal) cell chemotaxis. S1P treatment of mesenchymal (skeletal) cells activated RhoA GTPase, but this small G protein did not contribute to migration. Rather, two S1P receptors, S1PR1 and S1PR2, coordinately promoted migration through activation of the JAK/STAT3 and FAK/PI3K/AKT signaling pathways, respectively. These data demonstrate that the chemokine S1P couples bone formation to bone resorption through activation of kinase signaling pathways. PMID:23300082

Cbf11 and Cbf12, the fission yeast CSL proteins, play opposing roles in cell adhesion and coordination of cell and nuclear division

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

Prevorovsky, Martin; Grousl, Tomas; Stanurova, Jana

The CSL (CBF1/RBP-J{kappa}/Suppressor of Hairless/LAG-1) family is comprised of transcription factors essential for metazoan development, mostly due to their involvement in the Notch receptor signaling pathway. Recently, we identified two novel classes of CSL genes in the genomes of several fungal species, organisms lacking the Notch pathway. In this study, we characterized experimentally cbf11{sup +} and cbf12{sup +}, the two CSL genes of Schizosaccharomyces pombe, in order to elucidate the CSL function in fungi. We provide evidence supporting their identity as genuine CSL genes. Both cbf11{sup +} and cbf12{sup +} are non-essential; they have distinct expression profiles and code formore » nuclear proteins with transcription activation potential. Significantly, we demonstrated that Cbf11 recognizes specifically the canonical CSL response element GTG{sup A}/{sub G}GAA in vitro. The deletion of cbf11{sup +} is associated with growth phenotypes and altered colony morphology. Furthermore, we found that Cbf11 and Cbf12 play opposite roles in cell adhesion, nuclear and cell division and their coordination. Disturbed balance of the two CSL proteins leads to cell separation defects (sep phenotype), cut phenotype, and high-frequency diploidization in heterothallic strains. Our data show that CSL proteins operate in an organism predating the Notch pathway, which should be of relevance to the understanding of (Notch-independent) CSL functions in metazoans.« less

Conservation of Planar Polarity Pathway Function Across the Animal Kingdom.

PubMed

Hale, Rosalind; Strutt, David

2015-01-01

Planar polarity is a well-studied phenomenon resulting in the directional coordination of cells in the plane of a tissue. In invertebrates and vertebrates, planar polarity is established and maintained by the largely independent core and Fat/Dachsous/Four-jointed (Ft-Ds-Fj) pathways. Loss of function of these pathways can result in a wide range of developmental or cellular defects, including failure of gastrulation and problems with placement and function of cilia. This review discusses the conservation of these pathways across the animal kingdom. The lack of vital core pathway components in basal metazoans suggests that the core planar polarity pathway evolved shortly after, but not necessarily alongside, the emergence of multicellularity.

Chronophin coordinates cell leading edge dynamics by controlling active cofilin levels

PubMed Central

Delorme-Walker, Violaine; Seo, Ji-Yeon; Gohla, Antje; Fowler, Bruce; Bohl, Ben; DerMardirossian, Céline

2015-01-01

Cofilin, a critical player of actin dynamics, is spatially and temporally regulated to control the direction and force of membrane extension required for cell locomotion. In carcinoma cells, although the signaling pathways regulating cofilin activity to control cell direction have been established, the molecular machinery required to generate the force of the protrusion remains unclear. We show that the cofilin phosphatase chronophin (CIN) spatiotemporally regulates cofilin activity at the cell edge to generate persistent membrane extension. We show that CIN translocates to the leading edge in a PI3-kinase–, Rac1-, and cofilin-dependent manner after EGF stimulation to activate cofilin, promotes actin free barbed end formation, accelerates actin turnover, and enhances membrane protrusion. In addition, we establish that CIN is crucial for the balance of protrusion/retraction events during cell migration. Thus, CIN coordinates the leading edge dynamics by controlling active cofilin levels to promote MTLn3 cell protrusion. PMID:26324884

Coordinated activation of the secretory pathway during notochord formation in the Xenopus embryo.

PubMed

Tanegashima, Kosuke; Zhao, Hui; Rebbert, Martha L; Dawid, Igor B

2009-11-01

We compared the transcriptome in the developing notochord of Xenopus laevis embryos with that of other embryonic regions. A coordinated and intense activation of a large set of secretory pathway genes was observed in the notochord, but not in notochord precursors in the axial mesoderm at early gastrula stage. The genes encoding Xbp1 and Creb3l2 were also activated in the notochord. These two transcription factors are implicated in the activation of secretory pathway genes during the unfolded protein response, where cells react to the stress of a build-up of unfolded proteins in their endoplasmic reticulum. Xbp1 and Creb3l2 are differentially expressed but not differentially activated in the notochord. Reduction of expression of Xbp1 or Creb3l2 by injection of antisense morpholinos led to strong deficits in notochord but not somitic muscle development. In addition, the expression of some, but not all, genes encoding secretory proteins was inhibited by injection of xbp1 morpholinos. Furthermore, expression of activated forms of Xbp1 or Creb3l2 in animal explants could activate a similar subset of secretory pathway genes. We conclude that coordinated activation of a battery of secretory pathway genes mediated by Xbp1 and Creb/ATF factors is a characteristic and necessary feature of notochord formation.

A histone H3K36 chromatin switch coordinates DNA double-strand break repair pathway choice.

PubMed

Pai, Chen-Chun; Deegan, Rachel S; Subramanian, Lakxmi; Gal, Csenge; Sarkar, Sovan; Blaikley, Elizabeth J; Walker, Carol; Hulme, Lydia; Bernhard, Eric; Codlin, Sandra; Bähler, Jürg; Allshire, Robin; Whitehall, Simon; Humphrey, Timothy C

2014-06-09

DNA double-strand break (DSB) repair is a highly regulated process performed predominantly by non-homologous end joining (NHEJ) or homologous recombination (HR) pathways. How these pathways are coordinated in the context of chromatin is unclear. Here we uncover a role for histone H3K36 modification in regulating DSB repair pathway choice in fission yeast. We find Set2-dependent H3K36 methylation reduces chromatin accessibility, reduces resection and promotes NHEJ, while antagonistic Gcn5-dependent H3K36 acetylation increases chromatin accessibility, increases resection and promotes HR. Accordingly, loss of Set2 increases H3K36Ac, chromatin accessibility and resection, while Gcn5 loss results in the opposite phenotypes following DSB induction. Further, H3K36 modification is cell cycle regulated with Set2-dependent H3K36 methylation peaking in G1 when NHEJ occurs, while Gcn5-dependent H3K36 acetylation peaks in S/G2 when HR prevails. These findings support an H3K36 chromatin switch in regulating DSB repair pathway choice.

Monte Carlo Study Elucidates the Type 1/Type 2 Choice in Apoptotic Death Signaling in Healthy and Cancer Cells

PubMed Central

Raychaudhuri, Subhadip; Raychaudhuri, Somkanya C

2013-01-01

Apoptotic cell death is coordinated through two distinct (type 1 and type 2) intracellular signaling pathways. How the type 1/type 2 choice is made remains a central problem in the biology of apoptosis and has implications for apoptosis related diseases and therapy. We study the problem of type 1/type 2 choice in silico utilizing a kinetic Monte Carlo model of cell death signaling. Our results show that the type 1/type 2 choice is linked to deterministic versus stochastic cell death activation, elucidating a unique regulatory control of the apoptotic pathways. Consistent with previous findings, our results indicate that caspase 8 activation level is a key regulator of the choice between deterministic type 1 and stochastic type 2 pathways, irrespective of cell types. Expression levels of signaling molecules downstream also regulate the type 1/type 2 choice. A simplified model of DISC clustering elucidates the mechanism of increased active caspase 8 generation and type 1 activation in cancer cells having increased sensitivity to death receptor activation. We demonstrate that rapid deterministic activation of the type 1 pathway can selectively target such cancer cells, especially if XIAP is also inhibited; while inherent cell-to-cell variability would allow normal cells stay protected. PMID:24709706

The impact of the unfolded protein response on human disease

PubMed Central

Wang, Shiyu

2012-01-01

A central function of the endoplasmic reticulum (ER) is to coordinate protein biosynthetic and secretory activities in the cell. Alterations in ER homeostasis cause accumulation of misfolded/unfolded proteins in the ER. To maintain ER homeostasis, eukaryotic cells have evolved the unfolded protein response (UPR), an essential adaptive intracellular signaling pathway that responds to metabolic, oxidative stress, and inflammatory response pathways. The UPR has been implicated in a variety of diseases including metabolic disease, neurodegenerative disease, inflammatory disease, and cancer. Signaling components of the UPR are emerging as potential targets for intervention and treatment of human disease. PMID:22733998

Modulation of Olfactory Bulb Network Activity by Serotonin: Synchronous Inhibition of Mitral Cells Mediated by Spatially Localized GABAergic Microcircuits

ERIC Educational Resources Information Center

Schmidt, Loren J.; Strowbridge, Ben W.

2014-01-01

Although inhibition has often been proposed as a central mechanism for coordinating activity in the olfactory system, relatively little is known about how activation of different inhibitory local circuit pathways can generate coincident inhibition of principal cells. We used serotonin (5-HT) as a pharmacological tool to induce spiking in ensembles…

CREB3L1-mediated functional and structural adaptation of the secretory pathway in hormone-stimulated thyroid cells.

PubMed

García, Iris A; Torres Demichelis, Vanina; Viale, Diego L; Di Giusto, Pablo; Ezhova, Yulia; Polishchuk, Roman S; Sampieri, Luciana; Martinez, Hernán; Sztul, Elizabeth; Alvarez, Cecilia

2017-12-15

Many secretory cells increase the synthesis and secretion of cargo proteins in response to specific stimuli. How cells couple increased cargo load with a coordinate rise in secretory capacity to ensure efficient transport is not well understood. We used thyroid cells stimulated with thyrotropin (TSH) to demonstrate a coordinate increase in the production of thyroid-specific cargo proteins and ER-Golgi transport factors, and a parallel expansion of the Golgi complex. TSH also increased expression of the CREB3L1 transcription factor, which alone caused amplified transport factor levels and Golgi enlargement. Furthermore, CREB3L1 potentiated the TSH-induced increase in Golgi volume. A dominant-negative CREB3L1 construct hampered the ability of TSH to induce Golgi expansion, implying that this transcription factor contributes to Golgi expansion. Our findings support a model in which CREB3L1 acts as a downstream effector of TSH to regulate the expression of cargo proteins, and simultaneously increases the synthesis of transport factors and the expansion of the Golgi to synchronize the rise in cargo load with the amplified capacity of the secretory pathway. © 2017. Published by The Company of Biologists Ltd.

Arabidopsis AIP1-2 restricted by WER-mediated patterning modulates planar polarity.

PubMed

Kiefer, Christian S; Claes, Andrea R; Nzayisenga, Jean-Claude; Pietra, Stefano; Stanislas, Thomas; Hüser, Anke; Ikeda, Yoshihisa; Grebe, Markus

2015-01-01

The coordination of cell polarity within the plane of the tissue layer (planar polarity) is crucial for the development of diverse multicellular organisms. Small Rac/Rho-family GTPases and the actin cytoskeleton contribute to planar polarity formation at sites of polarity establishment in animals and plants. Yet, upstream pathways coordinating planar polarity differ strikingly between kingdoms. In the root of Arabidopsis thaliana, a concentration gradient of the phytohormone auxin coordinates polar recruitment of Rho-of-plant (ROP) to sites of polar epidermal hair initiation. However, little is known about cytoskeletal components and interactions that contribute to this planar polarity or about their relation to the patterning machinery. Here, we show that ACTIN7 (ACT7) represents a main actin isoform required for planar polarity of root hair positioning, interacting with the negative modulator ACTIN-INTERACTING PROTEIN1-2 (AIP1-2). ACT7, AIP1-2 and their genetic interaction are required for coordinated planar polarity of ROP downstream of ethylene signalling. Strikingly, AIP1-2 displays hair cell file-enriched expression, restricted by WEREWOLF (WER)-dependent patterning and modified by ethylene and auxin action. Hence, our findings reveal AIP1-2, expressed under control of the WER-dependent patterning machinery and the ethylene signalling pathway, as a modulator of actin-mediated planar polarity. © 2015. Published by The Company of Biologists Ltd.

Distinct but Concerted Roles of ATR, DNA-PK, and Chk1 in Countering Replication Stress during S Phase

PubMed Central

Buisson, Rémi; Boisvert, Jessica L.; Benes, Cyril H.; Zou, Lee

2015-01-01

The ATR-Chk1 pathway is critical for DNA damage responses and cell cycle progression. Chk1 inhibition is more deleterious to cycling cells than ATR inhibition, raising questions about ATR and Chk1 functions in the absence of extrinsic replication stress. Here, we show that a key role of ATR in S phase is to coordinate RRM2 accumulation and origin firing. ATR inhibitor (ATRi) induces massive ssDNA accumulation and replication catastrophe in a fraction of early S-phase cells. In other S-phase cells, however, ATRi induces moderate ssDNA and triggers a DNA-PK and Chk1-mediated backup pathway to suppress origin firing. The backup pathway creates a threshold such that ATRi selectively kills cells under high replication stress, whereas Chk1 inhibitor induces cell death at a lower threshold. The levels of ATRi-induced ssDNA correlate with ATRi sensitivity in a panel of cell lines, suggesting that ATRi-induced ssDNA could be predictive of ATRi sensitivity in cancer cells. PMID:26365377

Hsp90 Orchestrates Transcriptional Regulation by Hsf1 and Cell Wall Remodelling by MAPK Signalling during Thermal Adaptation in a Pathogenic Yeast

PubMed Central

Leach, Michelle D.; Budge, Susan; Walker, Louise; Munro, Carol; Cowen, Leah E.; Brown, Alistair J. P.

2012-01-01

Thermal adaptation is essential in all organisms. In yeasts, the heat shock response is commanded by the heat shock transcription factor Hsf1. Here we have integrated unbiased genetic screens with directed molecular dissection to demonstrate that multiple signalling cascades contribute to thermal adaptation in the pathogenic yeast Candida albicans. We show that the molecular chaperone heat shock protein 90 (Hsp90) interacts with and down-regulates Hsf1 thereby modulating short term thermal adaptation. In the longer term, thermal adaptation depends on key MAP kinase signalling pathways that are associated with cell wall remodelling: the Hog1, Mkc1 and Cek1 pathways. We demonstrate that these pathways are differentially activated and display cross talk during heat shock. As a result ambient temperature significantly affects the resistance of C. albicans cells to cell wall stresses (Calcofluor White and Congo Red), but not osmotic stress (NaCl). We also show that the inactivation of MAP kinase signalling disrupts this cross talk between thermal and cell wall adaptation. Critically, Hsp90 coordinates this cross talk. Genetic and pharmacological inhibition of Hsp90 disrupts the Hsf1-Hsp90 regulatory circuit thereby disturbing HSP gene regulation and reducing the resistance of C. albicans to proteotoxic stresses. Hsp90 depletion also affects cell wall biogenesis by impairing the activation of its client proteins Mkc1 and Hog1, as well as Cek1, which we implicate as a new Hsp90 client in this study. Therefore Hsp90 modulates the short term Hsf1-mediated activation of the classic heat shock response, coordinating this response with long term thermal adaptation via Mkc1- Hog1- and Cek1-mediated cell wall remodelling. PMID:23300438

Network inference reveals novel connections in pathways regulating growth and defense in the yeast salt response.

PubMed

MacGilvray, Matthew E; Shishkova, Evgenia; Chasman, Deborah; Place, Michael; Gitter, Anthony; Coon, Joshua J; Gasch, Audrey P

2018-05-01

Cells respond to stressful conditions by coordinating a complex, multi-faceted response that spans many levels of physiology. Much of the response is coordinated by changes in protein phosphorylation. Although the regulators of transcriptome changes during stress are well characterized in Saccharomyces cerevisiae, the upstream regulatory network controlling protein phosphorylation is less well dissected. Here, we developed a computational approach to infer the signaling network that regulates phosphorylation changes in response to salt stress. We developed an approach to link predicted regulators to groups of likely co-regulated phospho-peptides responding to stress, thereby creating new edges in a background protein interaction network. We then use integer linear programming (ILP) to integrate wild type and mutant phospho-proteomic data and predict the network controlling stress-activated phospho-proteomic changes. The network we inferred predicted new regulatory connections between stress-activated and growth-regulating pathways and suggested mechanisms coordinating metabolism, cell-cycle progression, and growth during stress. We confirmed several network predictions with co-immunoprecipitations coupled with mass-spectrometry protein identification and mutant phospho-proteomic analysis. Results show that the cAMP-phosphodiesterase Pde2 physically interacts with many stress-regulated transcription factors targeted by PKA, and that reduced phosphorylation of those factors during stress requires the Rck2 kinase that we show physically interacts with Pde2. Together, our work shows how a high-quality computational network model can facilitate discovery of new pathway interactions during osmotic stress.

Hippo circuitry and the redox modulation of hippo components in cancer cell fate decisions.

PubMed

Ashraf, Asma; Pervaiz, Shazib

2015-12-01

Meticulous and precise control of organ size is undoubtedly one of the most pivotal processes in mammalian development and regeneration along with cell differentiation, morphogenesis and programmed cell death. These processes are strictly regulated by complex and highly coordinated mechanisms to maintain a steady growth state. There are a number of extrinsic and intrinsic factors that dictate the total number and/or size of cells by influencing growth, proliferation, differentiation and cell death. Multiple pathways, such as those involved in promoting organ size and others that restrict disproportionate tissue growth act simultaneously to maintain cellular and tissue homeostasis. Aberrations at any level in these organ size-regulating processes can lead to various pathological states with cancers being the most formidable one (Yin and Zhang, 2011). Extensive research in the realm of growth control has led to the identification of the Hippo-signaling pathway as a critical network in modulating tissue growth via its effect on multiple signaling pathways and through intricate crosstalk with proteins that regulate cell polarity, adhesion and cell-cell interactions (Zhao et al., 2011b). The Hippo pathway controls cell number and organ size by transducing signals from the plasma membrane to the nucleus to regulate the expression of genes involved in cell fate determination (Shi et al., 2015). In this review, we summarize the recent discoveries concerning Hippo pathway, its diversiform regulation in mammals as well as its implications in cancers, and highlight the possible role of oxidative stress in Hippo pathway regulation. Copyright © 2015 Elsevier Ltd. All rights reserved.

Disruption of the Fanconi anemia-BRCA pathway in cisplatin-sensitive ovarian tumors.

PubMed

Taniguchi, Toshiyasu; Tischkowitz, Marc; Ameziane, Najim; Hodgson, Shirley V; Mathew, Christopher G; Joenje, Hans; Mok, Samuel C; D'Andrea, Alan D

2003-05-01

Ovarian tumor cells are often genomically unstable and hypersensitive to cisplatin. To understand the molecular basis for this phenotype, we examined the integrity of the Fanconi anemia-BRCA (FANC-BRCA) pathway in those cells. This pathway regulates cisplatin sensitivity and is governed by the coordinate activity of six genes associated with Fanconi anemia (FANCA, FANCC, FANCD2, FANCE, FANCF and FANCG) as well as BRCA1 and BRCA2 (FANCD1). Here we show that the FANC-BRCA pathway is disrupted in a subset of ovarian tumor lines. Mono-ubiquitination of FANCD2, a measure of the function of this pathway, and cisplatin resistance were restored by functional complementation with FANCF, a gene that is upstream in this pathway. FANCF inactivation in ovarian tumors resulted from methylation of its CpG island, and acquired cisplatin resistance correlated with demethylation of FANCF. We propose a model for ovarian tumor progression in which the initial methylation of FANCF is followed by FANCF demethylation and ultimately results in cisplatin resistance.

Pavement cells: a model system for non-transcriptional auxin signalling and crosstalks

PubMed Central

Chen, Jisheng; Wang, Fei; Zheng, Shiqin; Xu, Tongda; Yang, Zhenbiao

2015-01-01

Auxin (indole acetic acid) is a multifunctional phytohormone controlling various developmental patterns, morphogenetic processes, and growth behaviours in plants. The transcription-based pathway activated by the nuclear TRANSPORT INHIBITOR RESISTANT 1/auxin-related F-box auxin receptors is well established, but the long-sought molecular mechanisms of non-transcriptional auxin signalling remained enigmatic until very recently. Along with the establishment of the Arabidopsis leaf epidermal pavement cell (PC) as an exciting and amenable model system in the past decade, we began to gain insight into non-transcriptional auxin signalling. The puzzle-piece shape of PCs forms from intercalated or interdigitated cell growth, requiring local intra- and inter-cellular coordination of lobe and indent formation. Precise coordination of this interdigitated pattern requires auxin and an extracellular auxin sensing system that activates plasma membrane-associated Rho GTPases from plants and subsequent downstream events regulating cytoskeletal reorganization and PIN polarization. Apart from auxin, mechanical stress and cytokinin have been shown to affect PC interdigitation, possibly by interacting with auxin signals. This review focuses upon signalling mechanisms for cell polarity formation in PCs, with an emphasis on non-transcriptional auxin signalling in polarized cell expansion and pattern formation and how different auxin pathways interplay with each other and with other signals. PMID:26047974

Zyxin-Siah2–Lats2 axis mediates cooperation between Hippo and TGF-β signalling pathways

PubMed Central

Ma, Biao; Cheng, Hongcheng; Gao, Ruize; Mu, Chenglong; Chen, Ling; Wu, Shian; Chen, Quan; Zhu, Yushan

2016-01-01

The evolutionarily conserved Hippo pathway is a regulator that controls organ size, cell growth and tissue homeostasis. Upstream signals of the Hippo pathway have been widely studied, but how microenvironmental factors coordinately regulate this pathway remains unclear. In this study, we identify LIM domain protein Zyxin, as a scaffold protein, that in response to hypoxia and TGF-β stimuli, forms a ternary complex with Lats2 and Siah2 and stabilizes their interaction. This interaction facilitates Lats2 ubiquitination and degradation, Yap dephosphorylation and subsequently activation. We show that Zyxin is required for TGF-β and hypoxia-induced Lats2 downregulation and deactivation of Hippo signalling in MDA-MB-231 cells. Depletion of Zyxin impairs the capability of cell migration, proliferation and tumourigenesis in a xenograft model. Zyxin is upregulated in human breast cancer and positively correlates with histological stages and metastasis. Our study demonstrates that Zyxin-Lats2–Siah2 axis may serve as a potential therapeutic target in cancer treatment. PMID:27030211

The reorientation of cell nucleus promotes the establishment of front-rear polarity in migrating fibroblasts.

PubMed

Maninová, Miloslava; Klímová, Zuzana; Parsons, J Thomas; Weber, Michael J; Iwanicki, Marcin P; Vomastek, Tomáš

2013-06-12

The establishment of cell polarity is an essential step in the process of cell migration. This process requires precise spatiotemporal coordination of signaling pathways that in most cells create the typical asymmetrical profile of a polarized cell with nucleus located at the cell rear and the microtubule organizing center (MTOC) positioned between the nucleus and the leading edge. During cell polarization, nucleus rearward positioning promotes correct microtubule organizing center localization and thus the establishment of front-rear polarity and directional migration. We found that cell polarization and directional migration require also the reorientation of the nucleus. Nuclear reorientation is manifested as temporally restricted nuclear rotation that aligns the nuclear axis with the axis of cell migration. We also found that nuclear reorientation requires physical connection between the nucleus and cytoskeleton mediated by the LINC (linker of nucleoskeleton and cytoskeleton) complex. Nuclear reorientation is controlled by coordinated activity of lysophosphatidic acid (LPA)-mediated activation of GTPase Rho and the activation of integrin, FAK (focal adhesion kinase), Src, and p190RhoGAP signaling pathway. Integrin signaling is spatially induced at the leading edge as FAK and p190RhoGAP are predominantly activated or localized at this location. We suggest that integrin activation within lamellipodia defines cell front, and subsequent FAK, Src, and p190RhoGAP signaling represents the polarity signal that induces reorientation of the nucleus and thus promotes the establishment of front-rear polarity. Copyright © 2013 Elsevier Ltd. All rights reserved.

Murine craniofacial development requires Hdac3-mediated repression of Msx gene expression

PubMed Central

Singh, Nikhil; Gupta, Mudit; Trivedi, Chinmay M.; Singh, Manvendra K.; Li, Li; Epstein, Jonathan A.

2013-01-01

Craniofacial development is characterized by reciprocal interactions between neural crest cells and neighboring cell populations of ectodermal, endodermal and mesodermal origin. Various genetic pathways play critical roles in coordinating the development of cranial structures by modulating the growth, survival and differentiation of neural crest cells. However, the regulation of these pathways, particularly at the epigenomic level, remains poorly understood. Using murine genetics, we show that neural crest cells exhibit a requirement for the class I histone deacetylase Hdac3 during craniofacial development. Mice in which Hdac3 has been conditionally deleted in neural crest demonstrate fully penetrant craniofacial abnormalities, including microcephaly, cleft secondary palate and dental hypoplasia. Consistent with these abnormalities, we observe dysregulation of cell cycle genes and increased apoptosis in neural crest structures in mutant embryos. Known regulators of cell cycle progression and apoptosis in neural crest, including Msx1, Msx2 and Bmp4, are upregulated in Hdac3-deficient cranial mesenchyme. These results suggest that Hdac3 serves as a critical regulator of craniofacial morphogenesis, in part by repressing core apoptotic pathways in cranial neural crest cells. PMID:23506836

Structural basis for leucine sensing by the Sestrin2-mTORC1 pathway.

PubMed

Saxton, Robert A; Knockenhauer, Kevin E; Wolfson, Rachel L; Chantranupong, Lynne; Pacold, Michael E; Wang, Tim; Schwartz, Thomas U; Sabatini, David M

2016-01-01

Eukaryotic cells coordinate growth with the availability of nutrients through the mechanistic target of rapamycin complex 1 (mTORC1), a master growth regulator. Leucine is of particular importance and activates mTORC1 via the Rag guanosine triphosphatases and their regulators GATOR1 and GATOR2. Sestrin2 interacts with GATOR2 and is a leucine sensor. Here we present the 2.7 angstrom crystal structure of Sestrin2 in complex with leucine. Leucine binds through a single pocket that coordinates its charged functional groups and confers specificity for the hydrophobic side chain. A loop encloses leucine and forms a lid-latch mechanism required for binding. A structure-guided mutation in Sestrin2 that decreases its affinity for leucine leads to a concomitant increase in the leucine concentration required for mTORC1 activation in cells. These results provide a structural mechanism of amino acid sensing by the mTORC1 pathway. Copyright © 2016, American Association for the Advancement of Science.

Dual role for DOCK7 in tangential migration of interneuron precursors in the postnatal forebrain.

PubMed

Nakamuta, Shinichi; Yang, Yu-Ting; Wang, Chia-Lin; Gallo, Nicholas B; Yu, Jia-Ray; Tai, Yilin; Van Aelst, Linda

2017-12-04

Throughout life, stem cells in the ventricular-subventricular zone generate neuroblasts that migrate via the rostral migratory stream (RMS) to the olfactory bulb, where they differentiate into local interneurons. Although progress has been made toward identifying extracellular factors that guide the migration of these cells, little is known about the intracellular mechanisms that govern the dynamic reshaping of the neuroblasts' morphology required for their migration along the RMS. In this study, we identify DOCK7, a member of the DOCK180-family, as a molecule essential for tangential neuroblast migration in the postnatal mouse forebrain. DOCK7 regulates the migration of these cells by controlling both leading process (LP) extension and somal translocation via distinct pathways. It controls LP stability/growth via a Rac-dependent pathway, likely by modulating microtubule networks while also regulating F-actin remodeling at the cell rear to promote somal translocation via a previously unrecognized myosin phosphatase-RhoA-interacting protein-dependent pathway. The coordinated action of both pathways is required to ensure efficient neuroblast migration along the RMS. © 2017 Nakamuta et al.

Dual role for DOCK7 in tangential migration of interneuron precursors in the postnatal forebrain

PubMed Central

Yang, Yu-Ting; Yu, Jia-Ray; Tai, Yilin

2017-01-01

Throughout life, stem cells in the ventricular–subventricular zone generate neuroblasts that migrate via the rostral migratory stream (RMS) to the olfactory bulb, where they differentiate into local interneurons. Although progress has been made toward identifying extracellular factors that guide the migration of these cells, little is known about the intracellular mechanisms that govern the dynamic reshaping of the neuroblasts’ morphology required for their migration along the RMS. In this study, we identify DOCK7, a member of the DOCK180-family, as a molecule essential for tangential neuroblast migration in the postnatal mouse forebrain. DOCK7 regulates the migration of these cells by controlling both leading process (LP) extension and somal translocation via distinct pathways. It controls LP stability/growth via a Rac-dependent pathway, likely by modulating microtubule networks while also regulating F-actin remodeling at the cell rear to promote somal translocation via a previously unrecognized myosin phosphatase–RhoA–interacting protein-dependent pathway. The coordinated action of both pathways is required to ensure efficient neuroblast migration along the RMS. PMID:29089377

Cryptococcal titan cell formation is regulated by G-protein signaling in response to multiple stimuli.

PubMed

Okagaki, Laura H; Wang, Yina; Ballou, Elizabeth R; O'Meara, Teresa R; Bahn, Yong-Sun; Alspaugh, J Andrew; Xue, Chaoyang; Nielsen, Kirsten

2011-10-01

The titan cell is a recently described morphological form of the pathogenic fungus Cryptococcus neoformans. Occurring during the earliest stages of lung infection, titan cells are 5 to 10 times larger than the normal yeast-like cells, thereby resisting engulfment by lung phagocytes and favoring the persistence of infection. These enlarged cells exhibit an altered capsule structure, a thickened cell wall, increased ploidy, and resistance to nitrosative and oxidative stresses. We demonstrate that two G-protein-coupled receptors are important for induction of the titan cell phenotype: the Ste3a pheromone receptor (in mating type a cells) and the Gpr5 protein. Both receptors control titan cell formation through elements of the cyclic AMP (cAMP)/protein kinase A (PKA) pathway. This conserved signaling pathway, in turn, mediates its effect on titan cells through the PKA-regulated Rim101 transcription factor. Additional downstream effectors required for titan cell formation include the G(1) cyclin Pcl103, the Rho104 GTPase, and two GTPase-activating proteins, Gap1 and Cnc1560. These observations support developing models in which the PKA signaling pathway coordinately regulates many virulence-associated phenotypes in diverse human pathogens.

Cryptococcal Titan Cell Formation Is Regulated by G-Protein Signaling in Response to Multiple Stimuli▿†

PubMed Central

Okagaki, Laura H.; Wang, Yina; Ballou, Elizabeth R.; O'Meara, Teresa R.; Bahn, Yong-Sun; Alspaugh, J. Andrew; Xue, Chaoyang; Nielsen, Kirsten

2011-01-01

The titan cell is a recently described morphological form of the pathogenic fungus Cryptococcus neoformans. Occurring during the earliest stages of lung infection, titan cells are 5 to 10 times larger than the normal yeast-like cells, thereby resisting engulfment by lung phagocytes and favoring the persistence of infection. These enlarged cells exhibit an altered capsule structure, a thickened cell wall, increased ploidy, and resistance to nitrosative and oxidative stresses. We demonstrate that two G-protein-coupled receptors are important for induction of the titan cell phenotype: the Ste3a pheromone receptor (in mating type a cells) and the Gpr5 protein. Both receptors control titan cell formation through elements of the cyclic AMP (cAMP)/protein kinase A (PKA) pathway. This conserved signaling pathway, in turn, mediates its effect on titan cells through the PKA-regulated Rim101 transcription factor. Additional downstream effectors required for titan cell formation include the G1 cyclin Pcl103, the Rho104 GTPase, and two GTPase-activating proteins, Gap1 and Cnc1560. These observations support developing models in which the PKA signaling pathway coordinately regulates many virulence-associated phenotypes in diverse human pathogens. PMID:21821718

The epidermis coordinates auxin-induced stem growth in response to shade

PubMed Central

Procko, Carl; Burko, Yogev; Long, Jeff A.; Chory, Joanne

2016-01-01

Growth of a complex multicellular organism requires coordinated changes in diverse cell types. These cellular changes generate organs of the correct size, shape, and functionality. In plants, the growth hormone auxin induces stem elongation in response to shade; however, which cell types of the stem perceive the auxin signal and contribute to organ growth is poorly understood. Here, we blocked the transcriptional response to auxin within specific tissues to show that auxin signaling is required in many cell types for correct hypocotyl growth in shade, with a key role for the epidermis. Combining genetic manipulations in Arabidopsis thaliana with transcriptional profiling of the hypocotyl epidermis from Brassica rapa, we show that auxin acts in the epidermis in part by inducing activity of the locally acting, growth-promoting brassinosteroid pathway. Our findings clarify cell-specific auxin function in the hypocotyl and highlight the complexity of cell type interactions within a growing organ. PMID:27401556

Determinants of cell-to-cell variability in protein kinase signaling.

PubMed

Jeschke, Matthias; Baumgärtner, Stephan; Legewie, Stefan

2013-01-01

Cells reliably sense environmental changes despite internal and external fluctuations, but the mechanisms underlying robustness remain unclear. We analyzed how fluctuations in signaling protein concentrations give rise to cell-to-cell variability in protein kinase signaling using analytical theory and numerical simulations. We characterized the dose-response behavior of signaling cascades by calculating the stimulus level at which a pathway responds ('pathway sensitivity') and the maximal activation level upon strong stimulation. Minimal kinase cascades with gradual dose-response behavior show strong variability, because the pathway sensitivity and the maximal activation level cannot be simultaneously invariant. Negative feedback regulation resolves this trade-off and coordinately reduces fluctuations in the pathway sensitivity and maximal activation. Feedbacks acting at different levels in the cascade control different aspects of the dose-response curve, thereby synergistically reducing the variability. We also investigated more complex, ultrasensitive signaling cascades capable of switch-like decision making, and found that these can be inherently robust to protein concentration fluctuations. We describe how the cell-to-cell variability of ultrasensitive signaling systems can be actively regulated, e.g., by altering the expression of phosphatase(s) or by feedback/feedforward loops. Our calculations reveal that slow transcriptional negative feedback loops allow for variability suppression while maintaining switch-like decision making. Taken together, we describe design principles of signaling cascades that promote robustness. Our results may explain why certain signaling cascades like the yeast pheromone pathway show switch-like decision making with little cell-to-cell variability.

Cell elongation is an adaptive response for clearing long chromatid arms from the cleavage plane

PubMed Central

Kotadia, Shaila; Montembault, Emilie; Sullivan, William

2012-01-01

Chromosome segregation must be coordinated with cell cleavage to ensure correct transmission of the genome to daughter cells. Here we identify a novel mechanism by which Drosophila melanogaster neuronal stem cells coordinate sister chromatid segregation with cleavage furrow ingression. Cells adapted to a dramatic increase in chromatid arm length by transiently elongating during anaphase/telophase. The degree of cell elongation correlated with the length of the trailing chromatid arms and was concomitant with a slight increase in spindle length and an enlargement of the zone of cortical myosin distribution. Rho guanine-nucleotide exchange factor (Pebble)–depleted cells failed to elongate during segregation of long chromatids. As a result, Pebble-depleted adult flies exhibited morphological defects likely caused by cell death during development. These studies reveal a novel pathway linking trailing chromatid arms and cortical myosin that ensures the clearance of chromatids from the cleavage plane at the appropriate time during cytokinesis, thus preserving genome integrity. PMID:23185030

Phosphatidylinositol 3-kinase coordinately activates the MEK/ERK and AKT/NFkappaB pathways to maintain osteoclast survival.

PubMed

Gingery, Anne; Bradley, Elizabeth; Shaw, Aubie; Oursler, Merry Jo

2003-05-01

We have examined highly purified osteoclasts that were generated in vitro from murine co-culture of marrow precursors with stromal support cells and have found evidence of activation of the MEK/ERK and AKT/NFkappaB survival pathways. Many mature marrow-derived osteoclasts survived for at least 48 h in culture whether or not they are maintained with stromal cells. Moreover, supplementing purified osteoclasts with RANKL and/or M-CSF had no impact on their survival pattern. In addition, spleen-derived osteoclasts generated with RANKL and M-CSF treatment exhibited a similar survival pattern. Blocking MEK, AKT, or NFkappaB activity resulted in apoptosis of many, but not all, of the osteoclasts in purified marrow-derived osteoclasts, marrow-derived osteoclasts co-cultured with stromal cells, and spleen-derived osteoclasts maintained with RANKL and M-CSF. These data support that both the MEK/ERK and AKT/NFkappaB pathways contribute to osteoclast survival. Since PI3K has been shown to activate either of these pathways, we have examined its role in osteoclast survival. PI3K inhibition caused apoptosis of nearly all osteoclasts in purified and co-cultured marrow-derived osteoclasts and spleen-derived osteoclasts maintained with RANKL and M-CSF. Interestingly, in marrow-derived co-cultures, the apoptotic response was restricted to osteoclasts as there was no evidence of stromal support cell apoptosis. PI3K inhibition also blocked MEK1/2, ERK1/2, and AKT phosphorylation and NFkappaB activation in purified osteoclasts. Simultaneous blockage of both AKT and MEK1/2 caused rapid apoptosis of nearly all osteoclasts, mimicking the response to PI3K inhibition. These data reveal that PI3K coordinately activates two distinct survival pathways that are both important in osteoclast survival. Copyright 2003 Wiley-Liss, Inc.

Glycolysis Is Governed by Growth Regime and Simple Enzyme Regulation in Adherent MDCK Cells

PubMed Central

Rehberg, Markus; Ritter, Joachim B.; Reichl, Udo

2014-01-01

Due to its vital importance in the supply of cellular pathways with energy and precursors, glycolysis has been studied for several decades regarding its capacity and regulation. For a systems-level understanding of the Madin-Darby canine kidney (MDCK) cell metabolism, we couple a segregated cell growth model published earlier with a structured model of glycolysis, which is based on relatively simple kinetics for enzymatic reactions of glycolysis, to explain the pathway dynamics under various cultivation conditions. The structured model takes into account in vitro enzyme activities, and links glycolysis with pentose phosphate pathway and glycogenesis. Using a single parameterization, metabolite pool dynamics during cell cultivation, glucose limitation and glucose pulse experiments can be consistently reproduced by considering the cultivation history of the cells. Growth phase-dependent glucose uptake together with cell-specific volume changes generate high intracellular metabolite pools and flux rates to satisfy the cellular demand during growth. Under glucose limitation, the coordinated control of glycolytic enzymes re-adjusts the glycolytic flux to prevent the depletion of glycolytic intermediates. Finally, the model's predictive power supports the design of more efficient bioprocesses. PMID:25329309

Glycolysis is governed by growth regime and simple enzyme regulation in adherent MDCK cells.

PubMed

Rehberg, Markus; Ritter, Joachim B; Reichl, Udo

2014-10-01

Due to its vital importance in the supply of cellular pathways with energy and precursors, glycolysis has been studied for several decades regarding its capacity and regulation. For a systems-level understanding of the Madin-Darby canine kidney (MDCK) cell metabolism, we couple a segregated cell growth model published earlier with a structured model of glycolysis, which is based on relatively simple kinetics for enzymatic reactions of glycolysis, to explain the pathway dynamics under various cultivation conditions. The structured model takes into account in vitro enzyme activities, and links glycolysis with pentose phosphate pathway and glycogenesis. Using a single parameterization, metabolite pool dynamics during cell cultivation, glucose limitation and glucose pulse experiments can be consistently reproduced by considering the cultivation history of the cells. Growth phase-dependent glucose uptake together with cell-specific volume changes generate high intracellular metabolite pools and flux rates to satisfy the cellular demand during growth. Under glucose limitation, the coordinated control of glycolytic enzymes re-adjusts the glycolytic flux to prevent the depletion of glycolytic intermediates. Finally, the model's predictive power supports the design of more efficient bioprocesses.

Crosstalk between Hippo signalling and miRNAs in tumour progression.

PubMed

Li, Nianshuang; Xie, Chuan; Lu, Nonghua

2017-04-01

The Hippo signalling pathway co-ordinately modulates cell regeneration and organ size, and its deregulation contributes to tumorigenesis through many cellular processes, including overproliferation, apoptosis resistance and cell migration. Recent discoveries have shed new light on how microRNAs (miRNAs) are closely linked to the Hippo pathway in tumour progression. Hippo signalling has been reported to affect widespread miRNA biogenesis. In turn, several miRNAs regulate Hippo signalling, which contributes to carcinogenesis. This article will provide an overview of the crosstalk between Hippo signalling and miRNAs in the development of cancer and further appraise potential targets for therapeutic intervention. © 2016 Federation of European Biochemical Societies.

Developmental alterations in Huntington’s disease neural cells and pharmacological rescue in cells and mice

PubMed Central

2017-01-01

Neural cultures derived from Huntington’s disease (HD) patient-derived induced pluripotent stem cells were used for ‘omics’ analyses to identify mechanisms underlying neurodegeneration. RNA-seq analysis identified genes in glutamate and GABA signaling, axonal guidance and calcium influx whose expression was decreased in HD cultures. One-third of gene changes were in pathways regulating neuronal development and maturation. When mapped to stages of mouse striatal development, the profiles aligned with earlier embryonic stages of neuronal differentiation. We observed a strong correlation between HD-related histone marks, gene expression and unique peak profiles associated with dysregulated genes, suggesting a coordinated epigenetic program. Treatment with isoxazole-9, which targets key dysregulated pathways, led to amelioration of expanded polyglutamine repeat-associated phenotypes in neural cells and of cognitive impairment and synaptic pathology in HD model R6/2 mice. These data suggest that mutant huntingtin impairs neurodevelopmental pathways that could disrupt synaptic homeostasis and increase vulnerability to the pathologic consequence of expanded polyglutamine repeats over time. PMID:28319609

The history and regulatory mechanism of the Hippo pathway

PubMed Central

Kim, Wantae; Jho, Eek-hoon

2018-01-01

How the organ size is adjusted to the proper size during development and how organs know that they reach the original size during regeneration remain long-standing questions. Based on studies using multiple model organisms and approaches for over 20 years, a consensus has been established that the Hippo pathway plays crucial roles in controlling organ size and maintaining tissue homeostasis. Given the significance of these processes, the dysregulation of the Hippo pathway has also implicated various diseases, such as tissue degeneration and cancer. By regulating the downstream transcriptional coactivators YAP and TAZ, the Hippo pathway coordinates cell proliferation and apoptosis in response to a variety of signals including cell contact inhibition, polarity, mechanical sensation and soluble factors. Since the core components and their functions of the Hippo pathway are evolutionarily conserved, this pathway serves as a global regulator of organ size control. Therefore, further investigation of the regulatory mechanisms will provide physiological insights to better understand tissue homeostasis. In this review, the historical developments and current understandings of the regulatory mechanism of Hippo signaling pathway are discussed. PMID:29397869

Size uniformity of animal cells is actively maintained by a p38 MAPK-dependent regulation of G1-length.

PubMed

Liu, Shixuan; Ginzberg, Miriam Bracha; Patel, Nish; Hild, Marc; Leung, Bosco; Li, Zhengda; Chen, Yen-Chi; Chang, Nancy; Wang, Yuan; Tan, Ceryl; Diena, Shulamit; Trimble, William; Wasserman, Larry; Jenkins, Jeremy L; Kirschner, Marc W; Kafri, Ran

2018-03-29

Animal cells within a tissue typically display a striking regularity in their size. To date, the molecular mechanisms that control this uniformity are still unknown. We have previously shown that size uniformity in animal cells is promoted, in part, by size-dependent regulation of G1 length. To identify the molecular mechanisms underlying this process, we performed a large-scale small molecule screen and found that the p38 MAPK pathway is involved in coordinating cell size and cell cycle progression. Small cells display higher p38 activity and spend more time in G1 than larger cells. Inhibition of p38 MAPK leads to loss of the compensatory G1 length extension in small cells, resulting in faster proliferation, smaller cell size and increased size heterogeneity. We propose a model wherein the p38 pathway responds to changes in cell size and regulates G1 exit accordingly, to increase cell size uniformity. © 2017, Liu et al.

Transcript level coordination of carbon pathways during silicon starvation-induced lipid accumulation in the diatom Thalassiosira pseudonana

DOE PAGES

Smith, Sarah R.; Gle, Corine; Abbriano, Raffaela M.; ...

2016-02-04

Diatoms are one of the most productive and successful photosynthetic taxa on Earth and possess attributes such as rapid growth rates and production of lipids, making them candidate sources of renewable fuels. Despite their significance, few details of the mechanisms used to regulate growth and carbon metabolism are currently known, hindering metabolic engineering approaches to enhance productivity. To characterize the transcript level component of metabolic regulation, genome-wide changes in transcript abundance were documented in the model diatom Thalassiosira pseudonana on a time-course of silicon starvation. Growth, cell cycle progression, chloroplast replication, fatty acid composition, pigmentation, and photosynthetic parameters were characterizedmore » alongside lipid accumulation. Extensive coordination of large suites of genes was observed, highlighting the existence of clusters of coregulated genes as a key feature of global gene regulation in T. pseudonana. The identity of key enzymes for carbon metabolic pathway inputs (photosynthesis) and outputs (growth and storage) reveals these clusters are organized to synchronize these processes. Coordinated transcript level responses to silicon starvation are probably driven by signals linked to cell cycle progression and shifts in photophysiology. A mechanistic understanding of how this is accomplished will aid efforts to engineer metabolism for development of algal-derived biofuels.« less

A Systems Biology Approach Reveals that Tissue Tropism to West Nile Virus Is Regulated by Antiviral Genes and Innate Immune Cellular Processes

PubMed Central

Suthar, Mehul S.; Brassil, Margaret M.; Blahnik, Gabriele; McMillan, Aimee; Ramos, Hilario J.; Proll, Sean C.; Belisle, Sarah E.; Katze, Michael G.; Gale, Michael

2013-01-01

The actions of the RIG-I like receptor (RLR) and type I interferon (IFN) signaling pathways are essential for a protective innate immune response against the emerging flavivirus West Nile virus (WNV). In mice lacking RLR or IFN signaling pathways, WNV exhibits enhanced tissue tropism, indicating that specific host factors of innate immune defense restrict WNV infection and dissemination in peripheral tissues. However, the immune mechanisms by which the RLR and IFN pathways coordinate and function to impart restriction of WNV infection are not well defined. Using a systems biology approach, we defined the host innate immune response signature and actions that restrict WNV tissue tropism. Transcriptional profiling and pathway modeling to compare WNV-infected permissive (spleen) and nonpermissive (liver) tissues showed high enrichment for inflammatory responses, including pattern recognition receptors and IFN signaling pathways, that define restriction of WNV replication in the liver. Assessment of infected livers from Mavs−/−×Ifnar−/− mice revealed the loss of expression of several key components within the natural killer (NK) cell signaling pathway, including genes associated with NK cell activation, inflammatory cytokine production, and NK cell receptor signaling. In vivo analysis of hepatic immune cell infiltrates from WT mice demonstrated that WNV infection leads to an increase in NK cell numbers with enhanced proliferation, maturation, and effector action. In contrast, livers from Mavs−/−×Ifnar−/− infected mice displayed reduced immune cell infiltration, including a significant reduction in NK cell numbers. Analysis of cocultures of dendritic and NK cells revealed both cell-intrinsic and -extrinsic roles for the RLR and IFN signaling pathways to regulate NK cell effector activity. Taken together, these observations reveal a complex innate immune signaling network, regulated by the RLR and IFN signaling pathways, that drives tissue-specific antiviral effector gene expression and innate immune cellular processes that control tissue tropism to WNV infection. PMID:23544010

Molecular mechanisms of the mammalian Hippo signaling pathway.

PubMed

Ji, Xin-yan; Zhong, Guoxuan; Zhao, Bin

2017-07-20

The Hippo pathway plays an evolutionarily conserved fundamental role in controlling organ size in multicellular organisms. Importantly, evidence from studies of patient samples and mouse models clearly indicates that deregulation of the Hippo signaling pathway plays a crucial role in the initiation and progression of many different types of human cancers. The Hippo signaling pathway is regulated by various stimuli, such as mechanical stress, G-protein coupled receptor signaling, and cellular energy status. When activated, the Hippo kinase cascade phosphorylates and inhibits the transcription co-activator YAP (Yes-associated protein), and its paralog TAZ (transcriptional coactivator with PDZ-binding motif), resulting in their cytoplasmic retention and degradation. When the Hippo signaling pathway is inactive, dephosphorylated YAP/TAZ translocate into the nucleus and activate gene transcription through binding to TEAD (TEA domain) family and other transcription factors. Such changes in gene expression promote cell proliferation and stem cell/progenitor cell self-renewal but inhibit apoptosis, thereby coordinately promote increase in organ size, tissue regeneration, and tumorigenesis. In this review, we summarize the molecular mechanisms of the mammalian Hippo signaling pathway with special emphasis on the Hippo kinase cascade and its upstream signals, the Hippo signaling pathway regulation of YAP and the mechanisms of YAP in regulation of gene transcription.

Impaired coordination between signaling pathways is revealed in human colorectal cancer using single-cell mass cytometry of archival tissue blocks

PubMed Central

Simmons, Alan J.; Scurrah, Cherie’ R.; McKinley, Eliot T.; Herring, Charles A.; Irish, Jonathan M.; Washington, Mary K.; Coffey, Robert J.; Lau, Ken S.

2016-01-01

Cellular heterogeneity poses a significant challenge to understanding tissue level phenotypes and confounds conventional bulk analyses. To facilitate the analysis of signaling at the single-cell level in human tissues, we applied mass cytometry using CyTOF (Cytometry Time-of-Flight) to formalin-fixed paraffin-embedded (FFPE) normal and diseased intestinal specimens. We developed and validated a technique called FFPE-DISSECT (Disaggregation for Intracellular Signaling in Single Epithelial Cells from Tissue), a single-cell approach for characterizing native signaling states from embedded solid tissue samples. We applied FFPE-DISSECT coupled to mass cytometry and found differential signaling by tumor necrosis factor α (TNF-α) in intestinal enterocytes, goblet cells and enteroendocrine cells, implicating the role of the downstream RAS-RAF-MEK-ERK signaling pathway in dictating goblet cell identity. In addition, application of FFPE-DISSECT, mass cytometry, and data-driven computational analyses to human colon specimens confirmed reduced differentiation in colorectal cancer (CRC) compared to normal colon, and revealed quantitative increases in inter- and intra-tissue heterogeneity in CRC with regards to the modular regulation of signaling pathways. Specifically, modular co-regulation of the kinases P38 and ERK, the translation regulator 4EBP1, and the transcription factor CREB in the proliferative compartment of the normal colon was loss in CRC, as evidenced by their impaired coordination over samplings of single cells in tissue. Our data suggest that this single-cell approach, applied in conjunction with genomic annotation, such as microsatellite instability and mutations in KRAS and BRAF, allows rapid and detailed characterization of cellular heterogeneity from clinical repositories of embedded human tissues. FFPE-DISSECT coupled of mass cytometry can be used for deriving cellular landscapes from archived patient samples, beyond CRC, and as a high resolution tool for disease characterization and subtyping. PMID:27729552

Stress Inducibility of SIRT1 and Its Role in Cytoprotection and Cancer

PubMed Central

Raynes, Rachel; Brunquell, Jessica

2013-01-01

Cells must continuously respond to stressful insults via the upregulation of cytoprotective pathways. The longevity factor and deacetylase SIRT1 plays a critical role in coordinating this cellular response to stress. SIRT1 activity and levels are regulated by cellular stressors, including metabolic, genotoxic, oxidative, and proteotoxic stress. As a stress sensor, SIRT1 impacts cell survival by deacetylating substrate proteins to drive the cell towards a cytoprotective pathway. Extreme stress conditions, however, can cause SIRT1 to lead cells down an apoptotic pathway instead. SIRT1 is frequently dysregulated in cancer cells and has been characterized to have a dual role as both an oncogene and a tumor suppressor, likely due to its pivotal function in regulating cytoprotection. Recently, the ability of SIRT1 to regulate HSF1-dependent induction of the heat shock response has highlighted another pathway through which SIRT1 can modulate cytoprotection. Activation of HSF1 results in the production of cytoprotective chaperones that can facilitate the transformed phenotype of cancer cells. In this review, we discuss the stress-dependent regulation of SIRT1. We highlight the role of SIRT1 in stress management and cytoprotection and emphasize SIRT1-dependent activation of HSF1 as a potential mechanism for cancer promotion. PMID:24020008

Dishevelled links basal body docking and orientation in ciliated epithelial cells

PubMed Central

Vladar, Eszter K.; Axelrod, Jeffrey D.

2014-01-01

Some epithelia contain cells with multiple, motile cilia that beat in a concerted fashion. New tools and experimental systems have facilitated molecular studies of cilium biogenesis and of the coordinated planar polarization of cilia that leads to their concerted motility. Recent, elegant work by Park and colleagues, using embryonic frog epidermis, demonstrates that Dishevelled (Dvl), a key regulator of both the Wnt/β-catenin and Planar Cell Polarity (PCP) pathways, controls both the docking and planar polarization of ciliary basal bodies. PMID:18819800

The therapeutic potential of cell cycle targeting in multiple myeloma.

PubMed

Maes, Anke; Menu, Eline; Veirman, Kim De; Maes, Ken; Vand Erkerken, Karin; De Bruyne, Elke

2017-10-27

Proper cell cycle progression through the interphase and mitosis is regulated by coordinated activation of important cell cycle proteins (including cyclin-dependent kinases and mitotic kinases) and several checkpoint pathways. Aberrant activity of these cell cycle proteins and checkpoint pathways results in deregulation of cell cycle progression, which is one of the key hallmarks of cancer. Consequently, intensive research on targeting these cell cycle regulatory proteins identified several candidate small molecule inhibitors that are able to induce cell cycle arrest and even apoptosis in cancer cells. Importantly, several of these cell cycle regulatory proteins have also been proposed as therapeutic targets in the plasma cell malignancy multiple myeloma (MM). Despite the enormous progress in the treatment of MM the past 5 years, MM still remains most often incurable due to the development of drug resistance. Deregulated expression of the cyclins D is observed in virtually all myeloma patients, emphasizing the potential therapeutic interest of cyclin-dependent kinase inhibitors in MM. Furthermore, other targets have also been identified in MM, such as microtubules, kinesin motor proteins, aurora kinases, polo-like kinases and the anaphase promoting complex/cyclosome. This review will provide an overview of the cell cycle proteins and checkpoint pathways deregulated in MM and discuss the therapeutic potential of targeting proteins or protein complexes involved in cell cycle control in MM.

Hypoxia-driven angiogenesis: role of tip cells and extracellular matrix scaffolding.

PubMed

Germain, Stéphane; Monnot, Catherine; Muller, Laurent; Eichmann, Anne

2010-05-01

Angiogenesis is a highly coordinated tissue remodeling process leading to blood vessel formation. Hypoxia triggers angiogenesis via induction of expression of growth factors such as vascular endothelial growth factor (VEGF). VEGF instructs endothelial cells to form tip cells, which lead outgrowing capillary sprouts, whereas Notch signaling inhibits sprout formation. Basement membrane deposition and mechanical cues from the extracellular matrix (ECM) induced by hypoxia may participate to coordinated vessel sprouting in conjunction with the VEGF and Notch signaling pathways. Hypoxia regulates ECM composition, deposition, posttranslational modifications and rearrangement. In particular, hypoxia-driven vascular remodeling is dynamically regulated through modulation of ECM-modifying enzyme activities that eventually affect both matricellular proteins and growth factor availability. Better understanding of the complex interplay between endothelial cells and soluble growth factors and mechanical factors from the ECM will certainly have significant implications for understanding the regulation of developmental and pathological angiogenesis driven by hypoxia.

Pavement cells: a model system for non-transcriptional auxin signalling and crosstalks.

PubMed

Chen, Jisheng; Wang, Fei; Zheng, Shiqin; Xu, Tongda; Yang, Zhenbiao

2015-08-01

Auxin (indole acetic acid) is a multifunctional phytohormone controlling various developmental patterns, morphogenetic processes, and growth behaviours in plants. The transcription-based pathway activated by the nuclear TRANSPORT INHIBITOR RESISTANT 1/auxin-related F-box auxin receptors is well established, but the long-sought molecular mechanisms of non-transcriptional auxin signalling remained enigmatic until very recently. Along with the establishment of the Arabidopsis leaf epidermal pavement cell (PC) as an exciting and amenable model system in the past decade, we began to gain insight into non-transcriptional auxin signalling. The puzzle-piece shape of PCs forms from intercalated or interdigitated cell growth, requiring local intra- and inter-cellular coordination of lobe and indent formation. Precise coordination of this interdigitated pattern requires auxin and an extracellular auxin sensing system that activates plasma membrane-associated Rho GTPases from plants and subsequent downstream events regulating cytoskeletal reorganization and PIN polarization. Apart from auxin, mechanical stress and cytokinin have been shown to affect PC interdigitation, possibly by interacting with auxin signals. This review focuses upon signalling mechanisms for cell polarity formation in PCs, with an emphasis on non-transcriptional auxin signalling in polarized cell expansion and pattern formation and how different auxin pathways interplay with each other and with other signals. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Pericytes of the neurovascular unit: Key functions and signaling pathways

PubMed Central

Sweeney, Melanie D.; Ayyadurai, Shiva; Zlokovic, Berislav V.

2017-01-01

Pericytes are vascular mural cells embedded in the basement membrane of blood microvessels. They extend their processes along capillaries, pre-capillary arterioles, and post-capillary venules. The central nervous system (CNS) pericytes are uniquely positioned within the neurovascular unit between endothelial cells, astrocytes, and neurons. They integrate, coordinate, and process signals from their neighboring cells to generate diverse functional responses that are critical for CNS functions in health and disease including regulation of the blood-brain barrier permeability, angiogenesis, clearance of toxic metabolites, capillary hemodynamic responses, neuroinflammation, and stem cell activity. Here, we examine the key signaling pathways between pericytes and their neighboring endothelial cells, astrocytes, and neurons that control neurovascular functions. We also review the role of pericytes in different CNS disorders including rare monogenic diseases and complex neurological disorders such as Alzheimer's disease and brain tumors. Finally, we discuss directions for future studies. PMID:27227366

Metabolic and Epigenetic Coordination of T Cell and Macrophage Immunity.

PubMed

Phan, Anthony T; Goldrath, Ananda W; Glass, Christopher K

2017-05-16

Recognition of pathogens by innate and adaptive immune cells instructs rapid alterations of cellular processes to promote effective resolution of infection. To accommodate increased bioenergetic and biosynthetic demands, metabolic pathways are harnessed to maximize proliferation and effector molecule production. In parallel, activation initiates context-specific gene-expression programs that drive effector functions and cell fates that correlate with changes in epigenetic landscapes. Many chromatin- and DNA-modifying enzymes make use of substrates and cofactors that are intermediates of metabolic pathways, providing potential cross talk between metabolism and epigenetic regulation of gene expression. In this review, we discuss recent studies of T cells and macrophages supporting a role for metabolic activity in integrating environmental signals with activation-induced gene-expression programs through modulation of the epigenome and speculate as to how this may influence context-specific macrophage and T cell responses to infection. Copyright © 2017 Elsevier Inc. All rights reserved.

Metabolic and epigenetic coordination of T cell and Macrophage immunity

PubMed Central

Phan, Anthony T.; Goldrath, Ananda W.; Glass, Christopher K.

2017-01-01

Recognition of pathogens by innate and adaptive immune cells instructs rapid alterations of cellular processes to promote effective resolution of infection. To accommodate increased bioenergetic and biosynthetic demands, metabolic pathways are harnessed to maximize proliferation and effector molecule production. In parallel, activation initiates context-specific gene-expression programs that drive effector functions and cell fates that correlate with changes in epigenetic landscapes. Many chromatin- and DNA-modifying enzymes make use of substrates and cofactors that are intermediates of metabolic pathways, providing potential cross talk between metabolism and epigenetic regulation of gene expression. In this review, we discuss recent studies of T cells and macrophages supporting a role for metabolic activity in integrating environmental signals with activation-induced gene-expression programs through modulation of the epigenome and speculate as to how this may influence context-specific macrophage and T cell responses to infection. PMID:28514673

Emerging Imaging and Genomic Tools for Developmental Systems Biology.

PubMed

Liu, Zhe; Keller, Philipp J

2016-03-21

Animal development is a complex and dynamic process orchestrated by exquisitely timed cell lineage commitment, divisions, migration, and morphological changes at the single-cell level. In the past decade, extensive genetic, stem cell, and genomic studies provided crucial insights into molecular underpinnings and the functional importance of genetic pathways governing various cellular differentiation processes. However, it is still largely unknown how the precise coordination of these pathways is achieved at the whole-organism level and how the highly regulated spatiotemporal choreography of development is established in turn. Here, we discuss the latest technological advances in imaging and single-cell genomics that hold great promise for advancing our understanding of this intricate process. We propose an integrated approach that combines such methods to quantitatively decipher in vivo cellular dynamic behaviors and their underlying molecular mechanisms at the systems level with single-cell, single-molecule resolution. Copyright © 2016 Elsevier Inc. All rights reserved.

Dissecting Cell-Fate Determination Through Integrated Mathematical Modeling of the ERK/MAPK Signaling Pathway.

PubMed

Shin, Sung-Young; Nguyen, Lan K

2017-01-01

The past three decades have witnessed an enormous progress in the elucidation of the ERK/MAPK signaling pathway and its involvement in various cellular processes. Because of its importance and complex wiring, the ERK pathway has been an intensive subject for mathematical modeling, which facilitates the unraveling of key dynamic properties and behaviors of the pathway. Recently, however, it became evident that the pathway does not act in isolation but closely interacts with many other pathways to coordinate various cellular outcomes under different pathophysiological contexts. This has led to an increasing number of integrated, large-scale models that link the ERK pathway to other functionally important pathways. In this chapter, we first discuss the essential steps in model development and notable models of the ERK pathway. We then use three examples of integrated, multipathway models to investigate how crosstalk of ERK signaling with other pathways regulates cell-fate decision-making in various physiological and disease contexts. Specifically, we focus on ERK interactions with the phosphoinositide-3 kinase (PI3K), c-Jun N-terminal kinase (JNK), and β-adrenergic receptor (β-AR) signaling pathways. We conclude that integrated modeling in combination with wet-lab experimentation have been and will be instrumental in gaining an in-depth understanding of ERK signaling in multiple biological contexts.

T cell exit from quiescence and differentiation into Th2 cells depend on Raptor-mTORC1-mediated metabolic programming

PubMed Central

Yang, Kai; Shrestha, Sharad; Zeng, Hu; Karmaus, Peer W.F.; Neale, Geoffrey; Vogel, Peter; Guertin, David A.; Lamb, Richard F.; Chi, Hongbo

2014-01-01

SUMMARY Naïve T cells respond to antigen stimulation by exiting from quiescence and initiating clonal expansion and functional differentiation, but the control mechanism is elusive. Here we describe that Raptor-mTORC1-dependent metabolic programming is a central determinant of this transitional process. Loss of Raptor abrogated T cell priming and Th2 cell differentiation, although Raptor function is less important for continuous proliferation of actively cycling cells. mTORC1 coordinated multiple metabolic programs in T cells including glycolysis, lipid synthesis and oxidative phosphorylation to mediate antigen-triggered exit from quiescence. mTORC1 further linked glucose metabolism to the initiation of Th2 cell differentiation by orchestrating cytokine receptor expression and cytokine responsiveness. Activation of Raptor-mTORC1 integrated T cell receptor and CD28 co-stimulatory signals in antigen-stimulated T cells. Our studies identify a Raptor-mTORC1-dependent pathway linking signal-dependent metabolic reprogramming to quiescence exit, and this in turn coordinates lymphocyte activation and fate decisions in adaptive immunity. PMID:24315998

Clathrin-independent internalization and recycling

PubMed Central

Gong, Qiang; Huntsman, Christopher; Ma, Dzwokai

2008-01-01

Abstract The functionality of receptor and channel proteins depends directly upon their expression level on the plasma membrane. Therefore, the ability to selectively adjust the surface level of a particular receptor or channel protein is pivotal to many cellular signalling events. The internalization and recycling pathway plays a major role in the regulation of protein surface level, and thus has been a focus of research for many years. Although several endocytic pathways have been identified, most of our knowledge has come from the clathrin-dependent pathway, while the other pathways remain much less well defined. Considering that clathrin-independent internalization may account for as much as 50% of the total endocytic activity in the cell, the lack of such knowledge constitutes a major gap in our efforts to understand how different internalization pathways are utilized and co-ordinated. Recent studies have provided valuable insights into this area, yet many more questions still remain. In this review, we will give a panoramic introduction to the current knowledge of various internalization and recycling pathways, with an emphasis on the latest findings that have broadened our view of the clathrin-independent pathways. We will also dedicate one section to the emerging studies of the clathrin-independent internalization pathways in neuronal cells. PMID:18039352

Molecular Genetic Traits Influencing Maize Endosperm Development and Value: Closeout Report for DOE Grant DE-FG02-96ER20242

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

Brian A. Larkins

2012-09-12

Development of the endosperm in cereal grasses entails different phases characterized by cell division, endoreduplication, accumulation of storage metabolites and cell death, which need to be carried out in an orderly fashion. While correct regulation of the cell cycle plays an essential role in endosperm development, the key regulatory factors and how the cell cycle interfaces with other pathways in this developmental context are largely unknown. We investigated the cyclin-dependent kinase (CDK)-retinoblastoma pathway and how it controls the cell cycle and coordinates it with other processes during maize endosperm development. Retinoblastoma-related (RBR) proteins may be inactivated through CDK-mediated phosphorylation, butmore » the identity of the responsible kinase in maize is unknown. We have previously shown that down-regulation of CDKA;1 severely inhibits the endoreduplication cell cycle and suggested that CDK may be an up-stream regulator of the retinoblastoma pathway. We discovered two types of maize RBR genes, RBR1 and RBR3, which differ in terms of structure, regulation and function. Phylogenetic analyses indicate that these genes may be distinctive features of the Poaceae. We found that RBR3 plays a positive rather than a negative role in DNA replication, cell transformation, and the expression of the minichromosome maintenance (MCM)2-7 family of DNA replication factors. These features are a paradigm shift in RBR gene function and appear to be unique within the RBR gene family. They suggest the existence in maize and related cereal crops of specific RBR/E2F-dependent pathways impinging on the cell cycle and development. RBR1 was down-regulated in transgenic endosperm using RNAi approaches. This resulted in the de-repression of a number of down-stream E2F targets, including RBR3, the MCM2-7 gene family, DNA methyltransferase (MET)1, CDKB;1, and the recently identified RBR4 gene. It also increased endosperm ploidy levels, stimulated the production of a larger number of cells, reduced the average cell size, and promoted programmed cell death. To test whether CDKA;1 inhibits RBR1 (through phosphorylation) in the pathway that leads to DNA synthesis and endoreduplication, the two CDKA;1 and RBR1 down-regulated mutants were crossed and their progeny analyzed. Our results indicate that CDKA;1 controls endoreduplication through an RBR1-dependent pathway. However, the ability of RBR1 to repress gene expression programs is independent from CDKA1, suggesting the presence of two differently regulated RBR1 activities in developing endosperm. One type of RBR1 activity controls E2F-dependent gene expression and is largely independent from CDKA;1, while another suppresses endoreduplication and can be inhibited by CDKA;1. In addition, RBR1 is part of a regulatory feedback loop that impinges on CDK activity. Together, these results indicate that the CDKA;1-RBR1 pathway integrates and controls different processes associated with endosperm development. Genome-wide analyses of the transcriptome, metabolome, and epigenetic mechanisms to understand how the cell cycle is coordinated with other pathways at a systems biology level are currently underway.« less

Murine craniofacial development requires Hdac3-mediated repression of Msx gene expression.

PubMed

Singh, Nikhil; Gupta, Mudit; Trivedi, Chinmay M; Singh, Manvendra K; Li, Li; Epstein, Jonathan A

2013-05-15

Craniofacial development is characterized by reciprocal interactions between neural crest cells and neighboring cell populations of ectodermal, endodermal and mesodermal origin. Various genetic pathways play critical roles in coordinating the development of cranial structures by modulating the growth, survival and differentiation of neural crest cells. However, the regulation of these pathways, particularly at the epigenomic level, remains poorly understood. Using murine genetics, we show that neural crest cells exhibit a requirement for the class I histone deacetylase Hdac3 during craniofacial development. Mice in which Hdac3 has been conditionally deleted in neural crest demonstrate fully penetrant craniofacial abnormalities, including microcephaly, cleft secondary palate and dental hypoplasia. Consistent with these abnormalities, we observe dysregulation of cell cycle genes and increased apoptosis in neural crest structures in mutant embryos. Known regulators of cell cycle progression and apoptosis in neural crest, including Msx1, Msx2 and Bmp4, are upregulated in Hdac3-deficient cranial mesenchyme. These results suggest that Hdac3 serves as a critical regulator of craniofacial morphogenesis, in part by repressing core apoptotic pathways in cranial neural crest cells. Copyright © 2013 Elsevier Inc. All rights reserved.

Noncanonical ATM Activation and Signaling in Response to Transcription-Blocking DNA Damage.

PubMed

Marteijn, Jurgen A; Vermeulen, Wim; Tresini, Maria

2017-01-01

Environmental genotoxins and metabolic byproducts generate DNA lesions that can cause genomic instability and disrupt tissue homeostasis. To ensure genomic integrity, cells employ mechanisms that convert signals generated by stochastic DNA damage into organized responses, including activation of repair systems, cell cycle checkpoints, and apoptotic mechanisms. DNA damage response (DDR) signaling pathways coordinate these responses and determine cellular fates in part, by transducing signals that modulate RNA metabolism. One of the master DDR coordinators, the Ataxia Telangiectasia Mutated (ATM) kinase, has a fundamental role in mediating DNA damage-induced changes in mRNA synthesis. ATM acts by modulating a variety of RNA metabolic pathways including nascent RNA splicing, a process catalyzed by the spliceosome. Interestingly, ATM and the spliceosome influence each other's activity in a reciprocal manner by a pathway that initiates when transcribing RNA polymerase II (RNAPII) encounters DNA lesions that prohibit forward translocation. In response to stalling of RNAPII assembly of late-stage spliceosomes is disrupted resulting in increased splicing factor mobility. Displacement of spliceosomes from lesion-arrested RNA polymerases facilitates formation of R-loops between the nascent RNA and DNA adjacent to the transcription bubble. R-loops signal for noncanonical ATM activation which in quiescent cells occurs in absence of detectable dsDNA breaks. In turn, activated ATM signals to regulate spliceosome dynamics and AS genome wide.This chapter describes the use of fluorescence microscopy methods that can be used to evaluate noncanonical ATM activation by transcription-blocking DNA damage. First, we present an immunofluorescence-detection method that can be used to evaluate ATM activation by autophosphorylation, in fixed cells. Second, we present a protocol for Fluorescence Recovery After Photobleaching (FRAP) of GFP-tagged splicing factors, a highly sensitive and reproducible readout to measure in living cells, the ATM influence on the spliceosome. These approaches have been extensively used in our laboratory for a number of cell lines of various origins and are particularly informative when used in primary cells that can be synchronized in quiescence, to avoid generation of replication stress-induced dsDNA breaks and consequent ATM activation through its canonical pathway.

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

Railo, Antti; Nagy, Irina I.; Kilpelaeinen, Pekka

The Wnt family of glycoprotein growth factors controls a number of central cellular processes such as proliferation, differentiation and ageing. All the Wnt proteins analyzed so far either activate or inhibit the canonical {beta}-catenin signaling pathway that regulates transcription of the target genes. In addition, some of them activate noncanonical signaling pathways that involve components such as the JNK, heterotrimeric G proteins, protein kinase C, and calmodulin-dependent protein kinase II, although the precise signaling mechanisms are only just beginning to be revealed. We demonstrate here that Wnt-11 signaling is sufficient to inhibit not only the canonical {beta}-catenin mediated Wnt signalingmore » but also JNK/AP-1 and NF-{kappa}B signaling in the CHO cells, thus serving as a noncanonical Wnt ligand in this system. Inhibition of the JNK/AP-1 pathway is mediated in part by the MAPK kinase MKK4 and Akt. Moreover, protein kinase C is involved in the regulation of JNK/AP-1 by Wnt-11, but not of the NF-{kappa}B pathway. Consistent with the central role of Akt, JNK and NF-{kappa}B in cell survival and stress responses, Wnt-11 signaling promotes cell viability. Hence Wnt-11 is involved in coordination of key signaling pathways.« less

Ties that bind: the integration of plastid signalling pathways in plant cell metabolism.

PubMed

Brunkard, Jacob O; Burch-Smith, Tessa M

2018-04-13

Plastids are critical organelles in plant cells that perform diverse functions and are central to many metabolic pathways. Beyond their major roles in primary metabolism, of which their role in photosynthesis is perhaps best known, plastids contribute to the biosynthesis of phytohormones and other secondary metabolites, store critical biomolecules, and sense a range of environmental stresses. Accordingly, plastid-derived signals coordinate a host of physiological and developmental processes, often by emitting signalling molecules that regulate the expression of nuclear genes. Several excellent recent reviews have provided broad perspectives on plastid signalling pathways. In this review, we will highlight recent advances in our understanding of chloroplast signalling pathways. Our discussion focuses on new discoveries illuminating how chloroplasts determine life and death decisions in cells and on studies elucidating tetrapyrrole biosynthesis signal transduction networks. We will also examine the role of a plastid RNA helicase, ISE2, in chloroplast signalling, and scrutinize intriguing results investigating the potential role of stromules in conducting signals from the chloroplast to other cellular locations. © 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

Integrated compensatory network is activated in the absence of NCC phosphorylation.

PubMed

Grimm, P Richard; Lazo-Fernandez, Yoskaly; Delpire, Eric; Wall, Susan M; Dorsey, Susan G; Weinman, Edward J; Coleman, Richard; Wade, James B; Welling, Paul A

2015-05-01

Thiazide diuretics are used to treat hypertension; however, compensatory processes in the kidney can limit antihypertensive responses to this class of drugs. Here, we evaluated compensatory pathways in SPAK kinase-deficient mice, which are unable to activate the thiazide-sensitive sodium chloride cotransporter NCC (encoded by Slc12a3). Global transcriptional profiling, combined with biochemical, cell biological, and physiological phenotyping, identified the gene expression signature of the response and revealed how it establishes an adaptive physiology. Salt reabsorption pathways were created by the coordinate induction of a multigene transport system, involving solute carriers (encoded by Slc26a4, Slc4a8, and Slc4a9), carbonic anhydrase isoforms, and V-type H⁺-ATPase subunits in pendrin-positive intercalated cells (PP-ICs) and ENaC subunits in principal cells (PCs). A distal nephron remodeling process and induction of jagged 1/NOTCH signaling, which expands the cortical connecting tubule with PCs and replaces acid-secreting α-ICs with PP-ICs, were partly responsible for the compensation. Salt reabsorption was also activated by induction of an α-ketoglutarate (α-KG) paracrine signaling system. Coordinate regulation of a multigene α-KG synthesis and transport pathway resulted in α-KG secretion into pro-urine, as the α-KG-activated GPCR (Oxgr1) increased on the PP-IC apical surface, allowing paracrine delivery of α-KG to stimulate salt transport. Identification of the integrated compensatory NaCl reabsorption mechanisms provides insight into thiazide diuretic efficacy.

Integrated compensatory network is activated in the absence of NCC phosphorylation

PubMed Central

Grimm, P. Richard; Lazo-Fernandez, Yoskaly; Delpire, Eric; Wall, Susan M.; Dorsey, Susan G.; Weinman, Edward J.; Coleman, Richard; Wade, James B.; Welling, Paul A.

2015-01-01

Thiazide diuretics are used to treat hypertension; however, compensatory processes in the kidney can limit antihypertensive responses to this class of drugs. Here, we evaluated compensatory pathways in SPAK kinase–deficient mice, which are unable to activate the thiazide-sensitive sodium chloride cotransporter NCC (encoded by Slc12a3). Global transcriptional profiling, combined with biochemical, cell biological, and physiological phenotyping, identified the gene expression signature of the response and revealed how it establishes an adaptive physiology. Salt reabsorption pathways were created by the coordinate induction of a multigene transport system, involving solute carriers (encoded by Slc26a4, Slc4a8, and Slc4a9), carbonic anhydrase isoforms, and V-type H+-ATPase subunits in pendrin-positive intercalated cells (PP-ICs) and ENaC subunits in principal cells (PCs). A distal nephron remodeling process and induction of jagged 1/NOTCH signaling, which expands the cortical connecting tubule with PCs and replaces acid-secreting α-ICs with PP-ICs, were partly responsible for the compensation. Salt reabsorption was also activated by induction of an α-ketoglutarate (α-KG) paracrine signaling system. Coordinate regulation of a multigene α-KG synthesis and transport pathway resulted in α-KG secretion into pro-urine, as the α-KG–activated GPCR (Oxgr1) increased on the PP-IC apical surface, allowing paracrine delivery of α-KG to stimulate salt transport. Identification of the integrated compensatory NaCl reabsorption mechanisms provides insight into thiazide diuretic efficacy. PMID:25893600

Orchestrating liver development.

PubMed

Gordillo, Miriam; Evans, Todd; Gouon-Evans, Valerie

2015-06-15

The liver is a central regulator of metabolism, and liver failure thus constitutes a major health burden. Understanding how this complex organ develops during embryogenesis will yield insights into how liver regeneration can be promoted and how functional liver replacement tissue can be engineered. Recent studies of animal models have identified key signaling pathways and complex tissue interactions that progressively generate liver progenitor cells, differentiated lineages and functional tissues. In addition, progress in understanding how these cells interact, and how transcriptional and signaling programs precisely coordinate liver development, has begun to elucidate the molecular mechanisms underlying this complexity. Here, we review the lineage relationships, signaling pathways and transcriptional programs that orchestrate hepatogenesis. © 2015. Published by The Company of Biologists Ltd.

MAP kinase pathways in the yeast Saccharomyces cerevisiae

NASA Technical Reports Server (NTRS)

Gustin, M. C.; Albertyn, J.; Alexander, M.; Davenport, K.; McIntire, L. V. (Principal Investigator)

1998-01-01

A cascade of three protein kinases known as a mitogen-activated protein kinase (MAPK) cascade is commonly found as part of the signaling pathways in eukaryotic cells. Almost two decades of genetic and biochemical experimentation plus the recently completed DNA sequence of the Saccharomyces cerevisiae genome have revealed just five functionally distinct MAPK cascades in this yeast. Sexual conjugation, cell growth, and adaptation to stress, for example, all require MAPK-mediated cellular responses. A primary function of these cascades appears to be the regulation of gene expression in response to extracellular signals or as part of specific developmental processes. In addition, the MAPK cascades often appear to regulate the cell cycle and vice versa. Despite the success of the gene hunter era in revealing these pathways, there are still many significant gaps in our knowledge of the molecular mechanisms for activation of these cascades and how the cascades regulate cell function. For example, comparison of different yeast signaling pathways reveals a surprising variety of different types of upstream signaling proteins that function to activate a MAPK cascade, yet how the upstream proteins actually activate the cascade remains unclear. We also know that the yeast MAPK pathways regulate each other and interact with other signaling pathways to produce a coordinated pattern of gene expression, but the molecular mechanisms of this cross talk are poorly understood. This review is therefore an attempt to present the current knowledge of MAPK pathways in yeast and some directions for future research in this area.

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

Plasmodesmata in integrated cell signalling: insights from development and environmental signals and stresses

PubMed Central

Sager, Ross; Lee, Jung-Youn

2014-01-01

To survive as sedentary organisms built of immobile cells, plants require an effective intercellular communication system, both locally between neighbouring cells within each tissue and systemically across distantly located organs. Such a system enables cells to coordinate their intracellular activities and produce concerted responses to internal and external stimuli. Plasmodesmata, membrane-lined intercellular channels, are essential for direct cell-to-cell communication involving exchange of diffusible factors, including signalling and information molecules. Recent advances corroborate that plasmodesmata are not passive but rather highly dynamic channels, in that their density in the cell walls and gating activities are tightly linked to developmental and physiological processes. Moreover, it is becoming clear that specific hormonal signalling pathways play crucial roles in relaying primary cellular signals to plasmodesmata. In this review, we examine a number of studies in which plasmodesmal structure, occurrence, and/or permeability responses are found to be altered upon given cellular or environmental signals, and discuss common themes illustrating how plasmodesmal regulation is integrated into specific cellular signalling pathways. PMID:25262225

YAP regulates neuronal differentiation through Sonic hedgehog signaling pathway

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

Lin, Yi-Ting; Ding, Jing-Ya; Li, Ming-Yang

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 tomore » 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-overexpression phenotype in P19 cells. Black-Right-Pointing-Pointer Knockdown of Gli2 rescues the Yap-overexpression phenotype in cortical progenitors.« less

Epigenetic determinants of ovarian clear cell carcinoma biology

PubMed Central

Yamaguchi, Ken; Huang, Zhiqing; Matsumura, Noriomi; Mandai, Masaki; Okamoto, Takako; Baba, Tsukasa; Konishi, Ikuo; Berchuck, Andrew; Murphy, Susan K.

2015-01-01

Targeted approaches have revealed frequent epigenetic alterations in ovarian cancer, but the scope and relation of these changes to histologic subtype of disease is unclear. Genome-wide methylation and expression data for 14 clear cell carcinoma (CCC), 32 non-CCC, and 4 corresponding normal cell lines were generated to determine how methylation profiles differ between cells of different histological derivations of ovarian cancer. Consensus clustering showed that CCC is epigenetically distinct. Inverse relationships between expression and methylation in CCC were identified, suggesting functional regulation by methylation, and included 22 hypomethylated (UM) genes and 276 hypermethylated (HM) genes. Categorical and pathway analyses indicated that the CCC-specific UM genes were involved in response to stress and many contain hepatocyte nuclear factor (HNF) 1 binding sites, while the CCC-specific HM genes included members of the estrogen receptor alpha (ERalpha) network and genes involved in tumor development. We independently validated the methylation status of 17 of these pathway-specific genes, and confirmed increased expression of HNF1 network genes and repression of ERalpha pathway genes in CCC cell lines and primary cancer tissues relative to non-CCC specimens. Treatment of three CCC cell lines with the demethylating agent Decitabine significantly induced expression for all five genes analyzed. Coordinate changes in pathway expression were confirmed using two primary ovarian cancer datasets (p<0.0001 for both). Our results suggest that methylation regulates specific pathways and biological functions in CCC, with hypomethylation influencing the characteristic biology of the disease while hypermethylation contributes to the carcinogenic process. PMID:24382740

Disruption of stomatal lineage signaling or transcriptional regulators has differential effects on mesophyll development, but maintains coordination of gas exchange.

PubMed

Dow, Graham J; Berry, Joseph A; Bergmann, Dominique C

2017-10-01

Stomata are simultaneously tasked with permitting the uptake of carbon dioxide for photosynthesis while limiting water loss from the plant. This process is mainly regulated by guard cell control of the stomatal aperture, but recent advancements have highlighted the importance of several genes that control stomatal development. Using targeted genetic manipulations of the stomatal lineage and a combination of gas exchange and microscopy techniques, we show that changes in stomatal development of the epidermal layer lead to coupled changes in the underlying mesophyll tissues. This coordinated response tends to match leaf photosynthetic potential (V cmax ) with gas-exchange capacity (g smax ), and hence the uptake of carbon dioxide for water lost. We found that different genetic regulators systematically altered tissue coordination in separate ways: the transcription factor SPEECHLESS (SPCH) primarily affected leaf size and thickness, whereas peptides in the EPIDERMAL PATTERNING FACTOR (EPF) family altered cell density in the mesophyll. It was also determined that interlayer coordination required the cell-surface receptor TOO MANY MOUTHS (TMM). These results demonstrate that stomata-specific regulators can alter mesophyll properties, which provides insight into how molecular pathways can organize leaf tissues to coordinate gas exchange and suggests new strategies for improving plant water-use efficiency. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

Improving care coordination using organisational routines.

PubMed

Prætorius, Thim

2016-01-01

The purpose of this paper is to systematically apply theory of organisational routines to standardised care pathways. The explanatory power of routines is used to address open questions in the care pathway literature about their coordinating and organising role, the way they change and can be replicated, the way they are influenced by the organisation and the way they influence health care professionals. Theory of routines is systematically applied to care pathways in order to develop theoretically derived propositions. Care pathways mirror routines by being recurrent, collective and embedded and specific to an organisation. In particular, care pathways resemble standard operating procedures that can give rise to recurrent collective action patterns. In all, 11 propositions related to five categories are proposed by building on these insights: care pathways and coordination, change, replication, the organisation and health care professionals. Research limitations/implications - The paper is conceptual and uses care pathways as illustrative instances of hospital routines. The propositions provide a starting point for empirical research. The analysis highlights implications that health care professionals and managers have to consider in relation to coordination, change, replication, the way the organisation influences care pathways and the way care pathways influence health care professionals. Originality/value - Theory on organisational routines offers fundamental, yet unexplored, insights into hospital processes, including in particular care coordination.

The JNK/AP-1 pathway upregulates expression of the recycling endosome rab11a gene in B cells transformed by Theileria.

PubMed

Lizundia, Regina; Chaussepied, Marie; Naissant, Bernina; Masse, Guillemette X; Quevillon, Emmanuel; Michel, Fréderique; Monier, Solange; Weitzman, Jonathan B; Langsley, Gordon

2007-08-01

Lymphocyte transformation induced by Theileria parasites involves constitutive activation of c-Jun N-terminal kinase (JNK) and the AP-1 transcription factor. We found that JNK/AP-1 activation is associated with elevated levels of Rab11 protein in Theileria-transformed B cells. We show that AP-1 regulates rab11a promoter activity in B cells and that the induction of c-Jun activity in mouse fibroblasts also leads to increased transcription of the endogenous rab11a gene, consistent with it being an AP-1 target. Pharmacological inhibition of the JNK pathway reduced Rab11 protein levels and endosome recycling of transferrin receptor (TfR) and siRNA knockdown of JNK1 and Rab11A levels also reduced TfR surface expression. We propose a model, where activation of the JNK/AP-1 pathway during cell transformation might assure that the regulation of recycling endosomes is co-ordinated with cell-cycle progression. This might be achieved via the simultaneous upregulation of the cell cycle machinery (e.g. cyclin D1) and the recycling endosome regulators (e.g. Rab11A).

Apoptotic cells can induce non-autonomous apoptosis through the TNF pathway

PubMed Central

Pérez-Garijo, Ainhoa; Fuchs, Yaron; Steller, Hermann

2013-01-01

Apoptotic cells can produce signals to instruct cells in their local environment, including ones that stimulate engulfment and proliferation. We identified a novel mode of communication by which apoptotic cells induce additional apoptosis in the same tissue. Strong induction of apoptosis in one compartment of the Drosophila wing disc causes apoptosis of cells in the other compartment, indicating that dying cells can release long-range death factors. We identified Eiger, the Drosophila tumor necrosis factor (TNF) homolog, as the signal responsible for apoptosis-induced apoptosis (AiA). Eiger is produced in apoptotic cells and, through activation of the c-Jun N-terminal kinase (JNK) pathway, is able to propagate the initial apoptotic stimulus. We also show that during coordinated cell death of hair follicle cells in mice, TNF-α is expressed in apoptotic cells and is required for normal cell death. AiA provides a mechanism to explain cohort behavior of dying cells that is seen both in normal development and under pathological conditions. DOI: http://dx.doi.org/10.7554/eLife.01004.001 PMID:24066226

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

PubMed Central

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.

2015-01-01

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. PMID:26311851

NOVEL ROLES FOR INSULIN RECEPTOR (IR) IN ADIPOCYTES AND SKELETAL MUSCLE CELLS VIA NEW AND UNEXPECTED SUBSTRATES

PubMed Central

Ramalingam, Latha; Oh, Eunjin; Thurmond, Debbie C.

2012-01-01

The insulin signaling pathway regulates whole-body glucose homeostasis by transducing extracellular signals from the insulin receptor (IR) to downstream intracellular targets, thus coordinating a multitude of biological functions. Dysregulation of IR or its signal transduction is associated with insulin resistance, which may culminate in type 2 diabetes (T2D). Following initial stimulation of IR, insulin signaling diverges into different pathways, activating multiple substrates which have roles in various metabolic and cellular processes. The integration of multiple pathways arising from IR activation continues to expand as new IR substrates are identified and characterized. Accordingly, our review will focus on roles for IR substrates as they pertain to three primary areas: Metabolism/glucose uptake, Mitogenesis/growth, and Aging/Longevity. While IR functions in a seemingly pleotropic manner in many cell types, through these three main roles in fat and skeletal muscle cells, IR multi-tasks to regulate whole-body glucose homeostasis to impact healthspan and lifespan. PMID:23052216

Comprehensive analysis of pathway or functionally related gene expression in the National Cancer Institute's anticancer screen.

PubMed

Huang, Ruili; Wallqvist, Anders; Covell, David G

2006-03-01

We have analyzed the level of gene coregulation, using gene expression patterns measured across the National Cancer Institute's 60 tumor cell panels (NCI(60)), in the context of predefined pathways or functional categories annotated by KEGG (Kyoto Encyclopedia of Genes and Genomes), BioCarta, and GO (Gene Ontology). Statistical methods were used to evaluate the level of gene expression coherence (coordinated expression) by comparing intra- and interpathway gene-gene correlations. Our results show that gene expression in pathways, or groups of functionally related genes, has a significantly higher level of coherence than that of a randomly selected set of genes. Transcriptional-level gene regulation appears to be on a "need to be" basis, such that pathways comprising genes encoding closely interacting proteins and pathways responsible for vital cellular processes or processes that are related to growth or proliferation, specifically in cancer cells, such as those engaged in genetic information processing, cell cycle, energy metabolism, and nucleotide metabolism, tend to be more modular (lower degree of gene sharing) and to have genes significantly more coherently expressed than most signaling and regular metabolic pathways. Hierarchical clustering of pathways based on their differential gene expression in the NCI(60) further revealed interesting interpathway communications or interactions indicative of a higher level of pathway regulation. The knowledge of the nature of gene expression regulation and biological pathways can be applied to understanding the mechanism by which small drug molecules interfere with biological systems.

A Quantitative RNAi Screen for JNK Modifiers Identifies Pvr as a Novel Regulator of Drosophila Immune Signaling

PubMed Central

Bond, David; Foley, Edan

2009-01-01

Drosophila melanogaster responds to gram-negative bacterial challenges through the IMD pathway, a signal transduction cassette that is driven by the coordinated activities of JNK, NF-κB and caspase modules. While many modifiers of NF-κB activity were identified in cell culture and in vivo assays, the regulatory apparatus that determines JNK inputs into the IMD pathway is relatively unexplored. In this manuscript, we present the first quantitative screen of the entire genome of Drosophila for novel regulators of JNK activity in the IMD pathway. We identified a large number of gene products that negatively or positively impact on JNK activation in the IMD pathway. In particular, we identified the Pvr receptor tyrosine kinase as a potent inhibitor of JNK activation. In a series of in vivo and cell culture assays, we demonstrated that activation of the IMD pathway drives JNK-dependent expression of the Pvr ligands, Pvf2 and Pvf3, which in turn act through the Pvr/ERK MAP kinase pathway to attenuate the JNK and NF-κB arms of the IMD pathway. Our data illuminate a poorly understood arm of a critical and evolutionarily conserved innate immune response. Furthermore, given the pleiotropic involvement of JNK in eukaryotic cell biology, we believe that many of the novel regulators identified in this screen are of interest beyond immune signaling. PMID:19893628

Hippo Signaling in Mitosis: An Updated View in Light of the MEN Pathway.

PubMed

Hergovich, Alexander

2017-01-01

The Hippo pathway is an essential tumor suppressor signaling network that coordinates cell proliferation, death, and differentiation in higher eukaryotes. Intriguingly, the core components of the Hippo pathway are conserved from yeast to man, with the yeast analogs of mammalian MST1/2 (fly Hippo), MOB1 (fly Mats), LATS1/2 (fly Warts), and NDR1/2 (fly Tricornered) functioning as essential components of the mitotic exit network (MEN). Here, we update our previous summary of mitotic functions of Hippo core components in Drosophila melanogaster and mammals, with particular emphasis on similarities between the yeast MEN pathway and mitotic Hippo signaling. Mitotic functions of YAP and TAZ, the two main effectors of Hippo signaling, are also discussed.

AMP kinase promotes glioblastoma bioenergetics and tumour growth.

PubMed

Chhipa, Rishi Raj; Fan, Qiang; Anderson, Jane; Muraleedharan, Ranjithmenon; Huang, Yan; Ciraolo, Georgianne; Chen, Xiaoting; Waclaw, Ronald; Chow, Lionel M; Khuchua, Zaza; Kofron, Matthew; Weirauch, Matthew T; Kendler, Ady; McPherson, Christopher; Ratner, Nancy; Nakano, Ichiro; Dasgupta, Nupur; Komurov, Kakajan; Dasgupta, Biplab

2018-06-18

Stress is integral to tumour evolution, and cancer cell survival depends on stress management. We found that cancer-associated stress chronically activates the bioenergetic sensor AMP kinase (AMPK) and, to survive, tumour cells hijack an AMPK-regulated stress response pathway conserved in normal cells. Analysis of The Cancer Genome Atlas data revealed that AMPK isoforms are highly expressed in the lethal human cancer glioblastoma (GBM). We show that AMPK inhibition reduces viability of patient-derived GBM stem cells (GSCs) and tumours. In stressed (exercised) skeletal muscle, AMPK is activated to cooperate with CREB1 (cAMP response element binding protein-1) and promote glucose metabolism. We demonstrate that oncogenic stress chronically activates AMPK in GSCs that coopt the AMPK-CREB1 pathway to coordinate tumour bioenergetics through the transcription factors HIF1α and GABPA. Finally, we show that adult mice tolerate systemic deletion of AMPK, supporting the use of AMPK pharmacological inhibitors in the treatment of GBM.

Switching of metabolic programs in response to light availability is an essential function of the cyanobacterial circadian output pathway

PubMed Central

Puszynska, Anna M; O'Shea, Erin K

2017-01-01

The transcription factor RpaA is the master regulator of circadian transcription in cyanobacteria, driving genome-wide oscillations in mRNA abundance. Deletion of rpaA has no effect on viability in constant light conditions, but renders cells inviable in cycling conditions when light and dark periods alternate. We investigated the mechanisms underlying this viability defect, and demonstrate that the rpaA- strain cannot maintain appropriate energy status at night, does not accumulate carbon reserves during the day, and is defective in transcription of genes crucial for utilization of carbohydrate stores at night. Reconstruction of carbon utilization pathways combined with provision of an external carbon source restores energy charge and viability of the rpaA- strain in light/dark cycling conditions. Our observations highlight how a circadian output pathway controls and temporally coordinates essential pathways in carbon metabolism to maximize fitness of cells facing periodic energy limitations. DOI: http://dx.doi.org/10.7554/eLife.23210.001 PMID:28430105

Cross-presentation of IgG-containing immune complexes

PubMed Central

Baker, Kristi; Rath, Timo; Lencer, Wayne I.; Fiebiger, Edda

2012-01-01

IgG is a molecule that functionally combines facets of both innate and adaptive immunity and therefore bridges both arms of the immune system. On the one hand, IgG is created by adaptive immune cells, but can be generated by B cells independently of T cell help. On the other hand, once secreted, IgG can rapidly deliver antigens into intracellular processing pathways, which enable efficient priming of T cell responses towards epitopes from the cognate antigen initially bound by the IgG. While this process has long been known to participate in CD4+ T cell activation, IgG-mediated delivery of exogenous antigens into a major histocompatibility complex (MHC) class I processing pathway has received less attention. The coordinated engagement of IgG with IgG receptors expressed on the cell-surface (FcγR) and within the endolysosomal system (FcRn) is a highly potent means to deliver antigen into processing pathways that promote cross-presentation of MHC class I and presentation of MHC class II-restricted epitopes within the same dendritic cell. This review focuses on the mechanisms by which IgG-containing immune complexes mediate such cross-presentation and the implications that this understanding has for manipulation of immune-mediated diseases that depend upon or are due to the activities of CD8+ T cells. PMID:22847331

Combination of degradation pathways for naphthalene utilization in Rhodococcus sp. strain TFB

PubMed Central

Tomás-Gallardo, Laura; Gómez-Álvarez, Helena; Santero, Eduardo; Floriano, Belén

2014-01-01

Rhodococcus sp. strain TFB is a metabolic versatile bacterium able to grow on naphthalene as the only carbon and energy source. Applying proteomic, genetic and biochemical approaches, we propose in this paper that, at least, three coordinated but independently regulated set of genes are combined to degrade naphthalene in TFB. First, proteins involved in tetralin degradation are also induced by naphthalene and may carry out its conversion to salicylaldehyde. This is the only part of the naphthalene degradation pathway showing glucose catabolite repression. Second, a salicylaldehyde dehydrogenase activity that converts salicylaldehyde to salicylate is detected in naphthalene-grown cells but not in tetralin-or salicylate-grown cells. Finally, we describe the chromosomally located nag genes, encoding the gentisate pathway for salicylate conversion into fumarate and pyruvate, which are only induced by salicylate and not by naphthalene. This work shows how biodegradation pathways in Rhodococcus sp. strain TFB could be assembled using elements from different pathways mainly because of the laxity of the regulatory systems and the broad specificity of the catabolic enzymes. PMID:24325207

Evolution and Design Governing Signal Precision and Amplification in a Bacterial Chemosensory Pathway

PubMed Central

Espinosa, Leon; Baronian, Grégory; Molle, Virginie; Mauriello, Emilia M. F.; Brochier-Armanet, Céline; Mignot, Tâm

2015-01-01

Understanding the principles underlying the plasticity of signal transduction networks is fundamental to decipher the functioning of living cells. In Myxococcus xanthus, a particular chemosensory system (Frz) coordinates the activity of two separate motility systems (the A- and S-motility systems), promoting multicellular development. This unusual structure asks how signal is transduced in a branched signal transduction pathway. Using combined evolution-guided and single cell approaches, we successfully uncoupled the regulations and showed that the A-motility regulation system branched-off an existing signaling system that initially only controlled S-motility. Pathway branching emerged in part following a gene duplication event and changes in the circuit structure increasing the signaling efficiency. In the evolved pathway, the Frz histidine kinase generates a steep biphasic response to increasing external stimulations, which is essential for signal partitioning to the motility systems. We further show that this behavior results from the action of two accessory response regulator proteins that act independently to filter and amplify signals from the upstream kinase. Thus, signal amplification loops may underlie the emergence of new connectivity in signal transduction pathways. PMID:26291327

RhoA/ROCK pathway activity is essential for the correct localization of the germ plasm mRNAs in zebrafish embryos.

PubMed

Miranda-Rodríguez, Jerónimo Roberto; Salas-Vidal, Enrique; Lomelí, Hilda; Zurita, Mario; Schnabel, Denhi

2017-01-01

Zebrafish germ plasm is composed of mRNAs such as vasa and nanos and of proteins such as Bucky ball, all of which localize symmetrically in four aggregates at the distal region of the first two cleavage furrows. The coordination of actin microfilaments, microtubules and kinesin is essential for the correct localization of the germ plasm. Rho-GTPases, through their effectors, coordinate cytoskeletal dynamics. We address the participation of RhoA and its effector ROCK in germ plasm localization during the transition from two- to eight-cell embryos. We found that active RhoA is enriched along the cleavage furrow during the first two division cycles, whereas ROCK localizes at the distal region of the cleavage furrows in a similar pattern as the germ plasm mRNAs. Specific inhibition of RhoA and ROCK affected microtubules organization at the cleavage furrow; these caused the incorrect localization of the germ plasm mRNAs. The incorrect localization of the germ plasm led to a dramatic change in the number of germ cells during the blastula and 24hpf embryo stages without affecting any other developmental processes. We demonstrate that the Rho/ROCK pathway is intimately related to the determination of germ cells in zebrafish embryos. Copyright © 2016 Elsevier Inc. All rights reserved.

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 emerged from a bacterial population by acquiring specific functions that allowed them to outcompete their closest relatives. In conclusion, bacterial species could be defined not only as genomic species but also as ecological species. PMID:27060117

Reconstruction of the genome-scale co-expression network for the Hippo signaling pathway in colorectal cancer.

PubMed

Dehghanian, Fariba; Hojati, Zohreh; Hosseinkhan, Nazanin; Mousavian, Zaynab; Masoudi-Nejad, Ali

2018-05-26

The Hippo signaling pathway (HSP) has been identified as an essential and complex signaling pathway for tumor suppression that coordinates proliferation, differentiation, cell death, cell growth and stemness. In the present study, we conducted a genome-scale co-expression analysis to reconstruct the HSP in colorectal cancer (CRC). Five key modules were detected through network clustering, and a detailed discussion of two modules containing respectively 18 and 13 over and down-regulated members of HSP was provided. Our results suggest new potential regulatory factors in the HSP. The detected modules also suggest novel genes contributing to CRC. Moreover, differential expression analysis confirmed the differential expression pattern of HSP members and new suggested regulatory factors between tumor and normal samples. These findings can further reveal the importance of HSP in CRC. Copyright © 2018 Elsevier Ltd. All rights reserved.

Metabolic pathways for lipid synthesis under nitrogen stress in Chlamydomonas and Nannochloropsis.

PubMed

Banerjee, Avik; Maiti, Subodh K; Guria, Chandan; Banerjee, Chiranjib

2017-01-01

Microalgae are currently being considered as a clean, sustainable and renewable energy source. Enzymes that catalyse the metabolic pathways for biofuel production are specific and require strict regulation and co-ordination. Thorough knowledge of these key enzymes along with their regulatory molecules is essential to enable rational metabolic engineering, to drive the metabolic flux towards the desired metabolites of importance. This paper reviews two key enzymes that play their role in production of bio-oil: DGAT (acyl-CoA:diacylglycerol acyltransferase) and PDAT (phospholipid:diacylglycerol acyltransferase). It also deals with the transcription factors that control the enzymes while cell undergoes a metabolic shift under stress. The paper also discusses the association of other enzymes and pathways that provide substrates and precursors for oil accumulation. Finally a futuristic solution has been proposed about a synthetic algal cell platform that would be committed towards biofuel synthesis.

Phosphoglycerate Mutase 1 Coordinates Glycolysis and Biosynthesis to Promote Tumor Growth

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

Hitosugi, Taro; Zhou, Lu; Elf, Shannon

2012-11-12

It is unclear how cancer cells coordinate glycolysis and biosynthesis to support rapidly growing tumors. We found that the glycolytic enzyme phosphoglycerate mutase 1 (PGAM1), commonly upregulated in human cancers due to loss of TP53, contributes to biosynthesis regulation partially by controlling intracellular levels of its substrate, 3-phosphoglycerate (3-PG), and product, 2-phosphoglycerate (2-PG). 3-PG binds to and inhibits 6-phosphogluconate dehydrogenase in the oxidative pentose phosphate pathway (PPP), while 2-PG activates 3-phosphoglycerate dehydrogenase to provide feedback control of 3-PG levels. Inhibition of PGAM1 by shRNA or a small molecule inhibitor PGMI-004A results in increased 3-PG and decreased 2-PG levels in cancermore » cells, leading to significantly decreased glycolysis, PPP flux and biosynthesis, as well as attenuated cell proliferation and tumor growth.« less

Coordinate Regulation of Stem Cell Competition by Slit-Robo and JAK-STAT Signaling in the Drosophila Testis

PubMed Central

Stine, Rachel R.; Greenspan, Leah J.; Ramachandran, Kapil V.; Matunis, Erika L.

2014-01-01

Stem cells in tissues reside in and receive signals from local microenvironments called niches. Understanding how multiple signals within niches integrate to control stem cell function is challenging. The Drosophila testis stem cell niche consists of somatic hub cells that maintain both germline stem cells and somatic cyst stem cells (CySCs). Here, we show a role for the axon guidance pathway Slit-Roundabout (Robo) in the testis niche. The ligand Slit is expressed specifically in hub cells while its receptor, Roundabout 2 (Robo2), is required in CySCs in order for them to compete for occupancy in the niche. CySCs also require the Slit-Robo effector Abelson tyrosine kinase (Abl) to prevent over-adhesion of CySCs to the niche, and CySCs mutant for Abl outcompete wild type CySCs for niche occupancy. Both Robo2 and Abl phenotypes can be rescued through modulation of adherens junction components, suggesting that the two work together to balance CySC adhesion levels. Interestingly, expression of Robo2 requires JAK-STAT signaling, an important maintenance pathway for both germline and cyst stem cells in the testis. Our work indicates that Slit-Robo signaling affects stem cell function downstream of the JAK-STAT pathway by controlling the ability of stem cells to compete for occupancy in their niche. PMID:25375180

Comparative cell-specific transcriptomics reveals differentiation of C4 photosynthesis pathways in switchgrass and other C4 lineages

PubMed Central

Rao, Xiaolan; Lu, Nan; Li, Guifen; Nakashima, Jin; Tang, Yuhong; Dixon, Richard A.

2016-01-01

Almost all C4 plants require the co-ordination of the adjacent and fully differentiated cell types, mesophyll (M) and bundle sheath (BS). The C4 photosynthetic pathway operates through two distinct subtypes based on how malate is decarboxylated in BS cells; through NAD-malic enzyme (NAD-ME) or NADP-malic enzyme (NADP-ME). The diverse or unique cell-specific molecular features of M and BS cells from separate C4 subtypes of independent lineages remain to be determined. We here provide an M/BS cell type-specific transcriptome data set from the monocot NAD-ME subtype switchgrass (Panicum virgatum). A comparative transcriptomics approach was then applied to compare the M/BS mRNA profiles of switchgrass, monocot NADP-ME subtype C4 plants maize and Setaria viridis, and dicot NAD-ME subtype Cleome gynandra. We evaluated the convergence in the transcript abundance of core components in C4 photosynthesis and transcription factors to establish Kranz anatomy, as well as gene distribution of biological functions, in these four independent C4 lineages. We also estimated the divergence between NAD-ME and NADP-ME subtypes of C4 photosynthesis in the two cell types within C4 species, including differences in genes encoding decarboxylating enzymes, aminotransferases, and metabolite transporters, and differences in the cell-specific functional enrichment of RNA regulation and protein biogenesis/homeostasis. We suggest that C4 plants of independent lineages in both monocots and dicots underwent convergent evolution to establish C4 photosynthesis, while distinct C4 subtypes also underwent divergent processes for the optimization of M and BS cell co-ordination. The comprehensive data sets in our study provide a basis for further research on evolution of C4 species. PMID:26896851

Differential expression of immune-regulatory genes associated with PD-L1 display in melanoma: implications for PD-1 pathway blockade

PubMed Central

Taube, Janis M.; Young, Geoffrey D.; McMiller, Tracee L.; Chen, Shuming; Salas, January T.; Pritchard, Theresa S.; Xu, Haiying; Meeker, Alan K.; Fan, Jinshui; Cheadle, Chris; Berger, Alan E.; Pardoll, Drew M.; Topalian, Suzanne L.

2015-01-01

Purpose Blocking the immunosuppressive PD-1/PD-L1 pathway has anti-tumor activity in multiple cancer types, and PD-L1 expression on tumor cells and infiltrating myeloid cells correlates with the likelihood of response. We previously found that IFNG (interferon-gamma) was over-expressed by TILs in PD-L1+ vs. PD-L1(−) melanomas, creating adaptive immune resistance by promoting PD-L1 display. The current study was undertaken to identify additional factors in the PD-L1+ melanoma microenvironment coordinately contributing to immunosuppression. Experimental design Archived, formalin-fixed paraffin-embedded melanoma specimens were assessed for PD-L1 protein expression at the tumor cell surface with immunohistochemistry (IHC). Whole genome expression analysis, quantitative (q)RT-PCR, immunohistochemistry, and functional in vitro validation studies were employed to assess factors differentially expressed in PD-L1+ versus PD-L1(−) melanomas. Results Functional annotation clustering based on whole genome expression profiling revealed pathways up-regulated in PD-L1+ melanomas, involving immune cell activation, inflammation, and antigen processing and presentation. Analysis by qRT-PCR demonstrated over-expression of functionally related genes in PD-L1+ melanomas, involved in CD8+ T cell activation (CD8A, IFNG, PRF1, CCL5), antigen presentation (CD163, TLR3, CXCL1, LYZ), and immunosuppression [PDCD1 (PD-1), CD274(PD-L1), LAG3, IL10]. Functional studies demonstrated that some factors, including IL-10 and IL-32-gamma, induced PD-L1 expression on monocytes but not tumor cells. Conclusions These studies elucidate the complexity of immune checkpoint regulation in the tumor microenvironment, identifying multiple factors likely contributing to coordinated immunosuppression. These factors may provide tumor escape mechanisms from anti-PD-1/PD-L1 therapy, and should be considered for co-targeting in combinatorial immunomodulation treatment strategies. PMID:25944800

Computational modeling of heterogeneity and function of CD4+ T cells

PubMed Central

Carbo, Adria; Hontecillas, Raquel; Andrew, Tricity; Eden, Kristin; Mei, Yongguo; Hoops, Stefan; Bassaganya-Riera, Josep

2014-01-01

The immune system is composed of many different cell types and hundreds of intersecting molecular pathways and signals. This large biological complexity requires coordination between distinct pro-inflammatory and regulatory cell subsets to respond to infection while maintaining tissue homeostasis. CD4+ T cells play a central role in orchestrating immune responses and in maintaining a balance between pro- and anti- inflammatory responses. This tight balance between regulatory and effector reactions depends on the ability of CD4+ T cells to modulate distinct pathways within large molecular networks, since dysregulated CD4+ T cell responses may result in chronic inflammatory and autoimmune diseases. The CD4+ T cell differentiation process comprises an intricate interplay between cytokines, their receptors, adaptor molecules, signaling cascades and transcription factors that help delineate cell fate and function. Computational modeling can help to describe, simulate, analyze, and predict some of the behaviors in this complicated differentiation network. This review provides a comprehensive overview of existing computational immunology methods as well as novel strategies used to model immune responses with a particular focus on CD4+ T cell differentiation. PMID:25364738

Computational design of a Zn2+ receptor that controls bacterial gene expression

NASA Astrophysics Data System (ADS)

Dwyer, M. A.; Looger, L. L.; Hellinga, H. W.

2003-09-01

The control of cellular physiology and gene expression in response to extracellular signals is a basic property of living systems. We have constructed a synthetic bacterial signal transduction pathway in which gene expression is controlled by extracellular Zn2+. In this system a computationally designed Zn2+-binding periplasmic receptor senses the extracellular solute and triggers a two-component signal transduction pathway via a chimeric transmembrane protein, resulting in transcriptional up-regulation of a -galactosidase reporter gene. The Zn2+-binding site in the designed receptor is based on a four-coordinate, tetrahedral primary coordination sphere consisting of histidines and glutamates. In addition, mutations were introduced in a secondary coordination sphere to satisfy the residual hydrogen-bonding potential of the histidines coordinated to the metal. The importance of the secondary shell interactions is demonstrated by their effect on metal affinity and selectivity, as well as protein stability. Three designed protein sequences, comprising two distinct metal-binding positions, were all shown to bind Zn2+ and to function in the cell-based assay, indicating the generality of the design methodology. These experiments demonstrate that biological systems can be manipulated with computationally designed proteins that have drastically altered ligand-binding specificities, thereby extending the repertoire of genetic control by extracellular signals.

Metabolomic analysis of pancreatic β-cell insulin release in response to glucose.

PubMed

Huang, Mei; Joseph, Jamie W

2012-01-01

Defining the key metabolic pathways that are important for fuel-regulated insulin secretion is critical to providing a complete picture of how nutrients regulate insulin secretion. We have performed a detailed metabolomics study of the clonal β-cell line 832/13 using a gas chromatography-mass spectrometer (GC-MS) to investigate potential coupling factors that link metabolic pathways to insulin secretion. Mid-polar and polar metabolites, extracted from the 832/13 β-cells, were derivatized and then run on a GC/MS to identify and quantify metabolite concentrations. Three hundred fifty-five out of 527 chromatographic peaks could be identified as metabolites by our metabolomic platform. These identified metabolites allowed us to perform a systematic analysis of key pathways involved in glucose-stimulated insulin secretion (GSIS). Of these metabolites, 41 were consistently identified as biomarker for GSIS by orthogonal partial least-squares (OPLS). Most of the identified metabolites are from common metabolic pathways including glycolytic, sorbitol-aldose reductase pathway, pentose phosphate pathway, and the TCA cycle suggesting these pathways play an important role in GSIS. Lipids and related products were also shown to contribute to the clustering of high glucose sample groups. Amino acids lysine, tyrosine, alanine and serine were upregulated by glucose whereas aspartic acid was downregulated by glucose suggesting these amino acids might play a key role in GSIS. In summary, a coordinated signaling cascade elicited by glucose metabolism in pancreatic β-cells is revealed by our metabolomics platform providing a new conceptual framework for future research and/or drug discovery.

Evasion of anti-growth signaling: a key step in tumorigenesis and potential target for treatment and prophylaxis by natural compounds

PubMed Central

Amin, A.R.M. Ruhul; Karpowicz, Phillip A.; Carey, Thomas E.; Arbiser, Jack; Nahta, Rita; Chen, Zhuo G.; Dong, Jin-Tang; Kucuk, Omer; Khan, Gazala N.; Huang, Gloria S.; Mi, Shijun; Lee, Ho-Young; Reichrath, Joerg; Honoki, Kanya; Georgakilas, Alexandros G.; Amedei, Amedeo; Amin, Amr; Helferich, Bill; Boosani, Chandra S.; Ciriolo, Maria Rosa; Chen, Sophie; Mohammed, Sulma I.; Azmi, Asfar S.; Keith, W Nicol; Bhakta, Dipita; Halicka, Dorota; Niccolai, Elena; Fujii, Hiromasa; Aquilano, Katia; Ashraf, S. Salman; Nowsheen, Somaira; Yang, Xujuan; Bilsland, Alan; Shin, Dong M.

2015-01-01

The evasion of anti-growth signaling is an important characteristic of cancer cells. In order to continue to proliferate, cancer cells must somehow uncouple themselves from the many signals that exist to slow down cell growth. Here, we define the anti-growth signaling process, and review several important pathways involved in growth signaling: p53, phosphatase and tensin homolog (PTEN), retinoblastoma protein (Rb), Hippo, growth differentiation factor 15 (GDF15), AT-rich interactive domain 1A (ARID1A), Notch, insulin-like growth factor (IGF), and Krüppel-like factor 5 (KLF5) pathways. Aberrations in these processes in cancer cells involve mutations and thus the suppression of genes that prevent growth, as well as mutation and activation of genes involved in driving cell growth. Using these pathways as examples, we prioritize molecular targets that might be leveraged to promote anti-growth signaling in cancer cells. Interestingly, naturally-occurring phytochemicals found in human diets (either singly or as mixtures) may promote anti-growth signaling, and do so without the potentially adverse effects associated with synthetic chemicals. We review examples of naturally-occurring phytochemicals that may be applied to prevent cancer by antagonizing growth signaling, and propose one phytochemical for each pathway. These are: epigallocatechin-3-gallate (EGCG) for the Rb pathway, luteolin for p53, curcumin for PTEN, porphyrins for Hippo, genistein for GDF15, resveratrol for ARID1A, withaferin A for Notch and diguelin for the IGF1-receptor pathway. The coordination of anti-growth signaling and natural compound studies will provide insight into the future application of these compounds in the clinical setting. PMID:25749195

A mechanistic pan-cancer pathway model informed by multi-omics data interprets stochastic cell fate responses to drugs and mitogens

PubMed Central

Bouhaddou, Mehdi; Koch, Rick J.; DiStefano, Matthew S.; Tan, Annie L.; Mertz, Alex E.

2018-01-01

Most cancer cells harbor multiple drivers whose epistasis and interactions with expression context clouds drug and drug combination sensitivity prediction. We constructed a mechanistic computational model that is context-tailored by omics data to capture regulation of stochastic proliferation and death by pan-cancer driver pathways. Simulations and experiments explore how the coordinated dynamics of RAF/MEK/ERK and PI-3K/AKT kinase activities in response to synergistic mitogen or drug combinations control cell fate in a specific cellular context. In this MCF10A cell context, simulations suggest that synergistic ERK and AKT inhibitor-induced death is likely mediated by BIM rather than BAD, which is supported by prior experimental studies. AKT dynamics explain S-phase entry synergy between EGF and insulin, but simulations suggest that stochastic ERK, and not AKT, dynamics seem to drive cell-to-cell proliferation variability, which in simulations is predictable from pre-stimulus fluctuations in C-Raf/B-Raf levels. Simulations suggest MEK alteration negligibly influences transformation, consistent with clinical data. Tailoring the model to an alternate cell expression and mutation context, a glioma cell line, allows prediction of increased sensitivity of cell death to AKT inhibition. Our model mechanistically interprets context-specific landscapes between driver pathways and cell fates, providing a framework for designing more rational cancer combination therapy. PMID:29579036

Coordinated Proliferation and Differentiation of Human-Induced Pluripotent Stem Cell-Derived Cardiac Progenitor Cells Depend on Bone Morphogenetic Protein Signaling Regulation by GREMLIN 2

PubMed Central

Bylund, Jeffery B.; Trinh, Linh T.; Awgulewitsch, Cassandra P.; Paik, David T.; Jetter, Christopher; Jha, Rajneesh; Zhang, Jianhua; Nolan, Kristof; Xu, Chunhui; Thompson, Thomas B.; Kamp, Timothy J.

2017-01-01

Heart development depends on coordinated proliferation and differentiation of cardiac progenitor cells (CPCs), but how the two processes are synchronized is not well understood. Here, we show that the secreted Bone Morphogenetic Protein (BMP) antagonist GREMLIN 2 (GREM2) is induced in CPCs shortly after cardiac mesoderm specification during differentiation of human pluripotent stem cells. GREM2 expression follows cardiac lineage differentiation independently of the differentiation method used, or the origin of the pluripotent stem cells, suggesting that GREM2 is linked to cardiogenesis. Addition of GREM2 protein strongly increases cardiomyocyte output compared to established procardiogenic differentiation methods. Our data show that inhibition of canonical BMP signaling by GREM2 is necessary to promote proliferation of CPCs. However, canonical BMP signaling inhibition alone is not sufficient to induce cardiac differentiation, which depends on subsequent JNK pathway activation specifically by GREM2. These findings may have broader implications in the design of approaches to orchestrate growth and differentiation of pluripotent stem cell-derived lineages that depend on precise regulation of BMP signaling. PMID:28125926

Coordinated Proliferation and Differentiation of Human-Induced Pluripotent Stem Cell-Derived Cardiac Progenitor Cells Depend on Bone Morphogenetic Protein Signaling Regulation by GREMLIN 2.

PubMed

Bylund, Jeffery B; Trinh, Linh T; Awgulewitsch, Cassandra P; Paik, David T; Jetter, Christopher; Jha, Rajneesh; Zhang, Jianhua; Nolan, Kristof; Xu, Chunhui; Thompson, Thomas B; Kamp, Timothy J; Hatzopoulos, Antonis K

2017-05-01

Heart development depends on coordinated proliferation and differentiation of cardiac progenitor cells (CPCs), but how the two processes are synchronized is not well understood. Here, we show that the secreted Bone Morphogenetic Protein (BMP) antagonist GREMLIN 2 (GREM2) is induced in CPCs shortly after cardiac mesoderm specification during differentiation of human pluripotent stem cells. GREM2 expression follows cardiac lineage differentiation independently of the differentiation method used, or the origin of the pluripotent stem cells, suggesting that GREM2 is linked to cardiogenesis. Addition of GREM2 protein strongly increases cardiomyocyte output compared to established procardiogenic differentiation methods. Our data show that inhibition of canonical BMP signaling by GREM2 is necessary to promote proliferation of CPCs. However, canonical BMP signaling inhibition alone is not sufficient to induce cardiac differentiation, which depends on subsequent JNK pathway activation specifically by GREM2. These findings may have broader implications in the design of approaches to orchestrate growth and differentiation of pluripotent stem cell-derived lineages that depend on precise regulation of BMP signaling.

Coordination of planar cell polarity pathways through Spiny-legs

PubMed Central

Ambegaonkar, Abhijit A; Irvine, Kenneth D

2015-01-01

Morphogenesis and physiology of tissues and organs requires planar cell polarity (PCP) systems that orient and coordinate cells and their behaviors, but the relationship between PCP systems has been controversial. We have characterized how the Frizzled and Dachsous-Fat PCP systems are connected through the Spiny-legs isoform of the Prickle-Spiny-legs locus. Two different components of the Dachsous-Fat system, Dachsous and Dachs, can each independently interact with Spiny-legs and direct its localization in vivo. Through characterization of the contributions of Prickle, Spiny-legs, Dachsous, Fat, and Dachs to PCP in the Drosophila wing, eye, and abdomen, we define where Dachs-Spiny-legs and Dachsous-Spiny-legs interactions contribute to PCP, and provide a new understanding of the orientation of polarity and the basis of PCP phenotypes. Our results support the direct linkage of PCP systems through Sple in specific locales, while emphasizing that cells can be subject to and must ultimately resolve distinct, competing PCP signals. DOI: http://dx.doi.org/10.7554/eLife.09946.001 PMID:26505959

Repulsive guidance molecule B (RGMB) plays negative roles in breast cancer by coordinating BMP signaling.

PubMed

Li, Jin; Ye, Lin; Sanders, Andrew J; Jiang, Wen G

2012-07-01

Repulsive guidance molecules (RGMs) coordinate axon formation and iron homestasis. These molecules are also known as co-receptors of bone morphogenetic proteins (BMPs). However, the role played by RGMs in breast cancer remains unclear. The present study investigated the impact of RGMB on functions of breast cancer cells and corresponding mechanisms. RGMB was knocked down in breast cancer cells by way of an anti-RGMB ribozyme transgene. Knockdown of RGMB resulted in enhanced capacities of proliferation, adhesion, and migration in breast cancer cells. Further investigations demonstrated RGMB knockdown resulted in a reduced expression and activity of Caspase-3, accompanied with better survival in RGMB knockdown cells under serum starvation, which might be induced by its repression on MAPK JNK pathway. Up-regulations of Snai1, Twist, FAK, and Paxillin via enhanced Smad dependent sigaling led to increased capacities of adhesion and migration. Our current data firstly revealed that RGMB may act as a negative regulator in breast cancer through BMP signaling. Copyright © 2012 Wiley Periodicals, Inc.

A Glutaredoxin·BolA Complex Serves as an Iron-Sulfur Cluster Chaperone for the Cytosolic Cluster Assembly Machinery*♦

PubMed Central

Frey, Avery G.; Palenchar, Daniel J.; Wildemann, Justin D.; Philpott, Caroline C.

2016-01-01

Cells contain hundreds of proteins that require iron cofactors for activity. Iron cofactors are synthesized in the cell, but the pathways involved in distributing heme, iron-sulfur clusters, and ferrous/ferric ions to apoproteins remain incompletely defined. In particular, cytosolic monothiol glutaredoxins and BolA-like proteins have been identified as [2Fe-2S]-coordinating complexes in vitro and iron-regulatory proteins in fungi, but it is not clear how these proteins function in mammalian systems or how this complex might affect Fe-S proteins or the cytosolic Fe-S assembly machinery. To explore these questions, we use quantitative immunoprecipitation and live cell proximity-dependent biotinylation to monitor interactions between Glrx3, BolA2, and components of the cytosolic iron-sulfur cluster assembly system. We characterize cytosolic Glrx3·BolA2 as a [2Fe-2S] chaperone complex in human cells. Unlike complexes formed by fungal orthologs, human Glrx3-BolA2 interaction required the coordination of Fe-S clusters, whereas Glrx3 homodimer formation did not. Cellular Glrx3·BolA2 complexes increased 6–8-fold in response to increasing iron, forming a rapidly expandable pool of Fe-S clusters. Fe-S coordination by Glrx3·BolA2 did not depend on Ciapin1 or Ciao1, proteins that bind Glrx3 and are involved in cytosolic Fe-S cluster assembly and distribution. Instead, Glrx3 and BolA2 bound and facilitated Fe-S incorporation into Ciapin1, a [2Fe-2S] protein functioning early in the cytosolic Fe-S assembly pathway. Thus, Glrx3·BolA is a [2Fe-2S] chaperone complex capable of transferring [2Fe-2S] clusters to apoproteins in human cells. PMID:27519415

Rho GTPases Control Polarity, Protrusion, and Adhesion during Cell Movement

PubMed Central

Nobes, Catherine D.; Hall, Alan

1999-01-01

Cell movement is essential during embryogenesis to establish tissue patterns and to drive morphogenetic pathways and in the adult for tissue repair and to direct cells to sites of infection. Animal cells move by crawling and the driving force is derived primarily from the coordinated assembly and disassembly of actin filaments. The small GTPases, Rho, Rac, and Cdc42, regulate the organization of actin filaments and we have analyzed their contributions to the movement of primary embryo fibroblasts in an in vitro wound healing assay. Rac is essential for the protrusion of lamellipodia and for forward movement. Cdc42 is required to maintain cell polarity, which includes the localization of lamellipodial activity to the leading edge and the reorientation of the Golgi apparatus in the direction of movement. Rho is required to maintain cell adhesion during movement, but stress fibers and focal adhesions are not required. Finally, Ras regulates focal adhesion and stress fiber turnover and this is essential for cell movement. We conclude that the signal transduction pathways controlled by the four small GTPases, Rho, Rac, Cdc42, and Ras, cooperate to promote cell movement. PMID:10087266

Vascular remodeling: A redox-modulated mechanism of vessel caliber regulation.

PubMed

Tanaka, Leonardo Y; Laurindo, Francisco R M

2017-08-01

Vascular remodeling, i.e. whole-vessel structural reshaping, determines lumen caliber in (patho)physiology. Here we review mechanisms underlying vessel remodeling, with emphasis in redox regulation. First, we discuss confusing terminology and focus on strictu sensu remodeling. Second, we propose a mechanobiological remodeling paradigm based on the concept of tensional homeostasis as a setpoint regulator. We first focus on shear-mediated models as prototypes of remodeling closely dominated by highly redox-sensitive endothelial function. More detailed discussions focus on mechanosensors, integrins, extracellular matrix, cytoskeleton and inflammatory pathways as potential of mechanisms potentially coupling tensional homeostasis to redox regulation. Further discussion of remodeling associated with atherosclerosis and injury repair highlights important aspects of redox vascular responses. While neointima formation has not shown consistent responsiveness to antioxidants, vessel remodeling has been more clearly responsive, indicating that despite the multilevel redox signaling pathways, there is a coordinated response of the whole vessel. Among mechanisms that may orchestrate redox pathways, we discuss roles of superoxide dismutase activity and extracellular protein disulfide isomerase. We then discuss redox modulation of aneurysms, a special case of expansive remodeling. We propose that the redox modulation of vascular remodeling may reflect (1) remodeling pathophysiology is dominated by a particularly redox-sensitive cell type, e.g., endothelial cells (2) redox pathways are temporospatially coordinated at an organ level across distinct cellular and acellular structures or (3) the tensional homeostasis setpoint is closely connected to redox signaling. The mechanobiological/redox model discussed here can be a basis for improved understanding of remodeling and helps clarifying mechanisms underlying prevalent hard-to-treat diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

Control of B Lymphocyte Development and Functions by the mTOR Signaling Pathways

PubMed Central

Iwata, Terri N.; Ramírez-Komo, Julita A.; Park, Heon; Iritani, Brian M.

2017-01-01

Mechanistic target of rapamycin (mTOR) is a serine/threonine kinase originally discovered as the molecular target of the immunosuppressant rapamycin. mTOR forms two compositionally and functionally distinct complexes, mTORC1 and mTORC2, which are crucial for coordinating nutrient, energy, oxygen, and growth factor availability with cellular growth, proliferation, and survival. Recent studies have identified critical, non-redundant roles for mTORC1 and mTORC2 in controlling B cell development, differentiation, and functions, and have highlighted emerging roles of the Folliculin-Fnip protein complex in regulating mTOR and B cell development. In this review, we summarize the basic mechanisms of mTOR signaling; describe what is known about the roles of mTORC1, mTORC2, and the Folliculin/Fnip1 pathway in B cell development and functions; and briefly outline current clinical approaches for targeting mTOR in B cell neoplasms. We conclude by highlighting a few salient questions and future perspectives regarding mTOR in B lineage cells. PMID:28583723

BAM 1 and RECEPTOR-LIKE PROTEIN KINASE 2 constitute a signaling pathway and modulate CLE peptide-triggered growth inhibition in Arabidopsis root.

PubMed

Shimizu, Noriko; Ishida, Takashi; Yamada, Masashi; Shigenobu, Shuji; Tabata, Ryo; Kinoshita, Atsuko; Yamaguchi, Katsushi; Hasebe, Mitsuyasu; Mitsumasu, Kanako; Sawa, Shinichiro

2015-12-01

Ligand receptor-based signaling is a means of cell-to-cell communication for coordinating developmental and physiological processes in multicellular organisms. In plants, cell-producing meristems utilize this signaling to regulate their activities and ensure for proper development. Shoot and root systems share common requirements for carrying out this process; however, its molecular basis is largely unclear. It has been suggested that synthetic CLV3/EMBRYO SURROUNDING REGION (CLE) peptide shrinks the root meristem through the actions of CLAVATA2 (CLV2) and the RECEPTOR-LIKE PROTEIN KINASE 2 (RPK2) pathway in Arabidopsis thaliana. Our genetic screening for mutations that resist CLE peptide signaling in roots determined that BAM1, which is a member of the leucine-rich repeat receptor-like kinase (LRR-RLK) family, is also involved in this pathway. BAM1 is preferentially expressed in the root tip, including the quiescent center and its surrounding stem cells. Our genetic analysis revealed that BAM1 functions together with RPK2. Using coimmunoprecipitation assay, we showed that BAM1 is capable of forming heteromeric complexes with RPK2. These findings suggest that the BAM1 and RPK2 receptors constitute a signaling pathway that modulates cell proliferation in the root meristem and that related molecules are employed in root and shoot meristems. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.

Endodermal Hedgehog signals modulate Notch pathway activity in the developing digestive tract mesenchyme

PubMed Central

Kim, Tae-Hee; Kim, Byeong-Moo; Mao, Junhao; Rowan, Sheldon; Shivdasani, Ramesh A.

2011-01-01

The digestive tract epithelium and its adjoining mesenchyme undergo coordinated patterning and growth during development. The signals they exchange in the process are not fully characterized but include ligands of the Hedgehog (Hh) family, which originate in the epithelium and are necessary for mesenchymal cells to expand in number and drive elongation of the developing gut tube. The Notch signaling pathway has known requirements in fetal and adult intestinal epithelial progenitors. We detected Notch pathway activity in the embryonic gut mesenchyme and used conditional knockout mice to study its function. Selective disruption of the Notch effector gene RBP-Jκ (Rbpj) in the mesenchyme caused progressive loss of subepithelial fibroblasts and abbreviated gut length, revealing an unexpected requirement in this compartment. Surprisingly, constitutive Notch activity also induced rapid mesenchymal cell loss and impaired organogenesis, probably resulting from increased cell death and suggesting the need for a delicate balance in Notch signaling. Because digestive tract anomalies in mouse embryos with excess Notch activity phenocopy the absence of Hh signaling, we postulated that endodermal Hh restrains mesenchymal Notch pathway activity. Indeed, Hh-deficient embryos showed Notch overactivity in their defective gut mesenchyme and exposure to recombinant sonic hedgehog could override Notch-induced death of cultured fetal gut mesenchymal cells. These results reveal unexpected interactions between prominent signals in gastrointestinal development and provide a coherent explanation for Hh requirements in mesenchymal cell survival and organ growth. PMID:21750033

Six-coordinate manganese(3+) in catalysis by yeast manganese superoxide dismutase

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

Sheng, Yuewei; Gralla, Edith Butler; Schumacher, Mikhail

Reduction of superoxide (O{sub 2}{sup -}) by manganese-containing superoxide dismutase occurs through either a 'prompt protonation' pathway, or an 'inner-sphere' pathway, with the latter leading to formation of an observable Mn-peroxo complex. We recently reported that wild-type (WT) manganese superoxide dismutases (MnSODs) from Saccharomyces cerevisiae and Candida albicans are more gated toward the 'prompt protonation' pathway than human and bacterial MnSODs and suggested that this could result from small structural changes in the second coordination sphere of manganese. We report here that substitution of a second-sphere residue, Tyr34, by phenylalanine (Y34F) causes the MnSOD from S. cerevisiae to react exclusivelymore » through the 'inner-sphere' pathway. At neutral pH, we have a surprising observation that protonation of the Mn-peroxo complex in the mutant yeast enzyme occurs through a fast pathway, leading to a putative six-coordinate Mn3+ species, which actively oxidizes O{sub 2}{sup -} in the catalytic cycle. Upon increasing pH, the fast pathway is gradually replaced by a slow proton-transfer pathway, leading to the well-characterized five-coordinate Mn{sup 3+}. We here propose and compare two hypothetical mechanisms for the mutant yeast enzyme, diffeeing in the structure of the Mn-peroxo complex yet both involving formation of the active six-coordinate Mn{sup 3+} and proton transfer from a second-sphere water molecule, which has substituted for the -OH of Tyr34, to the Mn-peroxo complex. Because WT and the mutant yeast MnSOD both rest in the 2+ state and become six-coordinate when oxidized up from Mn{sup 2+}, six-coordinate Mn{sup 3+} species could also actively function in the mechanism of WT yeast MnSODs.« less

Stop Stalling: Mus81 Required for Efficient Replication | Center for Cancer Research

Cancer.gov

DNA replication is precisely controlled to ensure that daughter cells receive intact, accurate genetic information. Each segment of DNA must be copied only once, and the rate of replication coordinated genome-wide. Mild replication stress slows DNA synthesis and activates a pathway involving the Mus81 endonuclease, which generates a series of DNA breaks that are rapidly

SnoN regulates mammary gland alveologenesis and onset of lactation by promoting prolactin/Stat5 signaling

PubMed Central

Jahchan, Nadine S.; Wang, Douglas; Bissell, Mina J.; Luo, Kunxin

2012-01-01

Mammary epithelial cells undergo structural and functional differentiation at late pregnancy and parturition to produce and secrete milk. Both TGF-β and prolactin pathways are crucial regulators of this process. However, how the activities of these two antagonistic pathways are orchestrated to initiate lactation has not been well defined. Here, we show that SnoN, a negative regulator of TGF-β signaling, coordinates TGF-β and prolactin signaling to control alveologenesis and lactogenesis. SnoN expression is induced at late pregnancy by the coordinated actions of TGF-β and prolactin. The elevated SnoN promotes Stat5 signaling by enhancing its stability, thereby sharply increasing the activity of prolactin signaling at the onset of lactation. SnoN–/– mice display severe defects in alveologenesis and lactogenesis, and mammary epithelial cells from these mice fail to undergo proper morphogenesis. These defects can be rescued by an active Stat5. Thus, our study has identified a new player in the regulation of milk production and revealed a novel function of SnoN in mammary alveologenesis and lactogenesis in vivo through promotion of Stat5 signaling. PMID:22833129

Targeting the mevalonate cascade as a new therapeutic approach in heart disease, cancer and pulmonary disease.

PubMed

Yeganeh, Behzad; Wiechec, Emilia; Ande, Sudharsana R; Sharma, Pawan; Moghadam, Adel Rezaei; Post, Martin; Freed, Darren H; Hashemi, Mohammad; Shojaei, Shahla; Zeki, Amir A; Ghavami, Saeid

2014-07-01

The cholesterol biosynthesis pathway, also known as the mevalonate (MVA) pathway, is an essential cellular pathway that is involved in diverse cell functions. The enzyme 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase (HMGCR) is the rate-limiting step in cholesterol biosynthesis and catalyzes the conversion of HMG-CoA to MVA. Given its role in cholesterol and isoprenoid biosynthesis, the regulation of HMGCR has been intensely investigated. Because all cells require a steady supply of MVA, both the sterol (i.e. cholesterol) and non-sterol (i.e. isoprenoid) products of MVA metabolism exert coordinated feedback regulation on HMGCR through different mechanisms. The proper functioning of HMGCR as the proximal enzyme in the MVA pathway is essential under both normal physiologic conditions and in many diseases given its role in cell cycle pathways and cell proliferation, cholesterol biosynthesis and metabolism, cell cytoskeletal dynamics and stability, cell membrane structure and fluidity, mitochondrial function, proliferation, and cell fate. The blockbuster statin drugs ('statins') directly bind to and inhibit HMGCR, and their use for the past thirty years has revolutionized the treatment of hypercholesterolemia and cardiovascular diseases, in particular coronary heart disease. Initially thought to exert their effects through cholesterol reduction, recent evidence indicates that statins also have pleiotropic immunomodulatory properties independent of cholesterol lowering. In this review we will focus on the therapeutic applications and mechanisms involved in the MVA cascade including Rho GTPase and Rho kinase (ROCK) signaling, statin inhibition of HMGCR, geranylgeranyltransferase (GGTase) inhibition, and farnesyltransferase (FTase) inhibition in cardiovascular disease, pulmonary diseases (e.g. asthma and chronic obstructive pulmonary disease (COPD)), and cancer. Copyright © 2014 Elsevier Inc. All rights reserved.

Enforced hematopoietic cell E- and L-selectin ligand (HCELL) expression primes transendothelial migration of human mesenchymal stem cells.

PubMed

Thankamony, Sai P; Sackstein, Robert

2011-02-08

According to the multistep model of cell migration, chemokine receptor engagement (step 2) triggers conversion of rolling interactions (step 1) into firm adhesion (step 3), yielding transendothelial migration. We recently reported that glycosyltransferase-programmed stereosubstitution (GPS) of CD44 on human mesenchymal stem cells (hMSCs) creates the E-selectin ligand HCELL (hematopoietic cell E-selectin/L-selectin ligand) and, despite absence of CXCR4, systemically administered HCELL(+)hMSCs display robust osteotropism visualized by intravital microscopy. Here we performed studies to define the molecular effectors of this process. We observed that engagement of hMSC HCELL with E-selectin triggers VLA-4 adhesiveness, resulting in shear-resistant adhesion to ligand VCAM-1. This VLA-4 activation is mediated via a Rac1/Rap1 GTPase signaling pathway, resulting in transendothelial migration on stimulated human umbilical vein endothelial cells without chemokine input. These findings indicate that hMSCs coordinately integrate CD44 ligation and integrin activation, circumventing chemokine-mediated signaling, yielding a step 2-bypass pathway of the canonical multistep paradigm of cell migration.

Modes of Action and Functions of ERECTA-family Receptor-like Kinases in Plant Organ Growth and Development

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

TORII, Keiko U.

2012-05-01

Higher plants constitute the central resource for renewable lignocellulose biomass that can supplement for the world's depleting stores of fossil fuels. As such, understanding the molecular and genetic mechanisms of plant organ growth will provide key knowledge and genetic resources that enables manipulation of plant biomass feedstock for better growth and productivity. The goal of this proposal is to understand how cell proliferation and growth are coordinated during aboveground organ morphogenesis, and how cell-cell signaling mediated by a family of receptor kinases coordinates plant organogenesis. The well-established model plant Arabidopsis thaliana is used for our research to facilitate rapid progress.more » Specifically, we focus on how ERECTA-family leucine-rich repeat receptor kinases (LRR-RLKs) interact in a synergistic manner to promote organogenesis and pattern formation in Arabidopsis. This project was highly successful, resulted in fourteen publications including nine peer-reviewed original research articles. One provisional US patent has been filed through this DOE funding. We have addressed the critical roles for a family of receptor kinases in coordinating proliferation and differentiation of plants, and we successfully elucidated the downstream targets of this signaling pathway in specifying stomatal patterning.« less

Mammalian DNA single-strand break repair: an X-ra(y)ted affair.

PubMed

Caldecott, K W

2001-05-01

The genetic stability of living cells is continuously threatened by the presence of endogenous reactive oxygen species and other genotoxic molecules. Of particular threat are the thousands of DNA single-strand breaks that arise in each cell, each day, both directly from disintegration of damaged sugars and indirectly from the excision repair of damaged bases. If un-repaired, single-strand breaks can be converted into double-strand breaks during DNA replication, potentially resulting in chromosomal rearrangement and genetic deletion. Consequently, cells have adopted multiple pathways to ensure the rapid and efficient removal of single-strand breaks. A general feature of these pathways appears to be the extensive employment of protein-protein interactions to stimulate both the individual component steps and the overall repair reaction. Our current understanding of DNA single-strand break repair is discussed, and testable models for the architectural coordination of this important process are presented. Copyright 2001 John Wiley & Sons, Inc.

A Wingless and Notch double-repression mechanism regulates G1–S transition in the Drosophila wing

PubMed Central

Herranz, Héctor; Pérez, Lidia; Martín, Francisco A; Milán, Marco

2008-01-01

The control of tissue growth and patterning is orchestrated in various multicellular tissues by the coordinated activity of the signalling molecules Wnt/Wingless (Wg) and Notch, and mutations in these pathways can cause cancer. The role of these molecules in the control of cell proliferation and the crosstalk between their corresponding pathways remain poorly understood. Crosstalk between Notch and Wg has been proposed to organize pattern and growth in the Drosophila wing primordium. Here we report that Wg and Notch act in a surprisingly linear pathway to control G1–S progression. We present evidence that these molecules exert their function by regulating the expression of the dmyc proto-oncogene and the bantam micro-RNA, which positively modulated the activity of the E2F transcription factor. Our results demonstrate that Notch acts in this cellular context as a repressor of cell-cycle progression and Wg has a permissive role in alleviating Notch-mediated repression of G1–S progression in wing cells. PMID:18451803

Ankrd6 is a mammalian functional homolog of Drosophila planar cell polarity gene diego and regulates coordinated cellular orientation in the mouse inner ear.

PubMed

Jones, Chonnettia; Qian, Dong; Kim, Sun Myoung; Li, Shuangding; Ren, Dongdong; Knapp, Lindsey; Sprinzak, David; Avraham, Karen B; Matsuzaki, Fumio; Chi, Fanglu; Chen, Ping

2014-11-01

The coordinated polarization of neighboring cells within the plane of the tissue, known as planar cell polarity (PCP), is a recurring theme in biology. It is required for numerous developmental processes for the form and function of many tissues and organs across species. The genetic pathway regulating PCP was first discovered in Drosophila, and an analogous but distinct pathway is emerging in vertebrates. It consists of membrane protein complexes known as core PCP proteins that are conserved across species. Here we report that the over-expression of the murine Ankrd6 (mAnkrd6) gene that shares homology with Drosophila core PCP gene diego causes a typical PCP phenotype in Drosophila, and mAnkrd6 can rescue the loss of function of diego in Drosophila. In mice, mAnkrd6 protein is asymmetrically localized in cells of the inner ear sensory organs, characteristic of components of conserved core PCP complexes. The loss of mAnkrd6 causes PCP defects in the inner ear sensory organs. Moreover, canonical Wnt signaling is significantly increased in mouse embryonic fibroblasts from mAnkrd6 knockout mice in comparison to wild type controls. Together, these results indicated that mAnkrd6 is a functional homolog of the Drosophila diego gene for mammalian PCP regulation and act to suppress canonical Wnt signaling. Copyright © 2014 Elsevier Inc. All rights reserved.

Natural and modified promoters for tailored metabolic engineering of the yeast Saccharomyces cerevisiae.

PubMed

Hubmann, Georg; Thevelein, Johan M; Nevoigt, Elke

2014-01-01

The ease of highly sophisticated genetic manipulations in the yeast Saccharomyces cerevisiae has initiated numerous initiatives towards development of metabolically engineered strains for novel applications beyond its traditional use in brewing, baking, and wine making. In fact, baker's yeast has become a key cell factory for the production of various bulk and fine chemicals. Successful metabolic engineering requires fine-tuned adjustments of metabolic fluxes and coordination of multiple pathways within the cell. This has mostly been achieved by controlling gene expression at the transcriptional level, i.e., by using promoters with appropriate strengths and regulatory properties. Here we present an overview of natural and modified promoters, which have been used in metabolic pathway engineering of S. cerevisiae. Recent developments in creating promoters with tailor-made properties are also discussed.

Timing of Tissue-specific Cell Division Requires a Differential Onset of Zygotic Transcription during Metazoan Embryogenesis*

PubMed Central

Wong, Ming-Kin; Guan, Daogang; Ng, Kaoru Hon Chun; Ho, Vincy Wing Sze; An, Xiaomeng; Li, Runsheng; Ren, Xiaoliang

2016-01-01

Metazoan development demands not only precise cell fate differentiation but also accurate timing of cell division to ensure proper development. How cell divisions are temporally coordinated during development is poorly understood. Caenorhabditis elegans embryogenesis provides an excellent opportunity to study this coordination due to its invariant development and widespread division asynchronies. One of the most pronounced asynchronies is a significant delay of cell division in two endoderm progenitor cells, Ea and Ep, hereafter referred to as E2, relative to its cousins that mainly develop into mesoderm organs and tissues. To unravel the genetic control over the endoderm-specific E2 division timing, a total of 822 essential and conserved genes were knocked down using RNAi followed by quantification of cell cycle lengths using in toto imaging of C. elegans embryogenesis and automated lineage. Intriguingly, knockdown of numerous genes encoding the components of general transcription pathway or its regulatory factors leads to a significant reduction in the E2 cell cycle length but an increase in cell cycle length of the remaining cells, indicating a differential requirement of transcription for division timing between the two. Analysis of lineage-specific RNA-seq data demonstrates an earlier onset of transcription in endoderm than in other germ layers, the timing of which coincides with the birth of E2, supporting the notion that the endoderm-specific delay in E2 division timing demands robust zygotic transcription. The reduction in E2 cell cycle length is frequently associated with cell migration defect and gastrulation failure. The results suggest that a tissue-specific transcriptional activation is required to coordinate fate differentiation, division timing, and cell migration to ensure proper development. PMID:27056332

Multivesicular bodies: co-ordinated progression to maturity

PubMed Central

Woodman, Philip G; Futter, Clare E

2008-01-01

Multivesicular endosomes/bodies (MVBs) sort endocytosed proteins to different destinations. Many lysosomally directed membrane proteins are sorted onto intralumenal vesicles, whilst recycling proteins remain on the perimeter membrane from where they are removed via tubular extensions. MVBs move to the cell centre during this maturation process and, when all recycling proteins have been removed, fuse with lysosomes. Recent advances have identified endosomal-sorting complex required for transport (ESCRT)-dependent and ESCRT-independent pathways in intralumenal vesicle formation and mechanisms for sorting recycling cargo into tubules. Cytoskeletal motors, through interactions with these machineries and by regulating MVB movement, help to co-ordinate events leading to a mature, fusion-competent MVB. PMID:18502633

Hippo, TGF-β, and Src-MAPK pathways regulate transcription of the upd3 cytokine in Drosophila enterocytes upon bacterial infection.

PubMed

Houtz, Philip; Bonfini, Alessandro; Liu, Xi; Revah, Jonathan; Guillou, Aurélien; Poidevin, Mickael; Hens, Korneel; Huang, Hsin-Yi; Deplancke, Bart; Tsai, Yu-Chen; Buchon, Nicolas

2017-11-01

Cytokine signaling is responsible for coordinating conserved epithelial regeneration and immune responses in the digestive tract. In the Drosophila midgut, Upd3 is a major cytokine, which is induced in enterocytes (EC) and enteroblasts (EB) upon oral infection, and initiates intestinal stem cell (ISC) dependent tissue repair. To date, the genetic network directing upd3 transcription remains largely uncharacterized. Here, we have identified the key infection-responsive enhancers of the upd3 gene and show that distinct enhancers respond to various stresses. Furthermore, through functional genetic screening, bioinformatic analyses and yeast one-hybrid screening, we determined that the transcription factors Scalloped (Sd), Mothers against dpp (Mad), and D-Fos are principal regulators of upd3 expression. Our study demonstrates that upd3 transcription in the gut is regulated by the activation of multiple pathways, including the Hippo, TGF-β/Dpp, and Src, as well as p38-dependent MAPK pathways. Thus, these essential pathways, which are known to control ISC proliferation cell-autonomously, are also activated in ECs to promote tissue turnover the regulation of upd3 transcription.

Distinct regulation of Snail in two muscle lineages of the ascidian embryo achieves temporal coordination of muscle development.

PubMed

Tokuoka, Miki; Kobayashi, Kenji; Satou, Yutaka

2018-06-06

The transcriptional repressor Snail is required for proper differentiation of the tail muscle of ascidian tadpole larvae. Two muscle lineages (B5.1 and B6.4) contribute to the anterior tail muscle cells, and are consecutively separated from a transcriptionally quiescent germ cell lineage at the 16- and 32-cell stages. Concomitantly, cells of these lineages begin to express Tbx6.b ( Tbx6-r.b ) at the 16- and 32-cell stages, respectively. Meanwhile, Snail expression begins in these two lineages simultaneously at the 32-cell stage. Here, we show that Snail expression is regulated differently between these two lineages. In the B5.1 lineage, Snail was activated through Tbx6.b , which is activated by maternal factors, including Zic-r.a. In the B6.4 lineage, the MAPK pathway was cell-autonomously activated by a constitutively active form of Raf, enabling Zic-r.a to activate Snail independently of Tbx6.b As a result, Snail begins to be expressed at the 32-cell stage simultaneously in these two lineages. Such shortcuts might be required for coordinating developmental programs in embryos in which cells become separated progressively from stem cells, including germline cells. © 2018. Published by The Company of Biologists Ltd.

Stress-mediated translational control in cancer cells.

PubMed

Leprivier, Gabriel; Rotblat, Barak; Khan, Debjit; Jan, Eric; Sorensen, Poul H

2015-07-01

Tumor cells are continually subjected to diverse stress conditions of the tumor microenvironment, including hypoxia, nutrient deprivation, and oxidative or genotoxic stress. Tumor cells must evolve adaptive mechanisms to survive these conditions to ultimately drive tumor progression. Tight control of mRNA translation is critical for this response and the adaptation of tumor cells to such stress forms. This proceeds though a translational reprogramming process which restrains overall translation activity to preserve energy and nutrients, but which also stimulates the selective synthesis of major stress adaptor proteins. Here we present the different regulatory signaling pathways which coordinate mRNA translation in the response to different stress forms, including those regulating eIF2α, mTORC1 and eEF2K, and we explain how tumor cells hijack these pathways for survival under stress. Finally, mechanisms for selective mRNA translation under stress, including the utilization of upstream open reading frames (uORFs) and internal ribosome entry sites (IRESes) are discussed in the context of cell stress. This article is part of a Special Issue entitled: Translation and Cancer. Copyright © 2014 Elsevier B.V. All rights reserved.

PEPCK Coordinates the Regulation of Central Carbon Metabolism to Promote Cancer Cell Growth.

PubMed

Montal, Emily D; Dewi, Ruby; Bhalla, Kavita; Ou, Lihui; Hwang, Bor Jang; Ropell, Ashley E; Gordon, Chris; Liu, Wan-Ju; DeBerardinis, Ralph J; Sudderth, Jessica; Twaddel, William; Boros, Laszlo G; Shroyer, Kenneth R; Duraisamy, Sekhar; Drapkin, Ronny; Powers, R Scott; Rohde, Jason M; Boxer, Matthew B; Wong, Kwok-Kin; Girnun, Geoffrey D

2015-11-19

Phosphoenolpyruvate carboxykinase (PEPCK) is well known for its role in gluconeogenesis. However, PEPCK is also a key regulator of TCA cycle flux. The TCA cycle integrates glucose, amino acid, and lipid metabolism depending on cellular needs. In addition, biosynthetic pathways crucial to tumor growth require the TCA cycle for the processing of glucose and glutamine derived carbons. We show here an unexpected role for PEPCK in promoting cancer cell proliferation in vitro and in vivo by increasing glucose and glutamine utilization toward anabolic metabolism. Unexpectedly, PEPCK also increased the synthesis of ribose from non-carbohydrate sources, such as glutamine, a phenomenon not previously described. Finally, we show that the effects of PEPCK on glucose metabolism and cell proliferation are in part mediated via activation of mTORC1. Taken together, these data demonstrate a role for PEPCK that links metabolic flux and anabolic pathways to cancer cell proliferation. Copyright © 2015 Elsevier Inc. All rights reserved.

Robust Ordering of Anaphase Events by Adaptive Thresholds and Competing Degradation Pathways.

PubMed

Kamenz, Julia; Mihaljev, Tamara; Kubis, Armin; Legewie, Stefan; Hauf, Silke

2015-11-05

The splitting of chromosomes in anaphase and their delivery into the daughter cells needs to be accurately executed to maintain genome stability. Chromosome splitting requires the degradation of securin, whereas the distribution of the chromosomes into the daughter cells requires the degradation of cyclin B. We show that cells encounter and tolerate variations in the abundance of securin or cyclin B. This makes the concurrent onset of securin and cyclin B degradation insufficient to guarantee that early anaphase events occur in the correct order. We uncover that the timing of chromosome splitting is not determined by reaching a fixed securin level, but that this level adapts to the securin degradation kinetics. In conjunction with securin and cyclin B competing for degradation during anaphase, this provides robustness to the temporal order of anaphase events. Our work reveals how parallel cell-cycle pathways can be temporally coordinated despite variability in protein concentrations. Copyright © 2015 Elsevier Inc. All rights reserved.

AKAP-Lbc mobilizes a cardiac hypertrophy signaling pathway.

PubMed

Carnegie, Graeme K; Soughayer, Joseph; Smith, F Donelson; Pedroja, Benjamin S; Zhang, Fang; Diviani, Dario; Bristow, Michael R; Kunkel, Maya T; Newton, Alexandra C; Langeberg, Lorene K; Scott, John D

2008-10-24

Elevated catecholamines in the heart evoke transcriptional activation of the Myocyte Enhancer Factor (MEF) pathway to induce a cellular response known as pathological myocardial hypertrophy. We have discovered that the A-Kinase Anchoring Protein (AKAP)-Lbc is upregulated in hypertrophic cardiomyocytes. It coordinates activation and movement of signaling proteins that initiate MEF2-mediated transcriptional reprogramming events. Live-cell imaging, fluorescent kinase activity reporters, and RNA interference techniques show that AKAP-Lbc couples activation of protein kinase D (PKD) with the phosphorylation-dependent nuclear export of the class II histone deacetylase HDAC5. These studies uncover a role for AKAP-Lbc in which increased expression of the anchoring protein selectively amplifies a signaling pathway that drives cardiac myocytes toward a pathophysiological outcome.

Pathway to a Phenocopy: Heat Stress Effects in Early Embryogenesis

PubMed Central

Crews, Sarah M.; McCleery, W. Tyler; Hutson, M. Shane

2015-01-01

Background Heat shocks applied at the onset of gastrulation in early Drosophila embryos frequently lead to phenocopies of U-shaped mutants – having characteristic failures in the late morphogenetic processes of germband retraction and dorsal closure. The pathway from non-specific heat stress to phenocopied abnormalities is unknown. Results Drosophila embryos subjected to 30-min, 38-°C heat shocks at gastrulation appear to recover and restart morphogenesis. Post-heat-shock development appears normal, albeit slower, until a large fraction of embryos develop amnioserosa holes (diameters > 100 μm). These holes are positively correlated with terminal U-shaped phenocopies. They initiate between amnioserosa cells and open over tens of minutes by evading normal wound healing responses. They are not caused by tissue-wide increases in mechanical stress or decreases in cell-cell adhesion, but instead appear to initiate from isolated apoptosis of amnioserosa cells. Conclusions The pathway from heat shock to U-shaped phenocopies involves the opening of one or more large holes in the amnioserosa that compromise its structural integrity and lead to failures in morphogenetic processes that rely on amnioserosa-generated tensile forces. The proposed mechanism by which heat shock leads to hole initiation and expansion is heterochonicity – i.e., disruption of morphogenetic coordination between embryonic and extra-embryonic cell types. PMID:26498920

The Hippo signaling pathway in liver regeneration and tumorigenesis.

PubMed

Hong, Lixin; Cai, Yabo; Jiang, Mingting; Zhou, Dawang; Chen, Lanfen

2015-01-01

The Hippo signaling pathway is an evolutionarily conserved signaling module that plays critical roles in liver size control and tumorigenesis. The Hippo pathway consists of a core kinase cascade in which the mammalian Ste20-like kinases (Mst1/2, orthologs of Drosophila Hippo) and their cofactor Salvador (Sav1) form a complex to phosphorylate and activate the large tumor suppressor (Lats1/2). Lats1/2 kinases in turn phosphorylate and inhibit the transcription co-activators, the Yes-associated protein (YAP) and the transcriptional co-activator with PDZ-binding motif (TAZ), two major downstream effectors of the Hippo pathway. Losses of the Hippo pathway components induce aberrant hepatomegaly and tumorigenesis, in which YAP coordinates regulation of cell proliferation and apoptosis and plays an essential role. This review summarizes the current findings of the regulation of Hippo signaling in liver regeneration and tumorigenesis, focusing on how the loss of tumor suppressor components of the Hippo pathway results in liver cancers and discussing the molecular mechanisms that regulate the expression and activation of its downstream effector YAP in liver tumorigenesis. © The Author 2014. Published by ABBS Editorial Office in association with Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences.

Epidermal wound repair is regulated by the planar cell polarity signaling pathway.

PubMed

Caddy, Jacinta; Wilanowski, Tomasz; Darido, Charbel; Dworkin, Sebastian; Ting, Stephen B; Zhao, Quan; Rank, Gerhard; Auden, Alana; Srivastava, Seema; Papenfuss, Tony A; Murdoch, Jennifer N; Humbert, Patrick O; Parekh, Vishwas; Boulos, Nidal; Weber, Thomas; Zuo, Jian; Cunningham, John M; Jane, Stephen M

2010-07-20

The mammalian PCP pathway regulates diverse developmental processes requiring coordinated cellular movement, including neural tube closure and cochlear stereociliary orientation. Here, we show that epidermal wound repair is regulated by PCP signaling. Mice carrying mutant alleles of PCP genes Vangl2, Celsr1, PTK7, and Scrb1, and the transcription factor Grhl3, interact genetically, exhibiting failed wound healing, neural tube defects, and disordered cochlear polarity. Using phylogenetic analysis, ChIP, and gene expression in Grhl3(-)(/-) mice, we identified RhoGEF19, a homolog of a RhoA activator involved in PCP signaling in Xenopus, as a direct target of GRHL3. Knockdown of Grhl3 or RhoGEF19 in keratinocytes induced defects in actin polymerization, cellular polarity, and wound healing, and re-expression of RhoGEF19 rescued these defects in Grhl3-kd cells. These results define a role for Grhl3 in PCP signaling and broadly implicate this pathway in epidermal repair. (c) 2010 Elsevier Inc. All rights reserved.

Epidermal wound repair is regulated by the planar cell polarity signaling pathway

PubMed Central

Caddy, Jacinta; Wilanowski, Tomasz; Darido, Charbel; Dworkin, Sebastian; Ting, Stephen B.; Zhao, Quan; Rank, Gerhard; Auden, Alana; Srivastava, Seema; Papenfuss, Tony A.; Murdoch, Jennifer N.; Humbert, Patrick O.; Boulos, Nidal; Weber, Thomas; Zuo, Jian; Cunningham, John M.; Jane, Stephen M.

2010-01-01

SUMMARY The mammalian PCP pathway regulates diverse developmental processes requiring coordinated cellular movement, including neural tube closure and cochlear stereociliary orientation. Here, we show that epidermal wound repair is regulated by PCP signaling. Mice carrying mutant alleles of PCP genes Vangl2, Celsr1, PTK7, and Scrb1, and the transcription factor Grhl3, interact genetically, exhibiting failed wound healing, neural tube defects and disordered cochlear polarity. Using phylogenetic analysis, ChIP, and gene expression in Grhl3−/− mice, we identified RhoGEF19, a homologue of a RhoA activator involved in PCP signaling in Xenopus, as a direct target of GRHL3. Knockdown of Grhl3 or RhoGEF19 in keratinocytes induced defects in actin polymerisation, cellular polarity and wound healing, and re-expression of RhoGEF19 rescued these defects in Grhl3-kd cells. These results define a role for Grhl3 in PCP signaling, and broadly implicate this pathway in epidermal repair. PMID:20643356

A PHGDH inhibitor reveals coordination of serine synthesis and 1-carbon unit fate

PubMed Central

Pacold, Michael E.; Brimacombe, Kyle R.; Chan, Sze Ham; Rohde, Jason M.; Lewis, Caroline A.; Swier, Lotteke J.Y.M.; Possemato, Richard; Chen, Walter W.; Sullivan, Lucas B.; Fiske, Brian P.; Cho, Sung Won; Freinkman, Elizaveta; Birsoy, Kıvanç; Abu-Remaileh, Monther; Shaul, Yoav D.; Liu, Chieh Min; Zhou, Minerva; Koh, Min Jung; Chung, Haeyoon; Davidson, Shawn M.; Luengo, Alba; Wang, Amy Q.; Xu, Xin; Yasgar, Adam; Liu, Li; Rai, Ganesha; Westover, Kenneth D.; Vander Heiden, Matthew G.; Shen, Min; Gray, Nathanael S.; Boxer, Matthew B.; Sabatini, David M.

2016-01-01

Serine is a both a proteinogenic amino acid and the source of one-carbon units essential for de novo purine and deoxythymidine synthesis. In the canonical glucose-derived serine synthesis pathway, Homo sapiens phosphoglycerate dehydrogenase (PHGDH) catalyzes the first, rate-limiting step. Genetic loss of PHGDH is toxic towards PHGDH-overexpressing breast cancer cell lines even in the presence of exogenous serine. Here, we use a quantitative high-throughput screen to identify small molecule PHGDH inhibitors. These compounds reduce the production of glucose-derived serine in cells and suppress the growth of PHGDH-dependent cancer cells in culture and in orthotopic xenograft tumors. Surprisingly, PHGDH inhibition reduced the incorporation into nucleotides of one-carbon units from glucose-derived and exogenous serine. We conclude that glycolytic serine synthesis coordinates the use of one-carbon units from endogenous and exogenous serine in nucleotide synthesis, and suggest that one-carbon unit wasting may contribute to the efficacy of PHGDH inhibitors in vitro and in vivo. PMID:27110680

Phosphorylation of the Streptococcus pneumoniae cell wall biosynthesis enzyme MurC by a eukaryotic-like Ser/Thr kinase.

PubMed

Falk, Shaun P; Weisblum, Bernard

2013-03-01

Streptococcus pneumoniae contains a single Ser/Thr kinase-phosphatase pair known as StkP-PhpP. Here, we report the interaction of StkP-PhpP with S. pneumoniae UDP-N-acetylmuramoyl:L-alanine ligase, MurC, an enzyme that synthesizes an essential intermediate of the cell wall peptidoglycan pathway. Combinatorial phage display using StkP as target selected the peptide sequence YEVCGSDTVGC as an interacting partner and subsequently confirmed by ELISA. The phage peptide sequence YEVCGSDTVGC aligns closely with the MurC motif spanning S. pneumoniae amino acid coordinates 31-37. We show that MurC is phosphorylated by StkP and that phosphoMurC is dephosphorylated by PhpP. These data suggest a link between StkP-PhpP with the coordinated regulation of cell wall biosynthesis via MurC. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Biogenesis of the multifunctional lipid droplet: Lipids, proteins, and sites

PubMed Central

Gross, Steven P.

2014-01-01

Lipid droplets (LDs) are ubiquitous dynamic organelles that store and supply lipids in all eukaryotic and some prokaryotic cells for energy metabolism, membrane synthesis, and production of essential lipid-derived molecules. Interest in the organelle’s cell biology has exponentially increased over the last decade due to the link between LDs and prevalent human diseases and the discovery of new and unexpected functions of LDs. As a result, there has been significant recent progress toward understanding where and how LDs are formed, and the specific lipid pathways that coordinate LD biogenesis. PMID:24590170

The Hippo pathway in disease and therapy: cancer and beyond

PubMed Central

2014-01-01

The Hippo tumour suppressor pathway co-ordinates cell proliferation, cell death and cell differentiation to regulate tissue growth control. In mammals, a conserved core Hippo signalling module receives signal inputs on different levels to ensure the proper regulation of YAP/TAZ activities as transcriptional co-activators. While the core module members MST1/2, Salvador, LATS1/2 and MOB1 have been attributed tumour suppressive functions, YAP/TAZ have been mainly described to have oncogenic roles, although some reports provided evidence supporting growth suppressive roles of YAP/TAZ in certain cancer settings. Intriguingly, mammalian Hippo signalling is also implicated in non-cancer diseases and plays a role in tissue regeneration following injury. Cumulatively, these findings indicate that the pharmacological inhibition or activation of the Hippo pathway could be desirable depending on the disease context. In this review, we first summarise the functions of the mammalian Hippo pathway in tumour formation, and then discuss non-cancer diseases involving Hippo signalling core components with a specific focus on our current understanding of the non-cancer roles of MST1/2 and YAP/TAZ. In addition, the pros and cons of possible pharmacological interventions with Hippo signalling will be reviewed, with particular emphasis on anti-cancer drug development and regenerative medicine. PMID:25097725

Signaling network of dendritic cells in response to pathogens: a community-input supported knowledgebase.

PubMed

Patil, Sonali; Pincas, Hanna; Seto, Jeremy; Nudelman, German; Nudelman, Irina; Sealfon, Stuart C

2010-10-07

Dendritic cells are antigen-presenting cells that play an essential role in linking the innate and adaptive immune systems. Much research has focused on the signaling pathways triggered upon infection of dendritic cells by various pathogens. The high level of activity in the field makes it desirable to have a pathway-based resource to access the information in the literature. Current pathway diagrams lack either comprehensiveness, or an open-access editorial interface. Hence, there is a need for a dependable, expertly curated knowledgebase that integrates this information into a map of signaling networks. We have built a detailed diagram of the dendritic cell signaling network, with the goal of providing researchers with a valuable resource and a facile method for community input. Network construction has relied on comprehensive review of the literature and regular updates. The diagram includes detailed depictions of pathways activated downstream of different pathogen recognition receptors such as Toll-like receptors, retinoic acid-inducible gene-I-like receptors, C-type lectin receptors and nucleotide-binding oligomerization domain-like receptors. Initially assembled using CellDesigner software, it provides an annotated graphical representation of interactions stored in Systems Biology Mark-up Language. The network, which comprises 249 nodes and 213 edges, has been web-published through the Biological Pathway Publisher software suite. Nodes are annotated with PubMed references and gene-related information, and linked to a public wiki, providing a discussion forum for updates and corrections. To gain more insight into regulatory patterns of dendritic cell signaling, we analyzed the network using graph-theory methods: bifan, feedforward and multi-input convergence motifs were enriched. This emphasis on activating control mechanisms is consonant with a network that subserves persistent and coordinated responses to pathogen detection. This map represents a navigable aid for presenting a consensus view of the current knowledge on dendritic cell signaling that can be continuously improved through contributions of research community experts. Because the map is available in a machine readable format, it can be edited and may assist researchers in data analysis. Furthermore, the availability of a comprehensive knowledgebase might help further research in this area such as vaccine development. The dendritic cell signaling knowledgebase is accessible at http://tsb.mssm.edu/pathwayPublisher/DC_pathway/DC_pathway_index.html.

Care pathways across the primary-hospital care continuum: using the multi-level framework in explaining care coordination

PubMed Central

2013-01-01

Background Care pathways are widely used in hospitals for a structured and detailed planning of the care process. There is a growing interest in extending care pathways into primary care to improve quality of care by increasing care coordination. Evidence is sparse about the relationship between care pathways and care coordination. The multi-level framework explores care coordination across organizations and states that (inter)organizational mechanisms have an effect on the relationships between healthcare professionals, resulting in quality and efficiency of care. The aim of this study was to assess the extent to which care pathways support or create elements of the multi-level framework necessary to improve care coordination across the primary - hospital care continuum. Methods This study is an in-depth analysis of five existing local community projects located in four different regions in Flanders (Belgium) to determine whether the available empirical evidence supported or refuted the theoretical expectations from the multi-level framework. Data were gathered using mixed methods, including structured face-to-face interviews, participant observations, documentation and a focus group. Multiple cases were analyzed performing a cross case synthesis to strengthen the results. Results The development of a care pathway across the primary-hospital care continuum, supported by a step-by-step scenario, led to the use of existing and newly constructed structures, data monitoring and the development of information tools. The construction and use of these inter-organizational mechanisms had a positive effect on exchanging information, formulating and sharing goals, defining and knowing each other’s roles, expectations and competences and building qualitative relationships. Conclusion Care pathways across the primary-hospital care continuum enhance the components of care coordination. PMID:23919518

Care pathways across the primary-hospital care continuum: using the multi-level framework in explaining care coordination.

PubMed

Van Houdt, Sabine; Heyrman, Jan; Vanhaecht, Kris; Sermeus, Walter; De Lepeleire, Jan

2013-08-06

Care pathways are widely used in hospitals for a structured and detailed planning of the care process. There is a growing interest in extending care pathways into primary care to improve quality of care by increasing care coordination. Evidence is sparse about the relationship between care pathways and care coordination.The multi-level framework explores care coordination across organizations and states that (inter)organizational mechanisms have an effect on the relationships between healthcare professionals, resulting in quality and efficiency of care.The aim of this study was to assess the extent to which care pathways support or create elements of the multi-level framework necessary to improve care coordination across the primary-hospital care continuum. This study is an in-depth analysis of five existing local community projects located in four different regions in Flanders (Belgium) to determine whether the available empirical evidence supported or refuted the theoretical expectations from the multi-level framework. Data were gathered using mixed methods, including structured face-to-face interviews, participant observations, documentation and a focus group. Multiple cases were analyzed performing a cross case synthesis to strengthen the results. The development of a care pathway across the primary-hospital care continuum, supported by a step-by-step scenario, led to the use of existing and newly constructed structures, data monitoring and the development of information tools. The construction and use of these inter-organizational mechanisms had a positive effect on exchanging information, formulating and sharing goals, defining and knowing each other's roles, expectations and competences and building qualitative relationships. Care pathways across the primary-hospital care continuum enhance the components of care coordination.

Protein Design for Pathway Engineering

PubMed Central

Eriksen, Dawn T.; Lian, Jiazhang; Zhao, Huimin

2013-01-01

Design and construction of biochemical pathways has increased the complexity of biosynthetically-produced compounds when compared to single enzyme biocatalysis. However, the coordination of multiple enzymes can introduce a complicated set of obstacles to overcome in order to achieve a high titer and yield of the desired compound. Metabolic engineering has made great strides in developing tools to optimize the flux through a target pathway, but the inherent characteristics of a particular enzyme within the pathway can still limit the productivity. Thus, judicious protein design is critical for metabolic and pathway engineering. This review will describe various strategies and examples of applying protein design to pathway engineering to optimize the flux through the pathway. The proteins can be engineered for altered substrate specificity/selectivity, increased catalytic activity, reduced mass transfer limitations through specific protein localization, and reduced substrate/product inhibition. Protein engineering can also be expanded to design biosensors to enable high through-put screening and to customize cell signaling networks. These strategies have successfully engineered pathways for significantly increased productivity of the desired product or in the production of novel compounds. PMID:23558037

Protein design for pathway engineering.

PubMed

Eriksen, Dawn T; Lian, Jiazhang; Zhao, Huimin

2014-02-01

Design and construction of biochemical pathways has increased the complexity of biosynthetically-produced compounds when compared to single enzyme biocatalysis. However, the coordination of multiple enzymes can introduce a complicated set of obstacles to overcome in order to achieve a high titer and yield of the desired compound. Metabolic engineering has made great strides in developing tools to optimize the flux through a target pathway, but the inherent characteristics of a particular enzyme within the pathway can still limit the productivity. Thus, judicious protein design is critical for metabolic and pathway engineering. This review will describe various strategies and examples of applying protein design to pathway engineering to optimize the flux through the pathway. The proteins can be engineered for altered substrate specificity/selectivity, increased catalytic activity, reduced mass transfer limitations through specific protein localization, and reduced substrate/product inhibition. Protein engineering can also be expanded to design biosensors to enable high through-put screening and to customize cell signaling networks. These strategies have successfully engineered pathways for significantly increased productivity of the desired product or in the production of novel compounds. Copyright © 2013 Elsevier Inc. All rights reserved.

Signaling mechanisms regulating adult neural stem cells and neurogenesis

PubMed Central

Faigle, Roland; Song, Hongjun

2012-01-01

Background Adult neurogenesis occurs throughout life in discrete regions of the mammalian brain and is tightly regulated via both extrinsic environmental influences and intrinsic genetic factors. In recent years, several crucial signaling pathways have been identified in regulating self-renewal, proliferation, and differentiation of neural stem cells, as well as migration and functional integration of developing neurons in the adult brain. Scope of review Here we review our current understanding of signaling mechanisms, including Wnt, notch, sonic hedgehog, growth and neurotrophic factors, bone morphogenetic proteins, neurotransmitters, transcription factors, and epigenetic modulators, and crosstalk between these signaling pathways in the regulation of adult neurogenesis. We also highlight emerging principles in the vastly growing field of adult neural stem cell biology and neural plasticity. Major conclusions Recent methodological advances have enabled the field to identify signaling mechanisms that fine-tune and coordinate neurogenesis in the adult brain, leading to a better characterization of both cell-intrinsic and environmental cues defining the neurogenic niche. Significant questions related to niche cell identity and underlying regulatory mechanisms remain to be fully addressed and will be the focus of future studies. General significance A full understanding of the role and function of individual signaling pathways in regulating neural stem cells and generation and integration of newborn neurons in the adult brain may lead to targeted new therapies for neurological diseases in humans. PMID:22982587

A novel protein kinase D phosphorylation site in the tumor suppressor Rab interactor 1 is critical for coordination of cell migration.

PubMed

Ziegler, Susanne; Eiseler, Tim; Scholz, Rolf-Peter; Beck, Alexander; Link, Gisela; Hausser, Angelika

2011-03-01

The multifunctional signal adapter protein Ras and Rab interactor 1 (RIN1) is a Ras effector protein involved in the regulation of epithelial cell processes such as cell migration and endocytosis. RIN1 signals via two downstream pathways, namely the activation of Rab5 and Abl family kinases. Protein kinase D (PKD) phosphorylates RIN1 at serine 351 in vitro, thereby regulating interaction with 14-3-3 proteins. Here, we report the identification of serine 292 in RIN1 as an in vivo PKD phosphorylation site. PKD-mediated phosphorylation at this site was confirmed with a phospho-specific antibody and by mass spectrometry. We demonstrate that phosphorylation at serine 292 controls RIN1-mediated inhibition of cell migration by modulating the activation of Abl kinases. We further provide evidence that RIN1 in vivo phosphorylation at serine 351 occurs independently of PKD. Collectively, our data identify a novel PKD signaling pathway through RIN1 and Abl kinases that is involved in the regulation of actin remodeling and cell migration.

De-novo NAD+ synthesis regulates SIRT1-FOXO1 apoptotic pathway in response to NQO1 substrates in lung cancer cells.

PubMed

Liu, Huiying; Xing, Rong; Cheng, Xuefang; Li, Qingran; Liu, Fang; Ye, Hui; Zhao, Min; Wang, Hong; Wang, Guangji; Hao, Haiping

2016-09-20

Tryptophan metabolism is essential in diverse kinds of tumors via regulating tumor immunology. However, the direct role of tryptophan metabolism and its signaling pathway in cancer cells remain largely elusive. Here, we establish a mechanistic link from L-type amino acid transporter 1 (LAT1) mediated transport of tryptophan and the subsequent de-novo NAD+ synthesis to SIRT1-FOXO1 regulated apoptotic signaling in A549 cells in response to NQO1 activation. In response to NQO1 activation, SIRT1 is repressed leading to the increased cellular accumulation of acetylated FOXO1 that transcriptionally activates apoptotic signaling. Decreased uptake of tryptophan due to the downregulation of LAT1 coordinates with PARP-1 hyperactivation to induce rapid depletion of NAD+ pool. Particularly, the LAT1-NAD+-SIRT1 signaling is activated in tumor tissues of patients with non-small cell lung cancer. Because NQO1 activation is characterized with oxidative challenge induced DNA damage, these results suggest that LAT1 and de-novo NAD+ synthesis in NSCLC cells may play essential roles in sensing excessive oxidative stress.

The developmental origin of brain tumours: a cellular and molecular framework.

PubMed

Azzarelli, Roberta; Simons, Benjamin D; Philpott, Anna

2018-05-14

The development of the nervous system relies on the coordinated regulation of stem cell self-renewal and differentiation. The discovery that brain tumours contain a subpopulation of cells with stem/progenitor characteristics that are capable of sustaining tumour growth has emphasized the importance of understanding the cellular dynamics and the molecular pathways regulating neural stem cell behaviour. By focusing on recent work on glioma and medulloblastoma, we review how lineage tracing contributed to dissecting the embryonic origin of brain tumours and how lineage-specific mechanisms that regulate stem cell behaviour in the embryo may be subverted in cancer to achieve uncontrolled proliferation and suppression of differentiation. © 2018. Published by The Company of Biologists Ltd.

Remote Control of Intestinal Stem Cell Activity by Haemocytes in Drosophila

PubMed Central

Chakrabarti, Sveta; Li, Xiaoxue; Collas, Esther Jeanne; Boquete, Jean-Phillipe; Lemaitre, Bruno

2016-01-01

The JAK/STAT pathway is a key signaling pathway in the regulation of development and immunity in metazoans. In contrast to the multiple combinatorial JAK/STAT pathways in mammals, only one canonical JAK/STAT pathway exists in Drosophila. It is activated by three secreted proteins of the Unpaired family (Upd): Upd1, Upd2 and Upd3. Although many studies have established a link between JAK/STAT activation and tissue damage, the mode of activation and the precise function of this pathway in the Drosophila systemic immune response remain unclear. In this study, we used mutations in upd2 and upd3 to investigate the role of the JAK/STAT pathway in the systemic immune response. Our study shows that haemocytes express the three upd genes and that injury markedly induces the expression of upd3 by the JNK pathway in haemocytes, which in turn activates the JAK/STAT pathway in the fat body and the gut. Surprisingly, release of Upd3 from haemocytes upon injury can remotely stimulate stem cell proliferation and the expression of Drosomycin-like genes in the intestine. Our results also suggest that a certain level of intestinal epithelium renewal is required for optimal survival to septic injury. While haemocyte-derived Upd promotes intestinal stem cell activation and survival upon septic injury, haemocytes are dispensable for epithelium renewal upon oral bacterial infection. Our study also indicates that intestinal epithelium renewal is sensitive to insults from both the lumen and the haemocoel. It also reveals that release of Upds by haemocytes coordinates the wound-healing program in multiple tissues, including the gut, an organ whose integrity is critical to fly survival. PMID:27231872

Remote Control of Intestinal Stem Cell Activity by Haemocytes in Drosophila.

PubMed

Chakrabarti, Sveta; Dudzic, Jan Paul; Li, Xiaoxue; Collas, Esther Jeanne; Boquete, Jean-Phillipe; Lemaitre, Bruno

2016-05-01

The JAK/STAT pathway is a key signaling pathway in the regulation of development and immunity in metazoans. In contrast to the multiple combinatorial JAK/STAT pathways in mammals, only one canonical JAK/STAT pathway exists in Drosophila. It is activated by three secreted proteins of the Unpaired family (Upd): Upd1, Upd2 and Upd3. Although many studies have established a link between JAK/STAT activation and tissue damage, the mode of activation and the precise function of this pathway in the Drosophila systemic immune response remain unclear. In this study, we used mutations in upd2 and upd3 to investigate the role of the JAK/STAT pathway in the systemic immune response. Our study shows that haemocytes express the three upd genes and that injury markedly induces the expression of upd3 by the JNK pathway in haemocytes, which in turn activates the JAK/STAT pathway in the fat body and the gut. Surprisingly, release of Upd3 from haemocytes upon injury can remotely stimulate stem cell proliferation and the expression of Drosomycin-like genes in the intestine. Our results also suggest that a certain level of intestinal epithelium renewal is required for optimal survival to septic injury. While haemocyte-derived Upd promotes intestinal stem cell activation and survival upon septic injury, haemocytes are dispensable for epithelium renewal upon oral bacterial infection. Our study also indicates that intestinal epithelium renewal is sensitive to insults from both the lumen and the haemocoel. It also reveals that release of Upds by haemocytes coordinates the wound-healing program in multiple tissues, including the gut, an organ whose integrity is critical to fly survival.

Junction restructuring and spermatogenesis: the biology, regulation, and implication in male contraceptive development.

PubMed

Yan, Helen H N; Mruk, Dolores D; Cheng, C Yan

2008-01-01

Spermatogenesis that occurs in the seminiferous epithelium of adult mammalian testes is associated with extensive junction restructuring at the Sertoli-Sertoli cell, Sertoli-germ cell, and Sertoli-basement membrane interface. While this morphological phenomenon is known and has been described in great details for decades, the biochemical and molecular changes as well as the mechanisms/signaling pathways that define changes at the cell-cell and cell-matrix interface remain largely unknown until recently. In this chapter, we summarize and discuss findings in the field regarding the coordinated efforts of the anchoring [e.g., adherens junction (AJ), such as basal ectoplasmic specialization (basal ES)] and tight junctions (TJs) that are present in the same microenvironment, such as at the blood-testis barrier (BTB), or at distinctly opposite ends of the Sertoli cell epithelium, such as between apical ectoplasmic specialization (apical ES) in the apical compartment, and the BTB adjacent to the basal compartment of the epithelium. These efforts, in turn, regulate and coordinate different cellular events that occur during the seminiferous epithelial cycle. For instance, the events of spermiation and of preleptotene spermatocyte migration across the BTB both take place concurrently at stage VIII of the epithelial cycle of spermatogenesis. Recent findings suggest that these events are coordinated by protein complexes found at the apical and basal ES and TJ, which are located at different ends of the Sertoli cell epithelium. Besides, we highlight important areas of research that can now be undertaken, and functional studies that can be designed to tackle different issues pertinent to junction restructuring during spermatogenesis.

Phosphoinositide 3-Kinase Regulates Glycolysis through Mobilization of Aldolase from the Actin Cytoskeleton.

PubMed

Hu, Hai; Juvekar, Ashish; Lyssiotis, Costas A; Lien, Evan C; Albeck, John G; Oh, Doogie; Varma, Gopal; Hung, Yin Pun; Ullas, Soumya; Lauring, Josh; Seth, Pankaj; Lundquist, Mark R; Tolan, Dean R; Grant, Aaron K; Needleman, Daniel J; Asara, John M; Cantley, Lewis C; Wulf, Gerburg M

2016-01-28

The phosphoinositide 3-kinase (PI3K) pathway regulates multiple steps in glucose metabolism and also cytoskeletal functions, such as cell movement and attachment. Here, we show that PI3K directly coordinates glycolysis with cytoskeletal dynamics in an AKT-independent manner. Growth factors or insulin stimulate the PI3K-dependent activation of Rac, leading to disruption of the actin cytoskeleton, release of filamentous actin-bound aldolase A, and an increase in aldolase activity. Consistently, PI3K inhibitors, but not AKT, SGK, or mTOR inhibitors, cause a significant decrease in glycolysis at the step catalyzed by aldolase, while activating PIK3CA mutations have the opposite effect. These results point toward a master regulatory function of PI3K that integrates an epithelial cell's metabolism and its form, shape, and function, coordinating glycolysis with the energy-intensive dynamics of actin remodeling. Copyright © 2016 Elsevier Inc. All rights reserved.

Dissecting the chromatin interactome of microRNA genes.

PubMed

Chen, Dijun; Fu, Liang-Yu; Zhang, Zhao; Li, Guoliang; Zhang, Hang; Jiang, Li; Harrison, Andrew P; Shanahan, Hugh P; Klukas, Christian; Zhang, Hong-Yu; Ruan, Yijun; Chen, Ling-Ling; Chen, Ming

2014-03-01

Our knowledge of the role of higher-order chromatin structures in transcription of microRNA genes (MIRs) is evolving rapidly. Here we investigate the effect of 3D architecture of chromatin on the transcriptional regulation of MIRs. We demonstrate that MIRs have transcriptional features that are similar to protein-coding genes. RNA polymerase II-associated ChIA-PET data reveal that many groups of MIRs and protein-coding genes are organized into functionally compartmentalized chromatin communities and undergo coordinated expression when their genomic loci are spatially colocated. We observe that MIRs display widespread communication in those transcriptionally active communities. Moreover, miRNA-target interactions are significantly enriched among communities with functional homogeneity while depleted from the same community from which they originated, suggesting MIRs coordinating function-related pathways at posttranscriptional level. Further investigation demonstrates the existence of spatial MIR-MIR chromatin interacting networks. We show that groups of spatially coordinated MIRs are frequently from the same family and involved in the same disease category. The spatial interaction network possesses both common and cell-specific subnetwork modules that result from the spatial organization of chromatin within different cell types. Together, our study unveils an entirely unexplored layer of MIR regulation throughout the human genome that links the spatial coordination of MIRs to their co-expression and function.

Tuning of Nafion® by HKUST-1 as coordination network to enhance proton conductivity for fuel cell applications

NASA Astrophysics Data System (ADS)

Kim, Hee Jin; Talukdar, Krishan; Choi, Sang-June

2016-02-01

Metal-organic frameworks can be intentionally coordinated to achieve improved proton conductivity because they have highly ordered structures and modular nature that serve as a scaffold to anchor acidic groups and develop efficient proton transfer pathways for fuel cell application. Using the concept of a coordination network, the conductivity of Nafion® was tuned by the incorporation of HKUST-1. It has CuII-paddle wheel type nodes and 1,3,5-benzenetricarboxylate struts, feature accessible sites that provides an improved protonic channel depending on the water content. In spite of the fact that HKUST-1 is neutral, coordinated water molecules are contributed adequately acidic by CuII to supply protons to enhance proton conductivity. Water molecules play a vital part in transfer of proton as conducting media and serve as triggers to change proton conductivity through reforming hydrogen bonding networks by water adsorption/desorption process. Increased ion exchange capacity and proton conductivity with lower water uptake of the H3PO4-doped material, and improved thermal stability (as confirmed by thermogravimetric analysis) were achieved. The structure of HKUST-1 was confirmed via field emission scanning electron microscopy and X-ray diffraction, while the porosity and adsorption desorption capacity were characterized by porosity analysis.

Towards high resolution analysis of metabolic flux in cells and tissues.

PubMed

Sims, James K; Manteiga, Sara; Lee, Kyongbum

2013-10-01

Metabolism extracts chemical energy from nutrients, uses this energy to form building blocks for biosynthesis, and interconverts between various small molecules that coordinate the activities of cellular pathways. The metabolic state of a cell is increasingly recognized to determine the phenotype of not only metabolically active cell types such as liver, muscle, and adipose, but also other specialized cell types such as neurons and immune cells. This review focuses on methods to quantify intracellular reaction flux as a measure of cellular metabolic activity, with emphasis on studies involving cells of mammalian tissue. Two key areas are highlighted for future development, single cell metabolomics and noninvasive imaging, which could enable spatiotemporally resolved analysis and thereby overcome issues of heterogeneity, a distinctive feature of tissue metabolism. Copyright © 2013 Elsevier Ltd. All rights reserved.

Towards High Resolution Analysis of Metabolic Flux in Cells and Tissues

PubMed Central

Sims, James K; Manteiga, Sara; Lee, Kyongbum

2013-01-01

Metabolism extracts chemical energy from nutrients, uses this energy to form building blocks for biosynthesis, and interconverts between various small molecules that coordinate the activities of cellular pathways. The metabolic state of a cell is increasingly recognized to determine the phenotype of not only metabolically active cell types such as liver, muscle, and adipose, but also other specialized cell types such as neurons and immune cells. This review focuses on methods to quantify intracellular reaction flux as a measure of cellular metabolic activity, with emphasis on studies involving cells of mammalian tissue. Two key areas are highlighted for future development, single cell metabolomics and noninvasive imaging, which could enable spatiotemporally resolved analysis and thereby overcome issues of heterogeneity, a distinctive feature of tissue metabolism. PMID:23906926

Phospholipase D and the Maintenance of Phosphatidic Acid Levels for Regulation of Mammalian Target of Rapamycin (mTOR)*

PubMed Central

Foster, David A.; Salloum, Darin; Menon, Deepak; Frias, Maria A.

2014-01-01

Phosphatidic acid (PA) is a critical metabolite at the heart of membrane phospholipid biosynthesis. However, PA also serves as a critical lipid second messenger that regulates several proteins implicated in the control of cell cycle progression and cell growth. Three major metabolic pathways generate PA: phospholipase D (PLD), diacylglycerol kinase (DGK), and lysophosphatidic acid acyltransferase (LPAAT). The LPAAT pathway is integral to de novo membrane phospholipid biosynthesis, whereas the PLD and DGK pathways are activated in response to growth factors and stress. The PLD pathway is also responsive to nutrients. A key target for the lipid second messenger function of PA is mTOR, the mammalian/mechanistic target of rapamycin, which integrates both nutrient and growth factor signals to control cell growth and proliferation. Although PLD has been widely implicated in the generation of PA needed for mTOR activation, it is becoming clear that PA generated via the LPAAT and DGK pathways is also involved in the regulation of mTOR. In this minireview, we highlight the coordinated maintenance of intracellular PA levels that regulate mTOR signals stimulated by growth factors and nutrients, including amino acids, lipids, glucose, and Gln. Emerging evidence indicates compensatory increases in one source of PA when another source is compromised, highlighting the importance of being able to adapt to stressful conditions that interfere with PA production. The regulation of PA levels has important implications for cancer cells that depend on PA and mTOR activity for survival. PMID:24990952

Phospholipase D and the maintenance of phosphatidic acid levels for regulation of mammalian target of rapamycin (mTOR).

PubMed

Foster, David A; Salloum, Darin; Menon, Deepak; Frias, Maria A

2014-08-15

Phosphatidic acid (PA) is a critical metabolite at the heart of membrane phospholipid biosynthesis. However, PA also serves as a critical lipid second messenger that regulates several proteins implicated in the control of cell cycle progression and cell growth. Three major metabolic pathways generate PA: phospholipase D (PLD), diacylglycerol kinase (DGK), and lysophosphatidic acid acyltransferase (LPAAT). The LPAAT pathway is integral to de novo membrane phospholipid biosynthesis, whereas the PLD and DGK pathways are activated in response to growth factors and stress. The PLD pathway is also responsive to nutrients. A key target for the lipid second messenger function of PA is mTOR, the mammalian/mechanistic target of rapamycin, which integrates both nutrient and growth factor signals to control cell growth and proliferation. Although PLD has been widely implicated in the generation of PA needed for mTOR activation, it is becoming clear that PA generated via the LPAAT and DGK pathways is also involved in the regulation of mTOR. In this minireview, we highlight the coordinated maintenance of intracellular PA levels that regulate mTOR signals stimulated by growth factors and nutrients, including amino acids, lipids, glucose, and Gln. Emerging evidence indicates compensatory increases in one source of PA when another source is compromised, highlighting the importance of being able to adapt to stressful conditions that interfere with PA production. The regulation of PA levels has important implications for cancer cells that depend on PA and mTOR activity for survival. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Co-stimulatory function in primary germinal center responses: CD40 and B7 are required on distinct antigen-presenting cells.

PubMed

Watanabe, Masashi; Fujihara, Chiharu; Radtke, Andrea J; Chiang, Y Jeffrey; Bhatia, Sumeena; Germain, Ronald N; Hodes, Richard J

2017-09-04

T cell-dependent germinal center (GC) responses require coordinated interactions of T cells with two antigen-presenting cell (APC) populations, B cells and dendritic cells (DCs), in the presence of B7- and CD40-dependent co-stimulatory pathways. Contrary to the prevailing paradigm, we found unique cellular requirements for B7 and CD40 expression in primary GC responses to vaccine immunization with protein antigen and adjuvant: B7 was required on DCs but was not required on B cells, whereas CD40 was required on B cells but not on DCs in the generation of antigen-specific follicular helper T cells, antigen-specific GC B cells, and high-affinity class-switched antibody production. There was, in fact, no requirement for coexpression of B7 and CD40 on the same cell in these responses. Our findings support a substantially revised model for co-stimulatory function in the primary GC response, with crucial and distinct contributions of B7- and CD40-dependent pathways expressed by different APC populations and with important implications for understanding how to optimize vaccine responses or limit autoimmunity. This is a work of the U.S. Government and is not subject to copyright protection in the United States. Foreign copyrights may apply.

Antennally mediated negative feedback regulation of pheromone production in the pine engraver beetle, Ips pini

NASA Astrophysics Data System (ADS)

Ginzel, Matthew D.; Bearfield, Jeremy C.; Keeling, Christopher I.; McCormack, Colin C.; Blomquist, Gary J.; Tittiger, Claus

2007-01-01

Bark beetles use monoterpenoid aggregation pheromones to coordinate host colonization and mating. These chemical signals are produced de novo in midgut cells via the mevalonate pathway, and pheromone production may be regulated by a negative feedback system mediated through the antennae. In this study, we explored the effect of antennectomy on pheromone production and transcript levels of key mevalonate pathway genes in juvenile hormone III-treated male pine engraver beetles, Ips pini (Say). Antennectomized males produced significantly greater amounts of pheromone than podectomized males and those with intact antennae. Likewise, mRNA levels of three mevalonate pathway genes important in pheromone biosynthesis were measured by quantitative real-time PCR and found to be induced to a greater extent with antennectomy, suggesting a transcriptional regulation of pheromone production.

Unique Proton Transportation Pathway in a Robust Inorganic Coordination Polymer Leading to Intrinsically High and Sustainable Anhydrous Proton Conductivity.

PubMed

Gui, Daxiang; Dai, Xing; Tao, Zetian; Zheng, Tao; Wang, Xiangxiang; Silver, Mark A; Shu, Jie; Chen, Lanhua; Wang, Yanlong; Zhang, Tiantian; Xie, Jian; Zou, Lin; Xia, Yuanhua; Zhang, Jujia; Zhang, Jin; Zhao, Ling; Diwu, Juan; Zhou, Ruhong; Chai, Zhifang; Wang, Shuao

2018-05-16

Although comprehensive progress has been made in the area of coordination polymer (CP)/metal-organic framework (MOF)-based proton-conducting materials over the past decade, searching for a CP/MOF with stable, intrinsic, high anhydrous proton conductivity that can be directly used as a practical electrolyte in an intermediate-temperature proton-exchange membrane fuel cell assembly for durable power generation remains a substantial challenge. Here, we introduce a new proton-conducting CP, (NH 4 ) 3 [Zr(H 2/3 PO 4 ) 3 ] (ZrP), which consists of one-dimensional zirconium phosphate anionic chains and fully ordered charge-balancing NH 4 + cations. X-ray crystallography, neutron powder diffraction, and variable-temperature solid-state NMR spectroscopy suggest that protons are disordered within an inherent hydrogen-bonded infinite chain of acid-base pairs (N-H···O-P), leading to a stable anhydrous proton conductivity of 1.45 × 10 -3 S·cm -1 at 180 °C, one of the highest values among reported intermediate-temperature proton-conducting materials. First-principles and quantum molecular dynamics simulations were used to directly visualize the unique proton transport pathway involving very efficient proton exchange between NH 4 + and phosphate pairs, which is distinct from the common guest encapsulation/dehydration/superprotonic transition mechanisms. ZrP as the electrolyte was further assembled into a H 2 /O 2 fuel cell, which showed a record-high electrical power density of 12 mW·cm -2 at 180 °C among reported cells assembled from crystalline solid electrolytes, as well as a direct methanol fuel cell for the first time to demonstrate real applications. These cells were tested for over 15 h without notable power loss.

Reciprocal feedback regulation of PI3K and androgen receptor signaling in PTEN-deficient prostate cancer

PubMed Central

Carver, Brett S; Chapinski, Caren; Wongvipat, John; Hieronymus, Haley; Chen, Yu; Chandarlapaty, Sarat; Arora, Vivek K; Le, Carl; Koutcher, Jason; Scher, Howard; Scardino, Peter T; Rosen, Neal; Sawyers, Charles L

2011-01-01

Summary Prostate cancer is characterized by its dependence on androgen receptor and frequent activation of PI3K signaling. We find that AR transcriptional output is decreased in human and murine tumors with PTEN deletion and that PI3K pathway inhibition activates AR signaling by relieving feedback inhibition of HER kinases. Similarly, AR inhibition activates AKT signaling by reducing levels of the AKT phosphatase PHLPP. Thus, these two oncogenic pathways cross-regulate each other by reciprocal feedback. Inhibition of one activates the other, thereby maintaining tumor cell survival. However, combined pharmacologic inhibition of PI3K and AR signaling caused near complete prostate cancer regressions in a Pten-deficient murine prostate cancer model and in human prostate cancer xenografts, indicating that both pathways coordinately support survival. Significance The two most frequently activated signaling pathways in prostate cancer are driven by AR and PI3K. Inhibitors of the PI3K pathway are in early clinical trials and AR inhibitors confer clinical responses in most patients. However, these inhibitors rarely induce tumor regression in preclinical models. Here we show that these pathways regulate each other by reciprocal negative feedback, such that inhibition of one activates the other. Therefore, tumor cells can adapt and survive when either single pathway is inhibited pharmacologically. Our demonstration of profound tumor regressions with combined pathway inhibition in preclinical prostate tumor models provides rationale for combination therapy in patients. PMID:21575859

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

PubMed

Wang, Ming; Yang, Kezhen; Le, Jie

2015-03-01

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

mTORC1 as the main gateway to autophagy

PubMed Central

Rabanal-Ruiz, Yoana; Otten, Elsje G.; Korolchuk, Viktor I.

2017-01-01

Cells and organisms must coordinate their metabolic activity with changes in their environment to ensure their growth only when conditions are favourable. In order to maintain cellular homoeostasis, a tight regulation between the synthesis and degradation of cellular components is essential. At the epicentre of the cellular nutrient sensing is the mechanistic target of rapamycin complex 1 (mTORC1) which connects environmental cues, including nutrient and growth factor availability as well as stress, to metabolic processes in order to preserve cellular homoeostasis. Under nutrient-rich conditions mTORC1 promotes cell growth by stimulating biosynthetic pathways, including synthesis of proteins, lipids and nucleotides, and by inhibiting cellular catabolism through repression of the autophagic pathway. Its close signalling interplay with the energy sensor AMP-activated protein kinase (AMPK) dictates whether the cell actively favours anabolic or catabolic processes. Underlining the role of mTORC1 in the coordination of cellular metabolism, its deregulation is linked to numerous human diseases ranging from metabolic disorders to many cancers. Although mTORC1 can be modulated by a number of different inputs, amino acids represent primordial cues that cannot be compensated for by any other stimuli. The understanding of how amino acids signal to mTORC1 has increased considerably in the last years; however this area of research remains a hot topic in biomedical sciences. The current ideas and models proposed to explain the interrelationship between amino acid sensing, mTORC1 signalling and autophagy is the subject of the present review. PMID:29233869

Comparative cell cycle transcriptomics reveals synchronization of developmental transcription factor networks in cancer cells

PubMed Central

Johard, Helena; Mahdessian, Diana; Fedr, Radek; Marks, Carolyn; Medalová, Jiřina; Souček, Karel; Lundberg, Emma; Linnarsson, Sten; Bryja, Vítězslav; Sekyrova, Petra; Altun, Mikael; Andäng, Michael

2017-01-01

The cell cycle coordinates core functions such as replication and cell division. However, cell-cycle-regulated transcription in the control of non-core functions, such as cell identity maintenance through specific transcription factors (TFs) and signalling pathways remains unclear. Here, we provide a resource consisting of mapped transcriptomes in unsynchronized HeLa and U2OS cancer cells sorted for cell cycle phase by Fucci reporter expression. We developed a novel algorithm for data analysis that enables efficient visualization and data comparisons and identified cell cycle synchronization of Notch signalling and TFs associated with development. Furthermore, the cell cycle synchronizes with the circadian clock, providing a possible link between developmental transcriptional networks and the cell cycle. In conclusion we find that cell cycle synchronized transcriptional patterns are temporally compartmentalized and more complex than previously anticipated, involving genes, which control cell identity and development. PMID:29228002

mTORC1 and CK2 coordinate ternary and eIF4F complex assembly

PubMed Central

Gandin, Valentina; Masvidal, Laia; Cargnello, Marie; Gyenis, Laszlo; McLaughlan, Shannon; Cai, Yutian; Tenkerian, Clara; Morita, Masahiro; Balanathan, Preetika; Jean-Jean, Olivier; Stambolic, Vuk; Trost, Matthias; Furic, Luc; Larose, Louise; Koromilas, Antonis E.; Asano, Katsura; Litchfield, David; Larsson, Ola; Topisirovic, Ivan

2016-01-01

Ternary complex (TC) and eIF4F complex assembly are the two major rate-limiting steps in translation initiation regulated by eIF2α phosphorylation and the mTOR/4E-BP pathway, respectively. How TC and eIF4F assembly are coordinated, however, remains largely unknown. We show that mTOR suppresses translation of mRNAs activated under short-term stress wherein TC recycling is attenuated by eIF2α phosphorylation. During acute nutrient or growth factor stimulation, mTORC1 induces eIF2β phosphorylation and recruitment of NCK1 to eIF2, decreases eIF2α phosphorylation and bolsters TC recycling. Accordingly, eIF2β mediates the effect of mTORC1 on protein synthesis and proliferation. In addition, we demonstrate a formerly undocumented role for CK2 in regulation of translation initiation, whereby CK2 stimulates phosphorylation of eIF2β and simultaneously bolsters eIF4F complex assembly via the mTORC1/4E-BP pathway. These findings imply a previously unrecognized mode of translation regulation, whereby mTORC1 and CK2 coordinate TC and eIF4F complex assembly to stimulate cell proliferation. PMID:27040916

Divergence of macrophage phagocytic and antimicrobial programs in leprosy.

PubMed

Montoya, Dennis; Cruz, Daniel; Teles, Rosane M B; Lee, Delphine J; Ochoa, Maria Teresa; Krutzik, Stephan R; Chun, Rene; Schenk, Mirjam; Zhang, Xiaoran; Ferguson, Benjamin G; Burdick, Anne E; Sarno, Euzenir N; Rea, Thomas H; Hewison, Martin; Adams, John S; Cheng, Genhong; Modlin, Robert L

2009-10-22

Effective innate immunity against many microbial pathogens requires macrophage programs that upregulate phagocytosis and direct antimicrobial pathways, two functions generally assumed to be coordinately regulated. We investigated the regulation of these key functions in human blood-derived macrophages. Interleukin-10 (IL-10) induced the phagocytic pathway, including the C-type lectin CD209 and scavenger receptors, resulting in phagocytosis of mycobacteria and oxidized low-density lipoprotein. IL-15 induced the vitamin D-dependent antimicrobial pathway and CD209, yet the cells were less phagocytic. The differential regulation of macrophage functional programs was confirmed by analysis of leprosy lesions: the macrophage phagocytosis pathway was prominent in the clinically progressive, multibacillary form of the disease, whereas the vitamin D-dependent antimicrobial pathway predominated in the self-limited form and in patients undergoing reversal reactions from the multibacillary to the self-limited form. These data indicate that macrophage programs for phagocytosis and antimicrobial responses are distinct and differentially regulated in innate immunity to bacterial infections.

Transcriptional Profiling of Hypoxic Neural Stem Cells Identifies Calcineurin-NFATc4 Signaling as a Major Regulator of Neural Stem Cell Biology

PubMed Central

Moreno, Marta; Fernández, Virginia; Monllau, Josep M.; Borrell, Víctor; Lerin, Carles; de la Iglesia, Núria

2015-01-01

Summary Neural stem cells (NSCs) reside in a hypoxic microenvironment within the brain. However, the crucial transcription factors (TFs) that regulate NSC biology under physiologic hypoxia are poorly understood. Here we have performed gene set enrichment analysis (GSEA) of microarray datasets from hypoxic versus normoxic NSCs with the aim of identifying pathways and TFs that are activated under oxygen concentrations mimicking normal brain tissue microenvironment. Integration of TF target (TFT) and pathway enrichment analysis identified the calcium-regulated TF NFATc4 as a major candidate to regulate hypoxic NSC functions. Nfatc4 expression was coordinately upregulated by top hypoxia-activated TFs, while NFATc4 target genes were enriched in hypoxic NSCs. Loss-of-function analyses further revealed that the calcineurin-NFATc4 signaling axis acts as a major regulator of NSC self-renewal and proliferation in vitro and in vivo by promoting the expression of TFs, including Id2, that contribute to the maintenance of the NSC state. PMID:26235896

Positioning of centrioles is a conserved readout of Frizzled planar cell polarity signalling

PubMed Central

Carvajal-Gonzalez, Jose Maria; Roman, Angel-Carlos; Mlodzik, Marek

2016-01-01

Planar cell polarity (PCP) signalling is a well-conserved developmental pathway regulating cellular orientation during development. An evolutionarily conserved pathway readout is not established and, moreover, it is thought that PCP mediated cellular responses are tissue-specific. A key PCP function in vertebrates is to regulate coordinated centriole/cilia positioning, a function that has not been associated with PCP in Drosophila. Here we report instructive input of Frizzled-PCP (Fz/PCP) signalling into polarized centriole positioning in Drosophila wings. We show that centrioles are polarized in pupal wing cells as a readout of PCP signalling, with both gain and loss-of-function Fz/PCP signalling affecting centriole polarization. Importantly, loss or gain of centrioles does not affect Fz/PCP establishment, implicating centriolar positioning as a conserved PCP-readout, likely downstream of PCP-regulated actin polymerization. Together with vertebrate data, these results suggest a unifying model of centriole/cilia positioning as a common downstream effect of PCP signalling from flies to mammals. PMID:27021213

Positioning of centrioles is a conserved readout of Frizzled planar cell polarity signalling.

PubMed

Carvajal-Gonzalez, Jose Maria; Roman, Angel-Carlos; Mlodzik, Marek

2016-03-29

Planar cell polarity (PCP) signalling is a well-conserved developmental pathway regulating cellular orientation during development. An evolutionarily conserved pathway readout is not established and, moreover, it is thought that PCP mediated cellular responses are tissue-specific. A key PCP function in vertebrates is to regulate coordinated centriole/cilia positioning, a function that has not been associated with PCP in Drosophila. Here we report instructive input of Frizzled-PCP (Fz/PCP) signalling into polarized centriole positioning in Drosophila wings. We show that centrioles are polarized in pupal wing cells as a readout of PCP signalling, with both gain and loss-of-function Fz/PCP signalling affecting centriole polarization. Importantly, loss or gain of centrioles does not affect Fz/PCP establishment, implicating centriolar positioning as a conserved PCP-readout, likely downstream of PCP-regulated actin polymerization. Together with vertebrate data, these results suggest a unifying model of centriole/cilia positioning as a common downstream effect of PCP signalling from flies to mammals.

A flexible ontology for inference of emergent whole cell function from relationships between subcellular processes.

PubMed

Hansen, Jens; Meretzky, David; Woldesenbet, Simeneh; Stolovitzky, Gustavo; Iyengar, Ravi

2017-12-18

Whole cell responses arise from coordinated interactions between diverse human gene products functioning within various pathways underlying sub-cellular processes (SCP). Lower level SCPs interact to form higher level SCPs, often in a context specific manner to give rise to whole cell function. We sought to determine if capturing such relationships enables us to describe the emergence of whole cell functions from interacting SCPs. We developed the Molecular Biology of the Cell Ontology based on standard cell biology and biochemistry textbooks and review articles. Currently, our ontology contains 5,384 genes, 753 SCPs and 19,180 expertly curated gene-SCP associations. Our algorithm to populate the SCPs with genes enables extension of the ontology on demand and the adaption of the ontology to the continuously growing cell biological knowledge. Since whole cell responses most often arise from the coordinated activity of multiple SCPs, we developed a dynamic enrichment algorithm that flexibly predicts SCP-SCP relationships beyond the current taxonomy. This algorithm enables us to identify interactions between SCPs as a basis for higher order function in a context dependent manner, allowing us to provide a detailed description of how SCPs together can give rise to whole cell functions. We conclude that this ontology can, from omics data sets, enable the development of detailed SCP networks for predictive modeling of emergent whole cell functions.

Coordinated regulation by two VPS9 domain-containing guanine nucleotide exchange factors in small GTPase Rab5 signaling pathways in fission yeast

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

Tsukamoto, Yuta; Kagiwada, Satoshi; Shimazu, Sayuri

The small GTPase Rab5 is reported to regulate various cellular functions, such as vesicular transport and endocytosis. VPS9 domain-containing proteins are thought to activate Rab5(s) by their guanine-nucleotide exchange activities. Numerous VPS9 proteins have been identified and are structurally conserved from yeast to mammalian cells. However, the functional relationships among VPS9 proteins in cells remain unclear. Only one Rab5 and two VPS9 proteins were identified in the Schizosaccharomyces pombe genome. Here, we examined the cellular function of two VPS9 proteins and the relationship between these proteins in cellular functions. Vps901-GFP and Vps902-GFP exhibited dotted signals in vegetative and differentiated cells.more » vps901 deletion mutant (Δvps901) cells exhibited a phenotype deficient in the mating process and responses to high concentrations of ions, such as calcium and metals, and Δvps901Δvps902 double mutant cells exhibited round cell shapes similar to ypt5-909 (Rab5 mutant allele) cells. Deletion of both vps901 and vps902 genes completely abolished the mating process and responses to various stresses. A lack of vacuole formation and aberrant inner cell membrane structures were also observed in Δvps901Δvps902 cells by electron microscopy. These data strongly suggest that Vps901 and Vps902 are cooperatively involved in the regulation of cellular functions, such as cell morphology, sexual development, response to ion stresses, and vacuole formation, via Rab5 signaling pathways in fission yeast cells. - Highlights: • Roles of Rab5 activator VPS9 proteins in cellular functions. • Cooperation between VPS9 proteins in Rab5 signaling pathway. • Roles of each VPS9 protein in Rab5 signaling pathway are discussed.« less

Single-cell transcriptomes identify human islet cell signatures and reveal cell-type–specific expression changes in type 2 diabetes

PubMed Central

Bolisetty, Mohan; Kursawe, Romy; Sun, Lili; Sivakamasundari, V.; Kycia, Ina

2017-01-01

Blood glucose levels are tightly controlled by the coordinated action of at least four cell types constituting pancreatic islets. Changes in the proportion and/or function of these cells are associated with genetic and molecular pathophysiology of monogenic, type 1, and type 2 (T2D) diabetes. Cellular heterogeneity impedes precise understanding of the molecular components of each islet cell type that govern islet (dys)function, particularly the less abundant delta and gamma/pancreatic polypeptide (PP) cells. Here, we report single-cell transcriptomes for 638 cells from nondiabetic (ND) and T2D human islet samples. Analyses of ND single-cell transcriptomes identified distinct alpha, beta, delta, and PP/gamma cell-type signatures. Genes linked to rare and common forms of islet dysfunction and diabetes were expressed in the delta and PP/gamma cell types. Moreover, this study revealed that delta cells specifically express receptors that receive and coordinate systemic cues from the leptin, ghrelin, and dopamine signaling pathways implicating them as integrators of central and peripheral metabolic signals into the pancreatic islet. Finally, single-cell transcriptome profiling revealed genes differentially regulated between T2D and ND alpha, beta, and delta cells that were undetectable in paired whole islet analyses. This study thus identifies fundamental cell-type–specific features of pancreatic islet (dys)function and provides a critical resource for comprehensive understanding of islet biology and diabetes pathogenesis. PMID:27864352

The non-canonical BMP and Wnt/β-catenin signaling pathways orchestrate early tooth development

PubMed Central

Yuan, Guohua; Yang, Guobin; Zheng, Yuqian; Zhu, Xiaojing; Chen, Zhi; Zhang, Zunyi; Chen, YiPing

2015-01-01

BMP and Wnt signaling pathways play a crucial role in organogenesis, including tooth development. Despite extensive studies, the exact functions, as well as if and how these two pathways act coordinately in regulating early tooth development, remain elusive. In this study, we dissected regulatory functions of BMP and Wnt pathways in early tooth development using a transgenic noggin (Nog) overexpression model (K14Cre;pNog). It exhibits early arrested tooth development, accompanied by reduced cell proliferation and loss of odontogenic fate marker Pitx2 expression in the dental epithelium. We demonstrated that overexpression of Nog disrupted BMP non-canonical activity, which led to a dramatic reduction of cell proliferation rate but did not affect Pitx2 expression. We further identified a novel function of Nog by inhibiting Wnt/β-catenin signaling, causing loss of Pitx2 expression. Co-immunoprecipitation and TOPflash assays revealed direct binding of Nog to Wnts to functionally prevent Wnt/β-catenin signaling. In situ PLA and immunohistochemistry on Nog mutants confirmed in vivo interaction between endogenous Nog and Wnts and modulation of Wnt signaling by Nog in tooth germs. Genetic rescue experiments presented evidence that both BMP and Wnt signaling pathways contribute to cell proliferation regulation in the dental epithelium, with Wnt signaling also controlling the odontogenic fate. Reactivation of both BMP and Wnt signaling pathways, but not of only one of them, rescued tooth developmental defects in K14Cre;pNog mice, in which Wnt signaling can be substituted by transgenic activation of Pitx2. Our results reveal the orchestration of non-canonical BMP and Wnt/β-catenin signaling pathways in the regulation of early tooth development. PMID:25428587

ROP GTPase-mediated auxin signaling regulates pavement cell interdigitation in Arabidopsis thaliana.

PubMed

Lin, Deshu; Ren, Huibo; Fu, Ying

2015-01-01

In multicellular plant organs, cell shape formation depends on molecular switches to transduce developmental or environmental signals and to coordinate cell-to-cell communication. Plants have a specific subfamily of the Rho GTPase family, usually called Rho of Plants (ROP), which serve as a critical signal transducer involved in many cellular processes. In the last decade, important advances in the ROP-mediated regulation of plant cell morphogenesis have been made by using Arabidopsis thaliana leaf and cotyledon pavement cells. Especially, the auxin-ROP signaling networks have been demonstrated to control interdigitated growth of pavement cells to form jigsaw-puzzle shapes. Here, we review findings related to the discovery of this novel auxin-signaling mechanism at the cell surface. This signaling pathway is to a large extent independent of the well-known Transport Inhibitor Response (TIR)-Auxin Signaling F-Box (AFB) pathway, and instead requires Auxin Binding Protein 1 (ABP1) interaction with the plasma membrane-localized, transmembrane kinase (TMK) receptor-like kinase to regulate ROP proteins. Once activated, ROP influences cytoskeletal organization and inhibits endocytosis of the auxin transporter PIN1. The present review focuses on ROP signaling and its self-organizing feature allowing ROP proteins to serve as a bustling signal decoder and integrator for plant cell morphogenesis. © 2014 Institute of Botany, Chinese Academy of Sciences.

Riding the Waves: How Our Cells Send Signals | Center for Cancer Research

Cancer.gov

The ability of cells to perceive and respond to their environment is critical in order to maintain basic cellular functions such as development, tissue repair, and response to stress. This process happens through a complex system of communication, called cell signaling, which governs basic cellular activities and coordinates cell actions. Errors in cell signaling have been linked to numerous diseases, including cancer. NF-κB is a protein complex that plays a critical role in many cell signaling pathways by controlling gene activation. It is widely used by cells to regulate cell growth and survival and helps to protect the cell from conditions that would otherwise cause it to die. Many tumor cells have mutations in genes that cause NF-κB to become overactive. Blocking NF-κB could cause tumor cells to stop growing, die, or become more sensitive to therapeutics.

Late-phase synthesis of IκBα insulates the TLR4-activated canonical NF-κB pathway from noncanonical NF-κB signaling in macrophages

PubMed Central

Mukherjee, Tapas; Taye, Nandaraj; Vijayaragavan, Bharath; Chattopadhyay, Samit; Gomes, James; Basak, Soumen

2017-01-01

The nuclear factor κB (NF-κB) transcription factors coordinate the inflammatory immune response during microbial infection. Pathogenic substances engage canonical NF-κB signaling through the heterodimer RelA:p50, which is subjected to rapid negative feedback by inhibitor of κBα (IκBα). The noncanonical NF-κB pathway is required for the differentiation of immune cells; however, crosstalk between both pathways can occur. Concomitantly activated noncanonical signaling generates p52 from the p100 precursor. The synthesis of p100 is induced by canonical signaling, leading to formation of the late-acting RelA:p52 heterodimer. This crosstalk prolongs inflammatory RelA activity in epithelial cells to ensure pathogen clearance. We found that the Toll-like receptor 4 (TLR4)–activated canonical NF-κB signaling pathway is insulated from lymphotoxin β receptor (LTβR)–induced noncanonical signaling in mouse macrophage cell lines. Combined computational and biochemical studies indicated that the extent of NF-κB–responsive expression of Nfkbia, which encodes IκBα, inversely correlated with crosstalk. The Nfkbia promoter showed enhanced responsiveness to NF-κB activation in macrophages compared to that in fibroblasts. We found that this hyperresponsive promoter engaged the RelA:p52 dimer generated during costimulation of macrophages through TLR4 and LTβR to trigger synthesis of IκBα at late time points, which prevented the late-acting RelA crosstalk response. Together, these data suggest that despite the presence of identical signaling networks in cells of diverse lineages, emergent crosstalk between signaling pathways is subject to cell type–specific regulation. We propose that the insulation of canonical and noncanonical NF-κB pathways limits the deleterious effects of macrophage-mediated inflammation. PMID:27923915

In vivo functional mapping of the conserved protein domains within murine Themis1.

PubMed

Zvezdova, Ekaterina; Lee, Jan; El-Khoury, Dalal; Barr, Valarie; Akpan, Itoro; Samelson, Lawrence; Love, Paul E

2014-09-01

Thymocyte development requires the coordinated input of signals that originate from numerous cell surface molecules. Although the majority of thymocyte signal-initiating receptors are lineage-specific, most trigger 'ubiquitous' downstream signaling pathways. T-lineage-specific receptors are coupled to these signaling pathways by lymphocyte-restricted adapter molecules. We and others recently identified a new putative adapter protein, Themis1, whose expression is largely restricted to the T lineage. Mice lacking Themis1 exhibit a severe block in thymocyte development and a striking paucity of mature T cells revealing a critical role for Themis1 in T-cell maturation. Themis1 orthologs contain three conserved domains: a proline-rich region (PRR) that binds to the ubiquitous cytosolic adapter Grb2, a nuclear localization sequence (NLS), and two copies of a novel cysteine-containing globular (CABIT) domain. In the present study, we evaluated the functional importance of each of these motifs by retroviral reconstitution of Themis1(-/-) progenitor cells. The results demonstrate an essential requirement for the PRR and NLS motifs but not the conserved CABIT cysteines for Themis1 function.

Sirtuins and the metabolic hurdles in cancer

PubMed Central

German, Natalie J.; Haigis, Marcia C.

2017-01-01

The metabolic demands of cancer cannot be met by normal cell metabolism. Cancer cells undergo dramatic alteration of metabolic pathways in a process called reprogramming, characterized by increased nutrient uptake and re-purposing of these fuels for biosynthetic, bioenergetic or signaling pathways. Partitioning carbon sources toward growth and away from ATP production necessitates other means of generating energy for biosynthetic reactions. Additionally, cancer cell adaptations frequently leads to increased production of reactive oxygen species and lactic acid- metabolites which can be beneficial to cancer growth but also are potentially toxic and must be appropriately cleared. Sirtuins are a family of deacylases and ADP-ribosyltransferases with clear links to the regulation of cancer metabolism. Through their unique ability to integrate cellular stress and nutrient status with coordination of metabolic outputs, sirtuins are well poised to play pivotal roles in tumor metabolism. Here, we review the multi-faceted duties of sirtuins in tackling the metabolic hurdles in cancer. We focus on both beneficial and adverse effects of sirtuins in the regulation of energetic, biosynthetic and toxicity barriers faced by cancer cells. PMID:26126285

Joint Attention without Gaze Following: Human Infants and Their Parents Coordinate Visual Attention to Objects through Eye-Hand Coordination

PubMed Central

Yu, Chen; Smith, Linda B.

2013-01-01

The coordination of visual attention among social partners is central to many components of human behavior and human development. Previous research has focused on one pathway to the coordination of looking behavior by social partners, gaze following. The extant evidence shows that even very young infants follow the direction of another's gaze but they do so only in highly constrained spatial contexts because gaze direction is not a spatially precise cue as to the visual target and not easily used in spatially complex social interactions. Our findings, derived from the moment-to-moment tracking of eye gaze of one-year-olds and their parents as they actively played with toys, provide evidence for an alternative pathway, through the coordination of hands and eyes in goal-directed action. In goal-directed actions, the hands and eyes of the actor are tightly coordinated both temporally and spatially, and thus, in contexts including manual engagement with objects, hand movements and eye movements provide redundant information about where the eyes are looking. Our findings show that one-year-olds rarely look to the parent's face and eyes in these contexts but rather infants and parents coordinate looking behavior without gaze following by attending to objects held by the self or the social partner. This pathway, through eye-hand coupling, leads to coordinated joint switches in visual attention and to an overall high rate of looking at the same object at the same time, and may be the dominant pathway through which physically active toddlers align their looking behavior with a social partner. PMID:24236151

A Chemical Biology Approach to Interrogate Quorum Sensing Regulated Behaviors at the Molecular and Cellular Level

PubMed Central

Lowery, Colin A.; Matamouros, Susana; Niessen, Sherry; Zhu, Jie; Scolnick, Jonathan A.; Mee, Jenny M.; Cravatt, Benjamin F.; Miller, Samuel I.; Kaufmann, Gunnar F.; Janda, Kim D.

2013-01-01

SUMMARY Small molecule probes have been employed extensively to explore biological systems and elucidate cellular signaling pathways. In this study, we utilize an inhibitor of bacterial communication to monitor changes in the proteome of Salmonella enterica serovar Typhimurium with the aim of discovering new processes regulated by AI-2-based quorum sensing (QS), a mechanism of bacterial intracellular communication that allows for the coordination of gene expression in a cell density-dependent manner. In S. typhimurium, this system regulates the uptake and catabolism of intracellular signals and has been implicated in pathogenesis, including the invasion of host epithelial cells. We demonstrate that our QS antagonist is capable of selectively inhibiting the expression of known QS-regulated proteins in S. typhimurium, thus attesting that QS inhibitors may be used to confirm proposed and elucidate previously unidentified QS pathways without relying on genetic manipulation. PMID:23890008

Pathways of cellular proteostasis in aging and disease.

PubMed

Klaips, Courtney L; Jayaraj, Gopal Gunanathan; Hartl, F Ulrich

2018-01-02

Ensuring cellular protein homeostasis, or proteostasis, requires precise control of protein synthesis, folding, conformational maintenance, and degradation. A complex and adaptive proteostasis network coordinates these processes with molecular chaperones of different classes and their regulators functioning as major players. This network serves to ensure that cells have the proteins they need while minimizing misfolding or aggregation events that are hallmarks of age-associated proteinopathies, including neurodegenerative disorders such as Alzheimer's and Parkinson's diseases. It is now clear that the capacity of cells to maintain proteostasis undergoes a decline during aging, rendering the organism susceptible to these pathologies. Here we discuss the major proteostasis pathways in light of recent research suggesting that their age-dependent failure can both contribute to and result from disease. We consider different strategies to modulate proteostasis capacity, which may help develop urgently needed therapies for neurodegeneration and other age-dependent pathologies. © 2018 Klaips et al.

A chemical biology approach to interrogate quorum-sensing regulated behaviors at the molecular and cellular level.

PubMed

Lowery, Colin A; Matamouros, Susana; Niessen, Sherry; Zhu, Jie; Scolnick, Jonathan; Lively, Jenny M; Cravatt, Benjamin F; Miller, Samuel I; Kaufmann, Gunnar F; Janda, Kim D

2013-07-25

Small molecule probes have been used extensively to explore biologic systems and elucidate cellular signaling pathways. In this study, we use an inhibitor of bacterial communication to monitor changes in the proteome of Salmonella enterica serovar Typhimurium with the aim of discovering unrecognized processes regulated by AI-2-based quorum-sensing (QS), a mechanism of bacterial intercellular communication that allows for the coordination of gene expression in a cell density-dependent manner. In S. typhimurium, this system regulates the uptake and catabolism of intercellular signals and has been implicated in pathogenesis, including the invasion of host epithelial cells. We demonstrate that our QS antagonist is capable of selectively inhibiting the expression of known QS-regulated proteins in S. typhimurium, thus attesting that QS inhibitors may be used to confirm proposed and elucidate previously unidentified QS pathways without relying on genetic manipulation. Copyright © 2013 Elsevier Ltd. All rights reserved.

The apical complex couples cell fate and cell survival to cerebral cortical development

PubMed Central

Kim, Seonhee; Lehtinen, Maria K.; Sessa, Alessandro; Zappaterra, Mauro; Cho, Seo-Hee; Gonzalez, Dilenny; Boggan, Brigid; Austin, Christina A.; Wijnholds, Jan; Gambello, Michael J.; Malicki, Jarema; LaMantia, Anthony S.; Broccoli, Vania; Walsh, Christopher A.

2010-01-01

Cortical development depends upon tightly controlled cell fate and cell survival decisions that generate a functional neuronal population, but the coordination of these two processes is poorly understood. Here we show that conditional removal of a key apical complex protein, Pals1, causes premature withdrawal from the cell cycle, inducing excessive generation of early-born postmitotic neurons followed by surprisingly massive and rapid cell death, leading to the abrogation of virtually the entire cortical structure. Pals1 loss shows exquisite dosage sensitivity, so that heterozygote mutants show an intermediate phenotype on cell fate and cell death. Loss of Pals1 blocks essential cell survival signals, including the mammalian target of rapamycin (mTOR) pathway, while mTORC1 activation partially rescues Pals1 deficiency. These data highlight unexpected roles of the apical complex protein Pals1 in cell survival through interactions with mTOR signaling. PMID:20399730

Intracellular signaling by phospholipase D as a therapeutic target.

PubMed

Steed, P M; Chow, A H

2001-09-01

The pharmaceutical industry has recently focused on intracellular signaling as a means to integrate the multiple facets of complex disease states, such as inflammation, because these pathways respond to numerous extracellular signals and coordinate a collection of cell responses contributing to pathology. One critical aspect of intracellular signaling is regulation of key cell functions by lipid mediators, in particular the generation of a key mediator, phosphatidic acid (PA) via the hydrolysis of phosphatidylcholine by phospholipase D (PLD). Research in this field has intensified, due in part to the recent cloning and partial characterization of the two PLD isoforms in mammalian cells, and this work has contributed significantly to our understanding of events downstream of PA generation. It is these effector functions of PLD activity that make this pathway attractive as a therapeutic target while the biochemical properties of the PLD isozymes make them amenable to small molecule intervention. Recent studies indicate that PA, and its immediate metabolites diacylglycerol and lyso-PA, affect numerous cellular pathways including ligand-mediated secretion, cytoskeletal reorganisations, respiratory burst, prostaglandin release, cell migration, cytokine release, and mitogenesis. This review summarises the data implicating signaling via PLD in these cell functions, obtained from: (i) molecular analyses of PLD/effector interactions, (ii) correlation between PA production and cell responses, (iii) experimental manipulation of PA levels, (iv) inhibition of PLD regulators, and (v) direct inhibition of PA production. The utility of targeting PLD signaling for the treatment of acute/chronic inflammation and other indications is discussed in light of these data.

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

PubMed

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

2016-11-15

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

Guiding Principles for Data Architecture to Support the Pathways Community HUB Model

PubMed Central

Zeigler, Bernard P.; Redding, Sarah; Leath, Brenda A.; Carter, Ernest L.; Russell, Cynthia

2016-01-01

Introduction: The Pathways Community HUB Model provides a unique strategy to effectively supplement health care services with social services needed to overcome barriers for those most at risk of poor health outcomes. Pathways are standardized measurement tools used to define and track health and social issues from identification through to a measurable completion point. The HUB use Pathways to coordinate agencies and service providers in the community to eliminate the inefficiencies and duplication that exist among them. Pathways Community HUB Model and Formalization: Experience with the Model has brought out the need for better information technology solutions to support implementation of the Pathways themselves through decision-support tools for care coordinators and other users to track activities and outcomes, and to facilitate reporting. Here we provide a basis for discussing recommendations for such a data infrastructure by developing a conceptual model that formalizes the Pathway concept underlying current implementations. Requirements for Data Architecture to Support the Pathways Community HUB Model: The main contribution is a set of core recommendations as a framework for developing and implementing a data architecture to support implementation of the Pathways Community HUB Model. The objective is to present a tool for communities interested in adopting the Model to learn from and to adapt in their own development and implementation efforts. Problems with Quality of Data Extracted from the CHAP Database: Experience with the Community Health Access Project (CHAP) data base system (the core implementation of the Model) has identified several issues and remedies that have been developed to address these issues. Based on analysis of issues and remedies, we present several key features for a data architecture meeting the just mentioned recommendations. Implementation of Features: Presentation of features is followed by a practical guide to their implementation allowing an organization to consider either tailoring off-the-shelf generic systems to meet the requirements or offerings that are specialized for community-based care coordination. Discussion: Looking to future extensions, we discuss the utility and prospects for an ontology to include care coordination in the Unified Medical Language System (UMLS) of the National Library of Medicine and other existing medical and nursing taxonomies. Conclusions and Recommendations: Pathways structures are an important principle, not only for organizing the care coordination activities, but also for structuring the data stored in electronic form in the conduct of such care. We showed how the proposed architecture encourages design of effective decision support systems for coordinated care and suggested how interested organizations can set about acquiring such systems. Although the presentation focuses on the Pathways Community HUB Model, the principles for data architecture are stated in generic form and are applicable to any health information system for improving care coordination services and population health. PMID:26870743

A proposed route to independent measurements of tight junction conductance at discrete cell junctions

PubMed Central

Zhou, Lushan; Zeng, Yuhan; Baker, Lane A; Hou, Jianghui

2015-01-01

Direct recording of tight junction permeability is of pivotal importance to many biologic fields. Previous approaches bear an intrinsic disadvantage due to the difficulty of separating tight junction conductance from nearby membrane conductance. Here, we propose the design of Double whole-cell Voltage Clamp - Ion Conductance Microscopy (DVC-ICM) based on previously demonstrated potentiometric scanning of local conductive pathways. As proposed, DVC-ICM utilizes two coordinated whole-cell patch-clamps to neutralize the apical membrane current during potentiometric scanning, which in models described here will profoundly enhance the specificity of tight junction recording. Several potential pitfalls are considered, evaluated and addressed with alternative countermeasures. PMID:26716077

Interferon-inducible effector mechanisms in cell-autonomous immunity

PubMed Central

MacMicking, John D.

2014-01-01

Interferons (IFNs) induce the expression of hundreds of genes as part of an elaborate antimicrobial programme designed to combat infection in all nucleated cells — a process termed cell-autonomous immunity. As described in this Review, recent genomic and subgenomic analyses have begun to assign functional properties to novel IFN-inducible effector proteins that restrict bacteria, protozoa and viruses in different subcellular compartments and at different stages of the pathogen life cycle. Several newly described host defence factors also participate in canonical oxidative and autophagic pathways by spatially coordinating their activities to enhance microbial killing. Together, these IFN-induced effector networks help to confer vertebrate host resistance to a vast and complex microbial world. PMID:22531325

miR148b is a major coordinator of breast cancer progression in a relapse-associated microRNA signature by targeting ITGA5, ROCK1, PIK3CA, NRAS, and CSF1.

PubMed

Cimino, Daniela; De Pittà, Cristiano; Orso, Francesca; Zampini, Matteo; Casara, Silvia; Penna, Elisa; Quaglino, Elena; Forni, Marco; Damasco, Christian; Pinatel, Eva; Ponzone, Riccardo; Romualdi, Chiara; Brisken, Cathrin; De Bortoli, Michele; Biglia, Nicoletta; Provero, Paolo; Lanfranchi, Gerolamo; Taverna, Daniela

2013-03-01

Breast cancer is often fatal during its metastatic dissemination. To unravel the role of microRNAs (miRs) during malignancy, we analyzed miR expression in 77 primary breast carcinomas and identified 16 relapse-associated miRs that correlate with survival and/or distinguish tumor subtypes in different datasets. Among them, miR-148b, down-regulated in aggressive breast tumors, was found to be a major coordinator of malignancy. In fact, it is able to oppose various steps of tumor progression when overexpressed in cell lines by influencing invasion, survival to anoikis, extravasation, lung metastasis formation, and chemotherapy response. miR-148b controls malignancy by coordinating a novel pathway involving over 130 genes and, in particular, it directly targets players of the integrin signaling, such as ITGA5, ROCK1, PIK3CA/p110α, and NRAS, as well as CSF1, a growth factor for stroma cells. Our findings reveal the importance of the identified 16 miRs for disease outcome predictions and suggest a critical role for miR-148b in the control of breast cancer progression.

Endothelin and hepatic wound healing

PubMed Central

Khimji, Al-karim; Rockey, Don C.

2014-01-01

Liver wound healing is a coordinated response to injury caused by infections (hepatitis) or toxins (alcohol) or other processes where activation of hepatic stellate cells are a central component. During stellate cell activation, a major phenotypic transformation occurs which leads to increased production of increased extracellular matrix proteins and smooth muscle α-actin the results is organ dysfunction due to gross architectural disruption and impaired blood flow. Endothelin-1 (ET-1) is produced in increased amounts and the cellular source of ET-1 shifts from endothelial cells to stellate cells during liver injury thus setting a feedback loop which accentuates further activation, stellate cell proliferation, and production of extracellular matrix proteins. Therapy directed at intervening the ET-1 signaling pathway has significant therapeutic potential in patients with liver disease. PMID:21421048

Mesenchymal-epithelial interactions during hair follicle morphogenesis and cycling

PubMed Central

Sennett, Rachel; Rendl, Michael

2012-01-01

Embryonic hair follicle induction and formation are regulated by mesenchymal-epithelial interactions between specialized dermal cells and epidermal stem cells that switch to a hair fate. Similarly, during postnatal hair growth, communication between mesenchymal dermal papilla cells and surrounding epithelial matrix cells coordinates hair shaft production. Adult hair follicle regeneration in the hair cycle again is thought to be controlled by activating signals originating from the mesenchymal compartment and acting on hair follicle stem cells. Although many signaling pathways are implicated in hair follicle formation and growth, the precise nature, timing, and intersection of these inductive and regulatory signals remains elusive. The goal of this review is to summarize our current understanding and to discuss recent new insights into mesenchymal-epithelial interactions during hair follicle morphogenesis and cycling. PMID:22960356

R-spondin 1 is required for specification of hematopoietic stem cells through Wnt16 and Vegfa signaling pathways

PubMed Central

2017-01-01

Hematopoietic stem cells (HSCs) are the therapeutic component of bone marrow transplants, but finding immune-compatible donors limits treatment availability and efficacy. Recapitulation of endogenous specification during development is a promising approach to directing HSC specification in vitro, but current protocols are not capable of generating authentic HSCs with high efficiency. Across phyla, HSCs arise from hemogenic endothelium in the ventral floor of the dorsal aorta concurrent with arteriovenous specification and intersegmental vessel (ISV) sprouting, processes regulated by Notch and Wnt. We hypothesized that coordination of HSC specification with vessel patterning might involve modulatory regulatory factors such as R-spondin 1 (Rspo1), an extracellular protein that enhances β-catenin-dependent Wnt signaling and has previously been shown to regulate ISV patterning. We find that Rspo1 is required for HSC specification through control of parallel signaling pathways controlling HSC specification: Wnt16/DeltaC/DeltaD and Vegfa/Tgfβ1. Our results define Rspo1 as a key upstream regulator of two crucial pathways necessary for HSC specification. PMID:28087636

Reciprocal feedback regulation of PI3K and androgen receptor signaling in PTEN-deficient prostate cancer.

PubMed

Carver, Brett S; Chapinski, Caren; Wongvipat, John; Hieronymus, Haley; Chen, Yu; Chandarlapaty, Sarat; Arora, Vivek K; Le, Carl; Koutcher, Jason; Scher, Howard; Scardino, Peter T; Rosen, Neal; Sawyers, Charles L

2011-05-17

Prostate cancer is characterized by its dependence on androgen receptor (AR) and frequent activation of PI3K signaling. We find that AR transcriptional output is decreased in human and murine tumors with PTEN deletion and that PI3K pathway inhibition activates AR signaling by relieving feedback inhibition of HER kinases. Similarly, AR inhibition activates AKT signaling by reducing levels of the AKT phosphatase PHLPP. Thus, these two oncogenic pathways cross-regulate each other by reciprocal feedback. Inhibition of one activates the other, thereby maintaining tumor cell survival. However, combined pharmacologic inhibition of PI3K and AR signaling caused near-complete prostate cancer regressions in a Pten-deficient murine prostate cancer model and in human prostate cancer xenografts, indicating that both pathways coordinately support survival. Copyright © 2011 Elsevier Inc. All rights reserved.

FGF and retinoic acid activity gradients control the timing of neural crest cell emigration in the trunk

PubMed Central

Martínez-Morales, Patricia L.; Diez del Corral, Ruth; Olivera-Martínez, Isabel; Quiroga, Alejandra C.; Das, Raman M.; Barbas, Julio A.; Storey, Kate G.

2011-01-01

Coordination between functionally related adjacent tissues is essential during development. For example, formation of trunk neural crest cells (NCCs) is highly influenced by the adjacent mesoderm, but the molecular mechanism involved is not well understood. As part of this mechanism, fibroblast growth factor (FGF) and retinoic acid (RA) mesodermal gradients control the onset of neurogenesis in the extending neural tube. In this paper, using gain- and loss-of-function experiments, we show that caudal FGF signaling prevents premature specification of NCCs and, consequently, premature epithelial–mesenchymal transition (EMT) to allow cell emigration. In contrast, rostrally generated RA promotes EMT of NCCs at somitic levels. Furthermore, we show that FGF and RA signaling control EMT in part through the modulation of elements of the bone morphogenetic protein and Wnt signaling pathways. These data establish a clear role for opposition of FGF and RA signaling in control of the timing of NCC EMT and emigration and, consequently, coordination of the development of the central and peripheral nervous system during vertebrate trunk elongation. PMID:21807879

A central role for vesicle trafficking in epithelial neoplasia: Intracellular highways to carcinogenesis

PubMed Central

Goldenring, James R.

2014-01-01

Epithelial cell carcinogenesis involves the loss of polarity, alteration of polarized protein presentation, dynamic cell morphology changes, increased proliferation and increased cell motility and invasion. Elements of membrane vesicle trafficking underlie all of these processes. Specific membrane trafficking regulators, including Rab small GTPases, through the coordinated dynamics of intracellular trafficking along cytoskeletal pathways, determine cell surface presentation of proteins and overall function of both differentiated and neoplastic cells. While mutations in vesicle trafficking proteins may not be direct drivers of transformation, elements of the machinery of vesicle movement play critical roles in the phenotypes of neoplastic cells. Therefore, the regulators of membrane vesicle trafficking decisions are critical mediators of the full spectrum of cell physiologies driving cancer cell biology, including initial loss of polarity, invasion and metastasis. Targeting of these fundamental intracellular processes may provide important points for manipulation of cancer cell behaviour. PMID:24108097

Guiding Principles for Data Architecture to Support the Pathways Community HUB Model.

PubMed

Zeigler, Bernard P; Redding, Sarah; Leath, Brenda A; Carter, Ernest L; Russell, Cynthia

2016-01-01

The Pathways Community HUB Model provides a unique strategy to effectively supplement health care services with social services needed to overcome barriers for those most at risk of poor health outcomes. Pathways are standardized measurement tools used to define and track health and social issues from identification through to a measurable completion point. The HUB use Pathways to coordinate agencies and service providers in the community to eliminate the inefficiencies and duplication that exist among them. Experience with the Model has brought out the need for better information technology solutions to support implementation of the Pathways themselves through decision-support tools for care coordinators and other users to track activities and outcomes, and to facilitate reporting. Here we provide a basis for discussing recommendations for such a data infrastructure by developing a conceptual model that formalizes the Pathway concept underlying current implementations. The main contribution is a set of core recommendations as a framework for developing and implementing a data architecture to support implementation of the Pathways Community HUB Model. The objective is to present a tool for communities interested in adopting the Model to learn from and to adapt in their own development and implementation efforts. Experience with the Community Health Access Project (CHAP) data base system (the core implementation of the Model) has identified several issues and remedies that have been developed to address these issues. Based on analysis of issues and remedies, we present several key features for a data architecture meeting the just mentioned recommendations. Presentation of features is followed by a practical guide to their implementation allowing an organization to consider either tailoring off-the-shelf generic systems to meet the requirements or offerings that are specialized for community-based care coordination. Looking to future extensions, we discuss the utility and prospects for an ontology to include care coordination in the Unified Medical Language System (UMLS) of the National Library of Medicine and other existing medical and nursing taxonomies. Pathways structures are an important principle, not only for organizing the care coordination activities, but also for structuring the data stored in electronic form in the conduct of such care. We showed how the proposed architecture encourages design of effective decision support systems for coordinated care and suggested how interested organizations can set about acquiring such systems. Although the presentation focuses on the Pathways Community HUB Model, the principles for data architecture are stated in generic form and are applicable to any health information system for improving care coordination services and population health.

Comprehensive analysis of tobacco pollen transcriptome unveils common pathways in polar cell expansion and underlying heterochronic shift during spermatogenesis

PubMed Central

2012-01-01

Background Many flowering plants produce bicellular pollen. The two cells of the pollen grain are destined for separate fates in the male gametophyte, which provides a unique opportunity to study genetic interactions that govern guided single-cell polar expansion of the growing pollen tube and the coordinated control of germ cell division and sperm cell fate specification. We applied the Agilent 44 K tobacco gene chip to conduct the first transcriptomic analysis of the tobacco male gametophyte. In addition, we performed a comparative study of the Arabidopsis root-hair trichoblast transcriptome to evaluate genetic factors and common pathways involved in polarized cell-tip expansion. Results Progression of pollen grains from freshly dehisced anthers to pollen tubes 4 h after germination is accompanied with > 5,161 (14.9%) gametophyte-specific expressed probes active in at least one of the developmental stages. In contrast, > 18,821 (54.4%) probes were preferentially expressed in the sporophyte. Our comparative approach identified a subset of 104 pollen tube-expressed genes that overlap with root-hair trichoblasts. Reverse genetic analysis of selected candidates demonstrated that Cu/Zn superoxide dismutase 1 (CSD1), a WD-40 containing protein (BP130384), and Replication factor C1 (NtRFC1) are among the central regulators of pollen-tube tip growth. Extension of our analysis beyond the second haploid mitosis enabled identification of an opposing-dynamic accumulation of core regulators of cell proliferation and cell fate determinants in accordance with the progression of the germ cell cycle. Conclusions The current study provides a foundation to isolate conserved regulators of cell tip expansion and those that are unique for pollen tube growth to the female gametophyte. A transcriptomic data set is presented as a benchmark for future functional studies using developing pollen as a model. Our results demonstrated previously unknown functions of certain genes in pollen-tube tip growth. In addition, we highlighted the molecular dynamics of core cell-cycle regulators in the male gametophyte and postulated the first genetic model to account for the differential timing of spermatogenesis among angiosperms and its coordination with female gametogenesis. PMID:22340370

RhoA GTPase inhibition organizes contraction during epithelial morphogenesis

PubMed Central

Mason, Frank M.; Xie, Shicong; Vasquez, Claudia G.; Tworoger, Michael

2016-01-01

During morphogenesis, contraction of the actomyosin cytoskeleton within individual cells drives cell shape changes that fold tissues. Coordination of cytoskeletal contractility is mediated by regulating RhoA GTPase activity. Guanine nucleotide exchange factors (GEFs) activate and GTPase-activating proteins (GAPs) inhibit RhoA activity. Most studies of tissue folding, including apical constriction, have focused on how RhoA is activated by GEFs to promote cell contractility, with little investigation as to how GAPs may be important. Here, we identify a critical role for a RhoA GAP, Cumberland GAP (C-GAP), which coordinates with a RhoA GEF, RhoGEF2, to organize spatiotemporal contractility during Drosophila melanogaster apical constriction. C-GAP spatially restricts RhoA pathway activity to a central position in the apical cortex. RhoGEF2 pulses precede myosin, and C-GAP is required for pulsation, suggesting that contractile pulses result from RhoA activity cycling. Finally, C-GAP expression level influences the transition from reversible to irreversible cell shape change, which defines the onset of tissue shape change. Our data demonstrate that RhoA activity cycling and modulating the ratio of RhoGEF2 to C-GAP are required for tissue folding. PMID:27551058

Gap junctions in cells of the immune system: structure, regulation and possible functional roles.

PubMed

Sáez, J C; Brañes, M C; Corvalán, L A; Eugenín, E A; González, H; Martínez, A D; Palisson, F

2000-04-01

Gap junction channels are sites of cytoplasmic communication between contacting cells. In vertebrates, they consist of protein subunits denoted connexins (Cxs) which are encoded by a gene family. According to their Cx composition, gap junction channels show different gating and permeability properties that define which ions and small molecules permeate them. Differences in Cx primary sequences suggest that channels composed of different Cxs are regulated differentially by intracellular pathways under specific physiological conditions. Functional roles of gap junction channels could be defined by the relative importance of permeant substances, resulting in coordination of electrical and/or metabolic cellular responses. Cells of the native and specific immune systems establish transient homo- and heterocellular contacts at various steps of the immune response. Morphological and functional studies reported during the last three decades have revealed that many intercellular contacts between cells in the immune response present gap junctions or "gap junction-like" structures. Partial characterization of the molecular composition of some of these plasma membrane structures and regulatory mechanisms that control them have been published recently. Studies designed to elucidate their physiological roles suggest that they might permit coordination of cellular events which favor the effective and timely response of the immune system.

De-novo NAD+ synthesis regulates SIRT1-FOXO1 apoptotic pathway in response to NQO1 substrates in lung cancer cells

PubMed Central

Cheng, Xuefang; Li, Qingran; Liu, Fang; Ye, Hui; Zhao, Min; Wang, Hong; Wang, Guangji; Hao, Haiping

2016-01-01

Tryptophan metabolism is essential in diverse kinds of tumors via regulating tumor immunology. However, the direct role of tryptophan metabolism and its signaling pathway in cancer cells remain largely elusive. Here, we establish a mechanistic link from L-type amino acid transporter 1 (LAT1) mediated transport of tryptophan and the subsequent de-novo NAD+ synthesis to SIRT1-FOXO1 regulated apoptotic signaling in A549 cells in response to NQO1 activation. In response to NQO1 activation, SIRT1 is repressed leading to the increased cellular accumulation of acetylated FOXO1 that transcriptionally activates apoptotic signaling. Decreased uptake of tryptophan due to the downregulation of LAT1 coordinates with PARP-1 hyperactivation to induce rapid depletion of NAD+ pool. Particularly, the LAT1-NAD+-SIRT1 signaling is activated in tumor tissues of patients with non-small cell lung cancer. Because NQO1 activation is characterized with oxidative challenge induced DNA damage, these results suggest that LAT1 and de-novo NAD+ synthesis in NSCLC cells may play essential roles in sensing excessive oxidative stress. PMID:27566573

On the Evolution of the Cardiac Pacemaker

PubMed Central

Burkhard, Silja; van Eif, Vincent; Garric, Laurence; Christoffels, Vincent M.; Bakkers, Jeroen

2017-01-01

The rhythmic contraction of the heart is initiated and controlled by an intrinsic pacemaker system. Cardiac contractions commence at very early embryonic stages and coordination remains crucial for survival. The underlying molecular mechanisms of pacemaker cell development and function are still not fully understood. Heart form and function show high evolutionary conservation. Even in simple contractile cardiac tubes in primitive invertebrates, cardiac function is controlled by intrinsic, autonomous pacemaker cells. Understanding the evolutionary origin and development of cardiac pacemaker cells will help us outline the important pathways and factors involved. Key patterning factors, such as the homeodomain transcription factors Nkx2.5 and Shox2, and the LIM-homeodomain transcription factor Islet-1, components of the T-box (Tbx), and bone morphogenic protein (Bmp) families are well conserved. Here we compare the dominant pacemaking systems in various organisms with respect to the underlying molecular regulation. Comparative analysis of the pathways involved in patterning the pacemaker domain in an evolutionary context might help us outline a common fundamental pacemaker cell gene programme. Special focus is given to pacemaker development in zebrafish, an extensively used model for vertebrate development. Finally, we conclude with a summary of highly conserved key factors in pacemaker cell development and function. PMID:29367536

A coordinated phosphorylation cascade initiated by p38MAPK/MSK1 directs RARα to target promoters

PubMed Central

Bruck, Nathalie; Vitoux, Dominique; Ferry, Christine; Duong, Vanessa; Bauer, Annie; de Thé, Hughes; Rochette-Egly, Cécile

2009-01-01

The nuclear retinoic acid (RA) receptor alpha (RARα) is a transcriptional transregulator that controls the expression of specific gene subsets through binding at response elements and dynamic interactions with coregulators, which are coordinated by the ligand. Here, we highlighted a novel paradigm in which the transcription of RARα target genes is controlled by phosphorylation cascades initiated by the rapid RA activation of the p38MAPK/MSK1 pathway. We demonstrate that MSK1 phosphorylates RARα at S369 located in the ligand-binding domain, allowing the binding of TFIIH and thereby phosphorylation of the N-terminal domain at S77 by cdk7/cyclin H. MSK1 also phosphorylates histone H3 at S10. Finally, the phosphorylation cascade initiated by MSK1 controls the recruitment of RARα/TFIIH complexes to response elements and subsequently RARα target gene activation. Cancer cells characterized by a deregulated p38MAPK/MSK1 pathway, do not respond to RA, outlining the essential contribution of the RA-triggered phosphorylation cascade in RA signalling. PMID:19078967

Coordinated Activation of Cellulose and Repression of Lignin Biosynthesis Pathways in Rice1[C][W][OA

PubMed Central

Ambavaram, Madana M.R.; Krishnan, Arjun; Trijatmiko, Kurniawan R.; Pereira, Andy

2011-01-01

Cellulose from plant biomass is the largest renewable energy resource of carbon fixed from the atmosphere, which can be converted into fermentable sugars for production into ethanol. However, the cellulose present as lignocellulosic biomass is embedded in a hemicellulose and lignin matrix from which it needs to be extracted for efficient processing. Here, we show that expression of an Arabidopsis (Arabidopsis thaliana) transcription factor, SHINE (SHN), in rice (Oryza sativa), a model for the grasses, causes a 34% increase in cellulose and a 45% reduction in lignin content. The rice AtSHN lines also exhibit an altered lignin composition correlated with improved digestibility, with no compromise in plant strength and performance. Using a detailed systems-level analysis of global gene expression in rice, we reveal the SHN regulatory network coordinating down-regulation of lignin biosynthesis and up-regulation of cellulose and other cell wall biosynthesis pathway genes. The results thus support the development of nonfood crops and crop wastes with increased cellulose and low lignin with good agronomic performance that could improve the economic viability of lignocellulosic crop utilization for biofuels. PMID:21205614

The Notch pathway regulates the Second Mitotic Wave cell cycle independently of bHLH proteins.

PubMed

Bhattacharya, Abhishek; Li, Ke; Quiquand, Manon; Rimesso, Gerard; Baker, Nicholas E

2017-11-15

Notch regulates both neurogenesis and cell cycle activity to coordinate precursor cell generation in the differentiating Drosophila eye. Mosaic analysis with mitotic clones mutant for Notch components was used to identify the pathway of Notch signaling that regulates the cell cycle in the Second Mitotic Wave. Although S phase entry depends on Notch signaling and on the transcription factor Su(H), the transcriptional co-activator Mam and the bHLH repressor genes of the E(spl)-Complex were not essential, although these are Su(H) coactivators and targets during the regulation of neurogenesis. The Second Mitotic Wave showed little dependence on ubiquitin ligases neuralized or mindbomb, and although the ligand Delta is required non-autonomously, partial cell cycle activity occurred in the absence of known Notch ligands. We found that myc was not essential for the Second Mitotic Wave. The Second Mitotic Wave did not require the HLH protein Extra macrochaetae, and the bHLH protein Daughterless was required only cell-nonautonomously. Similar cell cycle phenotypes for Daughterless and Atonal were consistent with requirement for neuronal differentiation to stimulate Delta expression, affecting Notch activity in the Second Mitotic Wave indirectly. Therefore Notch signaling acts to regulate the Second Mitotic Wave without activating bHLH gene targets. Copyright © 2017 Elsevier Inc. All rights reserved.

Plant cell wall signalling and receptor-like kinases.

PubMed

Wolf, Sebastian

2017-02-15

Communication between the extracellular matrix and the cell interior is essential for all organisms as intrinsic and extrinsic cues have to be integrated to co-ordinate development, growth, and behaviour. This applies in particular to plants, the growth and shape of which is governed by deposition and remodelling of the cell wall, a rigid, yet dynamic, extracellular network. It is thus generally assumed that cell wall surveillance pathways exist to monitor the state of the wall and, if needed, elicit compensatory responses such as altered expression of cell wall remodelling and biosynthesis genes. Here, I highlight recent advances in the field of cell wall signalling in plants, with emphasis on the role of plasma membrane receptor-like kinase complexes. In addition, possible roles for cell wall-mediated signalling beyond the maintenance of cell wall integrity are discussed. © 2017 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

The role of the DNA sliding clamp in Okazaki fragment maturation in archaea and eukaryotes.

PubMed

Beattie, Thomas R; Bell, Stephen D

2011-01-01

Efficient processing of Okazaki fragments generated during discontinuous lagging-strand DNA replication is critical for the maintenance of genome integrity. In eukaryotes, a number of enzymes co-ordinate to ensure the removal of initiating primers from the 5'-end of each fragment and the generation of a covalently linked daughter strand. Studies in eukaryotic systems have revealed that the co-ordination of DNA polymerase δ and FEN-1 (Flap Endonuclease 1) is sufficient to remove the majority of primers. Other pathways such as that involving Dna2 also operate under certain conditions, although, notably, Dna2 is not universally conserved between eukaryotes and archaea, unlike the other core factors. In addition to the catalytic components, the DNA sliding clamp, PCNA (proliferating-cell nuclear antigen), plays a pivotal role in binding and co-ordinating these enzymes at sites of lagging-strand replication. Structural studies in eukaryotic and archaeal systems have revealed that PCNA-binding proteins can adopt different conformations when binding PCNA. This conformational malleability may be key to the co-ordination of these enzymes' activities.

Transcriptional control of monolignol biosynthesis in Pinus taeda: factors affecting monolignol ratios and carbon allocation in phenylpropanoid metabolism

NASA Technical Reports Server (NTRS)

Anterola, Aldwin M.; Jeon, Jae-Heung; Davin, Laurence B.; Lewis, Norman G.

2002-01-01

Transcriptional profiling of the phenylpropanoid pathway in Pinus taeda cell suspension cultures was carried out using quantitative real time PCR analyses of all known genes involved in the biosynthesis of the two monolignols, p-coumaryl and coniferyl alcohols (lignin/lignan precursors). When the cells were transferred to a medium containing 8% sucrose and 20 mm potassium iodide, the monolignol/phenylpropanoid pathway was induced, and transcript levels for phenylalanine ammonia lyase, cinnamate 4-hydroxylase, p-coumarate 3-hydroxylase, 4-coumarate:CoA ligase, caffeoyl-CoA O-methyltransferase, cinnamoyl-CoA reductase, and cinnamyl alcohol dehydrogenase were coordinately up-regulated. Provision of increasing levels of exogenously supplied Phe to saturating levels (40 mm) to the induction medium resulted in further up-regulation of their transcript levels in the P. taeda cell cultures; this in turn was accompanied by considerable increases in both p-coumaryl and coniferyl alcohol formation and excretion. By contrast, transcript levels for both cinnamate 4-hydroxylase and p-coumarate 3-hydroxylase were only slightly up-regulated. These data, when considered together with metabolic profiling results and genetic manipulation of various plant species, reveal that carbon allocation to the pathway and its differential distribution into the two monolignols is controlled by Phe supply and differential modulation of cinnamate 4-hydroxylase and p-coumarate 3-hydroxylase activities, respectively. The coordinated up-regulation of phenylalanine ammonia lyase, 4-coumarate:CoA ligase, caffeoyl-CoA O-methyltransferase, cinnamoyl-CoA reductase and cinnamyl alcohol dehydrogenase in the presence of increasing concentrations of Phe also indicates that these steps are not truly rate-limiting, because they are modulated according to metabolic demand. Finally, the transcript profile of a putative acid/ester O-methyltransferase, proposed as an alternative catalyst for O-methylation leading to coniferyl alcohol, was not up-regulated under any of the conditions employed, suggesting that it is not, in fact, involved in monolignol biosynthesis.

Transcriptional control of monolignol biosynthesis in Pinus taeda: factors affecting monolignol ratios and carbon allocation in phenylpropanoid metabolism.

PubMed

Anterola, Aldwin M; Jeon, Jae-Heung; Davin, Laurence B; Lewis, Norman G

2002-05-24

Transcriptional profiling of the phenylpropanoid pathway in Pinus taeda cell suspension cultures was carried out using quantitative real time PCR analyses of all known genes involved in the biosynthesis of the two monolignols, p-coumaryl and coniferyl alcohols (lignin/lignan precursors). When the cells were transferred to a medium containing 8% sucrose and 20 mm potassium iodide, the monolignol/phenylpropanoid pathway was induced, and transcript levels for phenylalanine ammonia lyase, cinnamate 4-hydroxylase, p-coumarate 3-hydroxylase, 4-coumarate:CoA ligase, caffeoyl-CoA O-methyltransferase, cinnamoyl-CoA reductase, and cinnamyl alcohol dehydrogenase were coordinately up-regulated. Provision of increasing levels of exogenously supplied Phe to saturating levels (40 mm) to the induction medium resulted in further up-regulation of their transcript levels in the P. taeda cell cultures; this in turn was accompanied by considerable increases in both p-coumaryl and coniferyl alcohol formation and excretion. By contrast, transcript levels for both cinnamate 4-hydroxylase and p-coumarate 3-hydroxylase were only slightly up-regulated. These data, when considered together with metabolic profiling results and genetic manipulation of various plant species, reveal that carbon allocation to the pathway and its differential distribution into the two monolignols is controlled by Phe supply and differential modulation of cinnamate 4-hydroxylase and p-coumarate 3-hydroxylase activities, respectively. The coordinated up-regulation of phenylalanine ammonia lyase, 4-coumarate:CoA ligase, caffeoyl-CoA O-methyltransferase, cinnamoyl-CoA reductase and cinnamyl alcohol dehydrogenase in the presence of increasing concentrations of Phe also indicates that these steps are not truly rate-limiting, because they are modulated according to metabolic demand. Finally, the transcript profile of a putative acid/ester O-methyltransferase, proposed as an alternative catalyst for O-methylation leading to coniferyl alcohol, was not up-regulated under any of the conditions employed, suggesting that it is not, in fact, involved in monolignol biosynthesis.

Cancer stem cells and cell size: A causal link?

PubMed

Li, Qiuhui; Rycaj, Kiera; Chen, Xin; Tang, Dean G

2015-12-01

The majority of normal animal cells are 10-20 μm in diameter. Many signaling mechanisms, notably PI3K/Akt/mTOR, Myc, and Hippo pathways, tightly control and coordinate cell growth, cell size, cell division, and cell number during homeostasis. These regulatory mechanisms are frequently deregulated during tumorigenesis resulting in wide variations in cell sizes and increased proliferation in cancer cells. Here, we first review the evidence that primitive stem cells in adult tissues are quiescent and generally smaller than their differentiated progeny, suggesting a correlation between small cell sizes with the stemness. Conversely, increased cell size positively correlates with differentiation phenotypes. We then discuss cancer stem cells (CSCs) and present some evidence that correlates cell sizes with CSC activity. Overall, a causal link between CSCs and cell size is relatively weak and remains to be rigorously assessed. In the future, optimizing methods for isolating cells based on size should help elucidate the connection between cancer cell size and CSC characteristics. Copyright © 2015 Elsevier Ltd. All rights reserved.

Hepatic stellate cell and myofibroblast-like cell gene expression in the explanted cirrhotic livers of patients undergoing liver transplantation.

PubMed

Estep, J Michael; O'Reilly, Linda; Grant, Geraldine; Piper, James; Jonsson, Johann; Afendy, Arian; Chandhoke, Vikas; Younossi, Zobair M

2010-02-01

Hepatic stellate cells (HSC) are involved in hepatic fibrogenesis. Cell signaling associated with an insult to the liver affects an HSC transdifferentiation to fibrogenic myofibroblast-like cells. To investigate the transcriptional expression distinguishing HSC and myofibroblast-like cells between livers with and without cirrhosis. Tissue from ten cirrhotic livers (undergoing transplant) and four non-cirrhotic livers from the National Disease Research Interchange underwent cell separation to extract HSC and myofibroblast-like cell populations. Separated cell types as well as LI-90 cells were subjected to microarray analysis. Selected microarray results were verified by quantitative real-time PCR. Differential expression of some genes, such as IL-1beta, IL-1alpha, and IL-6, was associated with both transdifferentiation and disease. Other genes, such as fatty acid 2-hydroxylase only show differential expression in association with disease. Functional analysis supported these findings, indicating some signal transduction pathways (IL-6) are involved in disease and activation, whereas retinoid X receptor signaling in HSC from cirrhotic and non-cirrhotic livers varies in scope and quality. These findings indicate distinct phenotypes for HSC from cirrhotic and non-cirrhotic livers. Furthermore, coordinated differential expression between genes involved in the same signal transduction pathways provides some insight into the mechanisms that may control the balance between fibrogenesis and fibrolysis.

Coordination of Actin- and Microtubule-Based Cytoskeletons Supports Transport of Spermatids and Residual Bodies/Phagosomes During Spermatogenesis in the Rat Testis

PubMed Central

Tang, Elizabeth I.; Lee, Will M.

2016-01-01

Germ cell transport across the seminiferous epithelium during spermatogenesis requires the intricate coordination of cell junctions, signaling proteins, and both actin- and microtubule (MT)-based cytoskeletons. Although the involvement of cytoskeletons in germ cell transport has been suggested, the precise mechanism(s) remains elusive. Based on growing evidence that actin and MT interactions underlie fundamental cellular processes, such as cell motility, it is unlikely that actin- and MT-based cytoskeletons work independently to regulate germ cell transport in the testis. Using rats treated with adjudin, a potential male contraceptive that disrupts spermatid adhesion and transport in the testis, as a study model, we show herein that actin- and MT-based cytoskeletons are both necessary for transport of spermatids and residual bodies/phagosomes across the seminiferous epithelium in adult rat testes. Analysis of intratubular expression of F-actin and tubulin revealed disruption of both actin and MT networks, concomitant with misdirected spermatids and phagosomes in rats treated with adjudin. Actin regulatory proteins, epidermal growth factor receptor pathway substrate 8 and actin-related protein 3, were mislocalized and down-regulated at the actin-rich anchoring junction between germ and Sertoli cells (apical ectoplasmic specialization) after adjudin treatment. Nonreceptor tyrosine kinase p-FAK-Tyr407, known to regulate F-actin nucleation via actin-related protein 3, was also mislocalized and down-regulated at the apical ectoplasmic specialization, corroborating the observation of actin cytoskeleton disruption. Additionally, spatiotemporal expression of MT regulatory protein end-binding protein 1, shown to be involved in MT-actin cross talk herein, was also disrupted after adjudin treatment. In summary, spermatid/phagosome transport across the epithelium during spermatogenesis requires the coordination between actin- and MT-based cytoskeletons. PMID:26894662

Azospirillum brasilense Chemotaxis Depends on Two Signaling Pathways Regulating Distinct Motility Parameters

PubMed Central

Mukherjee, Tanmoy; Kumar, Dhivya; Burriss, Nathan; Xie, Zhihong

2016-01-01

ABSTRACT The genomes of most motile bacteria encode two or more chemotaxis (Che) systems, but their functions have been characterized in only a few model systems. Azospirillum brasilense is a motile soil alphaproteobacterium able to colonize the rhizosphere of cereals. In response to an attractant, motile A. brasilense cells transiently increase swimming speed and suppress reversals. The Che1 chemotaxis pathway was previously shown to regulate changes in the swimming speed, but it has a minor role in chemotaxis and root surface colonization. Here, we show that a second chemotaxis system, named Che4, regulates the probability of swimming reversals and is the major signaling pathway for chemotaxis and wheat root surface colonization. Experimental evidence indicates that Che1 and Che4 are functionally linked to coordinate changes in the swimming motility pattern in response to attractants. The effect of Che1 on swimming speed is shown to enhance the aerotactic response of A. brasilense in gradients, likely providing the cells with a competitive advantage in the rhizosphere. Together, the results illustrate a novel mechanism by which motile bacteria utilize two chemotaxis pathways regulating distinct motility parameters to alter movement in gradients and enhance the chemotactic advantage. IMPORTANCE Chemotaxis provides motile bacteria with a competitive advantage in the colonization of diverse niches and is a function enriched in rhizosphere bacterial communities, with most species possessing at least two chemotaxis systems. Here, we identify the mechanism by which cells may derive a significant chemotactic advantage using two chemotaxis pathways that ultimately regulate distinct motility parameters. PMID:27068592

Azospirillum brasilense Chemotaxis Depends on Two Signaling Pathways Regulating Distinct Motility Parameters.

PubMed

Mukherjee, Tanmoy; Kumar, Dhivya; Burriss, Nathan; Xie, Zhihong; Alexandre, Gladys

2016-06-15

The genomes of most motile bacteria encode two or more chemotaxis (Che) systems, but their functions have been characterized in only a few model systems. Azospirillum brasilense is a motile soil alphaproteobacterium able to colonize the rhizosphere of cereals. In response to an attractant, motile A. brasilense cells transiently increase swimming speed and suppress reversals. The Che1 chemotaxis pathway was previously shown to regulate changes in the swimming speed, but it has a minor role in chemotaxis and root surface colonization. Here, we show that a second chemotaxis system, named Che4, regulates the probability of swimming reversals and is the major signaling pathway for chemotaxis and wheat root surface colonization. Experimental evidence indicates that Che1 and Che4 are functionally linked to coordinate changes in the swimming motility pattern in response to attractants. The effect of Che1 on swimming speed is shown to enhance the aerotactic response of A. brasilense in gradients, likely providing the cells with a competitive advantage in the rhizosphere. Together, the results illustrate a novel mechanism by which motile bacteria utilize two chemotaxis pathways regulating distinct motility parameters to alter movement in gradients and enhance the chemotactic advantage. Chemotaxis provides motile bacteria with a competitive advantage in the colonization of diverse niches and is a function enriched in rhizosphere bacterial communities, with most species possessing at least two chemotaxis systems. Here, we identify the mechanism by which cells may derive a significant chemotactic advantage using two chemotaxis pathways that ultimately regulate distinct motility parameters. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Human osteocalcin and bone sialoprotein mediating osteomimicry of prostate cancer cells: role of cAMP-dependent protein kinase A signaling pathway.

PubMed

Huang, Wen-Chin; Xie, Zhihui; Konaka, Hiroyuki; Sodek, Jaro; Zhau, Haiyen E; Chung, Leland W K

2005-03-15

Osteocalcin and bone sialoprotein are the most abundant noncollagenous bone matrix proteins expressed by osteoblasts. Surprisingly, osteocalcin and bone sialoprotein are also expressed by malignant but not normal prostate epithelial cells. The purpose of this study is to investigate how osteocalcin and bone sialoprotein expression is regulated in prostate cancer cells. Our investigation revealed that (a) human osteocalcin and bone sialoprotein promoter activities in an androgen-independent prostate cancer cell line of LNCaP lineage, C4-2B, were markedly enhanced 7- to 12-fold in a concentration-dependent manner by conditioned medium collected from prostate cancer and bone stromal cells. (b) Deletion analysis of human osteocalcin and bone sialoprotein promoter regions identified cyclic AMP (cAMP)-responsive elements (CRE) as the critical determinants for conditioned medium-mediated osteocalcin and bone sialoprotein gene expression in prostate cancer cells. Consistent with these results, the protein kinase A (PKA) pathway activators forskolin and dibutyryl cAMP and the PKA pathway inhibitor H-89, respectively, increased or repressed human osteocalcin and bone sialoprotein promoter activities. (c) Electrophoretic mobility shift assay showed that conditioned medium-mediated stimulation of human osteocalcin and bone sialoprotein promoter activities occurs through increased interaction between CRE and CRE-binding protein. (d) Conditioned medium was found to induce human osteocalcin and bone sialoprotein promoter activities via increased CRE/CRE-binding protein interaction in a cell background-dependent manner, with marked stimulation in selected prostate cancer but not bone stromal cells. Collectively, these results suggest that osteocalcin and bone sialoprotein expression is coordinated and regulated through cAMP-dependent PKA signaling, which may define the molecular basis of the osteomimicry exhibited by prostate cancer cells.

New insights into the pathways initiating and driving pancreatitis

PubMed Central

Gukovskaya, Anna S.; Pandol, Stephen J.; Gukovsky, Ilya

2016-01-01

Purpose of review In this article, we discuss recent studies that advance our understanding of molecular and cellular factors initiating and driving pancreatitis, with the emphasis on the role of acinar cell organelle disorders. Recent findings The central physiologic function of the pancreatic acinar cell – to synthesize, store, and secrete digestive enzymes – critically relies on coordinated actions of the endoplasmic reticulum (ER), the endolysosomal system, mitochondria, and autophagy. Recent studies begin to unravel the roles of these organelles’ disordering in the mechanism of pancreatitis. Mice deficient in key autophagy mediators Atg5 or Atg7, or lysosome-associated membrane protein-2, exhibit dysregulation of multiple signaling and metabolic pathways in pancreatic acinar cells and develop spontaneous pancreatitis. Mitochondrial dysfunction caused by sustained opening of the permeability transition pore is shown to mediate pancreatitis in several clinically relevant experimental models, and its inhibition by pharmacologic or genetic means greatly reduces local and systemic pathologic responses. Experimental pancreatitis is also alleviated with inhibitors of ORAI1, a key component of the plasma membrane channel mediating pathologic rise in acinar cell cytosolic Ca2+. Pancreatitis-promoting mutations are increasingly associated with the ER stress. These findings suggest novel pathways and drug targets for pancreatitis treatment. In addition, the recent studies identify new mediators (e.g., neutrophil extracellular traps) of the inflammatory and other responses of pancreatitis. Summary The recent findings illuminate a critical role of organelles regulating the autophagic, endolysosomal, mitochondrial, and ER pathways in maintaining pancreatic acinar cell homeostasis and secretory function; provide compelling evidence that organelle disordering is a key pathogenic mechanism initiating and driving pancreatitis; and identify molecular and cellular factors that could be targeted to restore organellar functions and thus alleviate or treat pancreatitis. PMID:27428704

Matriptase activation connects tissue factor-dependent coagulation initiation to epithelial proteolysis and signaling.

PubMed

Le Gall, Sylvain M; Szabo, Roman; Lee, Melody; Kirchhofer, Daniel; Craik, Charles S; Bugge, Thomas H; Camerer, Eric

2016-06-23

The coagulation cascade is designed to sense tissue injury by physical separation of the membrane-anchored cofactor tissue factor (TF) from inactive precursors of coagulation proteases circulating in plasma. Once TF on epithelial and other extravascular cells is exposed to plasma, sequential activation of coagulation proteases coordinates hemostasis and contributes to host defense and tissue repair. Membrane-anchored serine proteases (MASPs) play critical roles in the development and homeostasis of epithelial barrier tissues; how MASPs are activated in mature epithelia is unknown. We here report that proteases of the extrinsic pathway of blood coagulation transactivate the MASP matriptase, thus connecting coagulation initiation to epithelial proteolysis and signaling. Exposure of TF-expressing cells to factors (F) VIIa and Xa triggered the conversion of latent pro-matriptase to an active protease, which in turn cleaved the pericellular substrates protease-activated receptor-2 (PAR2) and pro-urokinase. An activation pathway-selective PAR2 mutant resistant to direct cleavage by TF:FVIIa and FXa was activated by these proteases when cells co-expressed pro-matriptase, and matriptase transactivation was necessary for efficient cleavage and activation of wild-type PAR2 by physiological concentrations of TF:FVIIa and FXa. The coagulation initiation complex induced rapid and prolonged enhancement of the barrier function of epithelial monolayers that was dependent on matriptase transactivation and PAR2 signaling. These observations suggest that the coagulation cascade engages matriptase to help coordinate epithelial defense and repair programs after injury or infection, and that matriptase may contribute to TF-driven pathogenesis in cancer and inflammation.

Analysis of the ERK1,2 transcriptome in mammary epithelial cells

PubMed Central

Grill, Constance; Gheyas, Ferdous; Dayananth, Priya; Jin, Weihong; Ding, Wei; Qiu, Ping; Wang, Luquan; Doll, Ronald J.; English, Jessie M.

2004-01-01

MAPK (mitogen-activated protein kinase) pathways constitute major regulators of cellular transcriptional programmes. We analysed the ERK1,2 (extracellular-signal-regulated kinase 1,2) transcriptome in a non-transformed MEC (mammary epithelial cell) line, MCF-12A, utilizing rAd MEK1EE, a recombinant adenovirus encoding constitutively active MEK1 (MAPK/ERK kinase 1). rAd MEK1EE infection induced morphological changes and DNA synthesis which were inhibited by the MEK1,2 inhibitor PD184352. Hierarchical clustering of data derived from seven time points over 24 h identified 430 and 305 co-ordinately up-regulated and down-regulated genes respectively. c-Myc binding sites were identified in the promoters of most of these up-regulated genes. A total of 46 candidate effectors of the Raf/MEK/ERK1,2 pathway in MECs were identified by comparing our dataset with previously reported Raf-1-regulated genes. These analyses led to the identification of a suite of growth factors co-ordinately induced by MEK1EE, including multiple ErbB ligands, vascular endothelial growth factor and PHRP (parathyroid hormone-related protein). PHRP is the primary mediator of humoral hypercalcaemia of malignancy, and has been implicated in metastasis to bone. We demonstrate that PHRP is secreted by MEK1EE-expressing cells. This secretion is inhibited by PD184352, but not by ErbB inhibitors. Our results suggest that, in addition to anti-proliferative properties, MEK1,2 inhibitors may be anti-angiogenic and possess therapeutic utility in the treatment of PHRP-positive tumours. PMID:15109307

Regulation of fibroblast growth factor receptor signalling and trafficking by Src and Eps8.

PubMed

Auciello, Giulio; Cunningham, Debbie L; Tatar, Tulin; Heath, John K; Rappoport, Joshua Z

2013-01-15

Fibroblast growth factor receptors (FGFRs) mediate a wide spectrum of cellular responses that are crucial for development and wound healing. However, aberrant FGFR activity leads to cancer. Activated growth factor receptors undergo stimulated endocytosis, but can continue to signal along the endocytic pathway. Endocytic trafficking controls the duration and intensity of signalling, and growth factor receptor signalling can lead to modifications of trafficking pathways. We have developed live-cell imaging methods for studying FGFR dynamics to investigate mechanisms that coordinate the interplay between receptor trafficking and signal transduction. Activated FGFR enters the cell following recruitment to pre-formed clathrin-coated pits (CCPs). However, FGFR activation stimulates clathrin-mediated endocytosis; FGF treatment increases the number of CCPs, including those undergoing endocytosis, and this effect is mediated by Src and its phosphorylation target Eps8. Eps8 interacts with the clathrin-mediated endocytosis machinery and depletion of Eps8 inhibits FGFR trafficking and immediate Erk signalling. Once internalized, FGFR passes through peripheral early endosomes en route to recycling and degredative compartments, through an Src- and Eps8-dependent mechanism. Thus Eps8 functions as a key coordinator in the interplay between FGFR signalling and trafficking. This work provides the first detailed mechanistic analysis of growth factor receptor clustering at the cell surface through signal transduction and endocytic trafficking. As we have characterised the Src target Eps8 as a key regulator of FGFR signalling and trafficking, and identified the early endocytic system as the site of Eps8-mediated effects, this work provides novel mechanistic insight into the reciprocal regulation of growth factor receptor signalling and trafficking.

The tangled web of non-canonical Wnt signalling in neural migration.

PubMed

Clark, Charlotte E J; Nourse, C Cathrin; Cooper, Helen M

2012-01-01

In all multicellular animals, successful embryogenesis is dependent on the ability of cells to detect the status of the local environment and respond appropriately. The nature of the extracellular environment is communicated to the intracellular compartment by ligand/receptor interactions at the cell surface. The Wnt canonical and non-canonical signalling pathways are found in the most primitive metazoans, and they play an essential role in the most fundamental developmental processes in all multicellular organisms. Vertebrates have expanded the number of Wnts and Frizzled receptors and have additionally evolved novel Wnt receptor families (Ryk, Ror). The multiplicity of potential interactions between Wnts, their receptors and downstream effectors has exponentially increased the complexity of the signal transduction network. Signalling through each of the Wnt pathways, as well as crosstalk between them, plays a critical role in the establishment of the complex architecture of the vertebrate central nervous system. In this review, we explore the signalling networks triggered by non-canonical Wnt/receptor interactions, focussing on the emerging roles of the non-conventional Wnt receptors Ryk and Ror. We describe the role of these pathways in neural tube formation and axon guidance where Wnt signalling controls tissue polarity, coordinated cell migration and axon guidance via remodelling of the cytoskeleton. Copyright © 2012 S. Karger AG, Basel.

Isothiocyanates: mechanism of cancer chemopreventive action.

PubMed

Thornalley, Paul J

2002-04-01

Dietary and synthetic isothiocyanates have cancer chemopreventive activity. Dietary isothiocyanates are formed from glucosinolate precursors of ingested green vegetables. Isothiocyanates are absorbed across intestinal cell membranes by passive diffusion and bind reversibly to plasma protein thiols by thiocarbamoylation. Free isothiocyanate enters cells and is converted to the glutathione conjugate by glutathione S-transferases (GSTs). The glutathione conjugate is exported from cells by multidrug resistance proteins (MRPs), and metabolized in the mercapturic acid pathway to the corresponding mercapturic acid. The isothiocyanate is reformed by fragmentation of mercapturic acid pathway metabolites; it is inactivated by slow hydrolysis to the corresponding amine that is inactive in chemoprevention. Depletion of cellular glutathione and protein thiocarbamoylation activates signal transduction for cancer chemoprevention. Isothiocyanates inhibited and inactivated cytochrome P450 isoforms. They induced increased expression of GST, NADPH: quinone oxidoreductase, aldo-keto reductase and gamma-glutamylcysteine synthetase. These responses were coordinated at the transcription level by nuclear factor-erythroid 2 p45-related factor-2 acting through the antioxidant/electrophile enhancer response element and stimulated by the mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase-1 and c-Jun N-terminal kinase-1 (JNK1) pathway. Isothiocyanates also induced apoptosis of pre-cancerous cells and tumor cells activated by caspase-8 and potentiated by JNK1. The chemopreventive activity of isothiocyanates is influenced by the isothiocyanate bioavailability-as is toxicity, GST polymorphism, protein thiocarbamoylation and probably also by MRP expression. These features of isothiocyanate metabolism and chemoprevention deserve further investigation.

Coordination of Satellite Cell Activation and Self-Renewal by Par-Complex-Dependent Asymmetric Activation of p38α/β MAPK

PubMed Central

Troy, Andrew; Cadwallader, Adam B.; Fedorov, Yuri; Tyner, Kristina; Tanaka, Kathleen Kelly; Olwin, Bradley B.

2014-01-01

SUMMARY In response to muscle injury, satellite cells activate the p38α/β MAPK pathway to exit quiescence, then proliferate, repair skeletal muscle, and self-renew, replenishing the quiescent satellite cell pool. Although satellite cells are capable of asymmetric division, the mechanisms regulating satellite cell self-renewal are not understood. We found that satellite cells, once activated, enter the cell cycle and a subset undergoes asymmetric division, renewing the satellite cell pool. Asymmetric localization of the Par complex activates p38α/β MAPK in only one daughter cell, inducing MyoD, which permits cell cycle entry and generates a proliferating myoblast. The absence of p38α/β MAPK signaling in the other daughter cell prevents MyoD induction, renewing the quiescent satellite cell. Thus, satellite cells employ a mechanism to generate distinct daughter cells, coupling the Par complex and p38α/β MAPK signaling to link the response to muscle injury with satellite cell self-renewal. PMID:23040480

Myc requires RhoA/SRF to reprogram glutamine metabolism.

PubMed

Haikala, Heidi M; Marques, Elsa; Turunen, Mikko; Klefström, Juha

2018-05-04

RhoA regulates actin cytoskeleton but recent evidence suggest a role for this conserved Rho GTPase also in other cellular processes, including transcriptional control of cell proliferation and survival. Interestingy, loss of RhoA is synthetic lethal with oncogenic Myc, a master transcription factor that turns on anabolic metabolism to promote cell growth in many cancers. We show evidence indicating that the synthetic lethal interaction between RhoA loss and Myc arises from deficiency in glutamine utilization, resulting from impaired co-regulation of glutaminase expression and anaplerosis by Myc and RhoA - serum response factor (SRF) pathway. The results suggest metabolic coordination between Myc and RhoA/SRF in sustaining cancer cell viability and indicate RhoA/SRF as a potential vulnerability in cancer cells for therapeutic targeting.

Cell-derived microparticles: new targets in the therapeutic management of disease.

PubMed

Roseblade, Ariane; Luk, Frederick; Rawling, Tristan; Ung, Alison; Grau, Georges E R; Bebawy, Mary

2013-01-01

Intercellular communication is essential to maintain vital physiological activities and to regulate the organism's phenotype. There are a number of ways in which cells communicate with one another. This can occur via autocrine signaling, endocrine signaling or by the transfer of molecular mediators across gap junctions. More recently communication via microvesicular shedding has gained important recognition as a significant pathway by which cells can coordinate the spread and dominance of selective traits within a population. Through this communication apparatus, cells can now acquire and secure a survival advantage, particularly in the context of malignant disease. This review aims to highlight some of the functions and implications of microparticles in physiology of various disease states, and present a novel therapeutic strategy through the regulation of microparticle production.

Expression of the transforming growth factor alpha protooncogene in differentiating human promyelocytic leukemia (HL-60) cells.

PubMed

Walz, T M; Malm, C; Wasteson, A

1993-01-01

The process of myeloid differentiation in human promyelocytic leukemia cells (HL-60) is accompanied by the coordinate expression of numerous protooncogenes. To investigate the expression of transforming growth factor alpha (TGF-alpha) in myeloid differentiation, HL-60 cells were induced to differentiate into granulocytes with 1.25% dimethyl sulfoxide, 0.2 microM all-trans retinoic acid, or 500 microM N6,O2-dibutyryladenosine-3'5'-cyclic monophosphate or differentiated along the monocyte/macrophage pathway with 0.1 microM phorbol-12-myristate-13-acetate. Using Northern blot analyses, TGF-alpha transcripts were detected within 24 h of treatment in cells differentiating toward granulocytes; maximal levels of gene expression were reached after 3 days or later and remained essentially constant throughout the observation period. These cells released TGF-alpha protein, as demonstrated by analysis of the incubation medium. In contrast, no TGF-alpha RNA or protein was detectable in HL-60 cell cultures when induced with phorbol-12-myristate-13-acetate. Epidermal growth factor receptor transcripts could not be detected either in undifferentiated or in differentiated HL-60 cells; therefore it appears as if an autocrine loop involving TGF-alpha in HL-60 cells is unlikely. In conclusion, the results demonstrate, for the first time, the expression of TGF-alpha in human granulocyte precursor cells. Our findings may indicate novel regulatory pathways in hematopoiesis.

Feedback Activation of Basic Fibroblast Growth Factor Signaling via the Wnt/β-Catenin Pathway in Skin Fibroblasts

PubMed Central

Wang, Xu; Zhu, Yuting; Sun, Congcong; Wang, Tao; Shen, Yingjie; Cai, Wanhui; Sun, Jia; Chi, Lisha; Wang, Haijun; Song, Na; Niu, Chao; Shen, Jiayi; Cong, Weitao; Zhu, Zhongxin; Xuan, Yuanhu; Li, Xiaokun; Jin, Litai

2017-01-01

Skin wound healing is a complex process requiring the coordinated behavior of many cell types, especially in the proliferation and migration of fibroblasts. Basic fibroblast growth factor (bFGF) is a member of the FGF family that promotes fibroblast migration, but the underlying molecular mechanism remains elusive. The present RNA sequencing study showed that the expression levels of several canonical Wnt pathway genes, including Wnt2b, Wnt3, Wnt11, T-cell factor 7 (TCF7), and Frizzled 8 (FZD8) were modified by bFGF stimulation in fibroblasts. Enzyme-linked immunosorbent assay (ELISA) analysis also showed that Wnt pathway was activated under bFGF treatment. Furthermore, treatment of fibroblasts with lithium chloride or IWR-1, an inducer and inhibitor of the Wnt signaling pathway, respectively, promoted and inhibited cell migration. Also, levels of cytosolic glycogen synthase kinase 3 beta phosphorylated at serine9 (pGSK3β Ser9) and nuclear β-catenin were increased upon exposure to bFGF. Molecular and biochemical assays indicated that phosphoinositide 3-kinase (PI3K) signaling activated the GSK3β/β-catenin/Wnt signaling pathway via activation of c-Jun N-terminal kinase (JNK), suggesting that PI3K and JNK act at the upstream of β-catenin. In contrast, knock-down of β-catenin delayed fibroblast cell migration even under bFGF stimulation. RNA sequencing analysis of β-catenin knock-down fibroblasts demonstrated that β-catenin positively regulated the transcription of bFGF and FGF21. Moreover, FGF21 treatment activated AKT and JNK, and accelerated fibroblast migration to a similar extent as bFGF does. In addition, ELISA analysis demonstrated that both of bFGF and FGF21 were auto secretion factor and be regulated by Wnt pathway stimulators. Taken together, our analyses define a feedback regulatory loop between bFGF (FGF21) and Wnt signaling acting through β-catenin in skin fibroblasts. PMID:28217097

The clearance of dying cells: table for two

PubMed Central

Green, D R; Oguin, T H; Martinez, J

2016-01-01

Phagocytic cells of the immune system must constantly survey for, recognize, and efficiently clear the billions of cellular corpses that arise as a result of development, stress, infection, or normal homeostasis. This process, termed efferocytosis, is critical for the prevention of autoimmune and inflammatory disorders, and persistence of dead cells in tissue is characteristic of many human autoimmune diseases, notably systemic lupus erythematosus. The most notable characteristic of the efferocytosis of apoptotic cells is its ‘immunologically silent' response. Although the mechanisms by which phagocytes facilitate engulfment of dead cells has been a well-studied area, the pathways that coordinate to process the ingested corpse and direct the subsequent immune response is an area of growing interest. The recently described pathway of LC3 (microtubule-associated protein 1A/1B-light chain 3)-associated phagocytosis (LAP) has shed some light on this issue. LAP is triggered when an extracellular particle, such as a dead cell, engages an extracellular receptor during phagocytosis, induces the translocation of autophagy machinery, and ultimately LC3 to the cargo-containing phagosome, termed the LAPosome. In this review, we will examine efferocytosis and the impact of LAP on efferocytosis, allowing us to reimagine the impact of the autophagy machinery on innate host defense mechanisms. PMID:26990661

Live-cell imaging for the assessment of the dynamics of autophagosome formation: focus on early steps.

PubMed

Karanasios, Eleftherios; Ktistakis, Nicholas T

2015-03-01

Autophagy is a cytosolic degradative pathway, which through a series of complicated membrane rearrangements leads to the formation of a unique double membrane vesicle, the autophagosome. The use of fluorescent proteins has allowed visualizing the autophagosome formation in live cells and in real time, almost 40 years after electron microscopy studies observed these structures for the first time. In the last decade, live-cell imaging has been extensively used to study the dynamics of autophagosome formation in cultured mammalian cells. Hereby we will discuss how the live-cell imaging studies have tried to settle the debate about the origin of the autophagosome membrane and how they have described the way different autophagy proteins coordinate in space and time in order to drive autophagosome formation. Copyright © 2014 Elsevier Inc. All rights reserved.

Emerging regulatory paradigms in glutathione metabolism

PubMed Central

Liu, Yilin; Hyde, Annastasia S.; Simpson, Melanie A.; Barycki, Joseph J.

2015-01-01

One of the hallmarks of cancer is the ability to generate and withstand unusual levels of oxidative stress. In part, this property of tumor cells is conferred by elevation of the cellular redox buffer glutathione. Though enzymes of the glutathione synthesis and salvage pathways have been characterized for several decades, we still lack a comprehensive understanding of their independent and coordinate regulatory mechanisms. Recent studies have further revealed that overall central metabolic pathways are frequently altered in various tumor types, resulting in significant increases in biosynthetic capacity, and feeding into glutathione synthesis. In this review, we will discuss the enzymes and pathways affecting glutathione flux in cancer, and summarize current models for regulating cellular glutathione through both de novo synthesis and efficient salvage. In addition, we examine the integration of glutathione metabolism with other altered fates of intermediary metabolites, and highlight remaining questions about molecular details of the accepted regulatory modes. PMID:24974179

Control systems and coordination protocols of the secretory pathway.

PubMed

Luini, Alberto; Mavelli, Gabriella; Jung, Juan; Cancino, Jorge

2014-01-01

Like other cellular modules, the secretory pathway and the Golgi complex are likely to be supervised by control systems that support homeostasis and optimal functionality under all conditions, including external and internal perturbations. Moreover, the secretory apparatus must be functionally connected with other cellular modules, such as energy metabolism and protein degradation, via specific rules of interaction, or "coordination protocols". These regulatory devices are of fundamental importance for optimal function; however, they are generally "hidden" at steady state. The molecular components and the architecture of the control systems and coordination protocols of the secretory pathway are beginning to emerge through studies based on the use of controlled transport-specific perturbations aimed specifically at the detection and analysis of these internal regulatory devices.

Histone methyltransferase SETDB1 maintains survival of mouse spermatogonial stem/progenitor cells via PTEN/AKT/FOXO1 pathway.

PubMed

Liu, Tiantian; Chen, Xiaoxu; Li, Tianjiao; Li, Xueliang; Lyu, Yinghua; Fan, Xiaoteng; Zhang, Pengfei; Zeng, Wenxian

2017-10-01

Spermatogonial stem cells (SSCs) possess the capacity of self-renewal and differentiation, which are the basis of spermatogenesis. In maintenance of SSC homeostasis, intrinsic/extrinsic factors and various signaling pathways tightly control the fate of SSCs. Methyltransferase SETDB1 (Set domain, bifurcated 1) catalyzes histone H3 lysine 9 (H3K9) trimethylation and represses gene expression. SETDB1 is required for maintaining the survival of spermatogonial stem cells in mice. However, the underlying molecular mechanism remains unclear. In the present study, we found that Setdb1 regulates PTEN/AKT/FOXO1 pathway to inhibit SSC apoptosis. Co-immunoprecipitation and reporter gene assay revealed that SETDB1 interacted and coordinated with AKT to regulate FOXO1 activity and expression of the downstream target genes Bim and Puma. Among the SETDB1-bound genes, the H3K9me3 levels on the promoter regions of Bim and Pten decreased in Setdb1-KD group; in contrast, H3K9me3 status on promoters of Bax and Puma remained unchanged. Therefore, SETDB1 was responsible for regulating the transcription activity of genes in the apoptotic pathway at least in part through modulating H3K9me3. This study replenishes the research on the epigenetic regulation of SSC survival, and provides a new insight for the future study of epigenetic regulation of spermatogenesis. Copyright © 2017 Elsevier B.V. All rights reserved.

Mitochondrial dynamics and optical conformation changes in DsRed as studied by Fourier imaging correlation spectroscopy

NASA Astrophysics Data System (ADS)

Senning, Eric Nicolas

Novel experiments that probe the dynamics of intracellular species, including the center-of-mass displacements and internal conformational transitions of biological macromolecules, have the potential to reveal the complex biochemical mechanisms operating within the cell. This work presents the implementation and development of Fourier imaging correlation spectroscopy (FICS), a phase-selective approach to fluorescence spectroscopy that measures the collective coordinate fluctuations of fluorescently labeled microscopic particles. In FICS experiments, a spatially modulated optical grating excites a fluorescently labeled sample. Phase-synchronous detection of the fluorescence, with respect to the phase of the exciting optical grating, can be used to monitor the fluctuations of partially averaged spatial coordinates. These data are then analyzed by two-point and four-point time correlation functions to provide a statistically meaningful understanding of the dynamics under observation. FICS represents a unique route to elevate signal levels, while acquiring detailed information about molecular coordinate trajectories. Mitochondria of mammalian cells are known to associate with cytoskeletal proteins, and their motions are affected by the stability of microtubules and microfilaments. Within the cell it is possible to fluorescently label the mitochondria and study its dynamic behavior with FICS. The dynamics of S. cerevisiae yeast mitochondria are characterized at four discrete length scales (ranging from 0.6--1.19 mum) and provide detailed information about the influence of specific cytoskeletal elements. Using the microtubule and microfilament destabilizing agents, Nocodazole and Latrunculin A, it is determined that microfilaments are required for normal yeast mitochondrial motion while microtubules have no effect. Experiments with specific actin mutants revealed that actin is responsible for enhanced mobility on length scales greater than 0.6 mum. The versatility of FICS expands when individual molecules are labeled with fluorescent chromophores. In recent experiments on the tetrameric fluorescent protein DsRed, polarization-modulated FICS (PM-FICS) is demonstrated to separate conformational dynamics from molecular translational dynamics. The optical switching pathways of DsRed, a tetrameric complex of fluorescent protein subunits, are examined. An analysis of PM-FICS coordinate trajectories, in terms of 2D spectra and joint probability distributions, provides detailed information about the transition pathways between distinct dipole-coupled DsRed conformations. This dissertation includes co-authored and previously published material.

Yersinia vs. host Immunity: how a pathogen evades or triggers a protective response

PubMed Central

Chung, Lawton K.; Bliska, James B.

2015-01-01

The human pathogenic Yersinia species cause diseases that represent a significant source of morbidity and mortality. Despite this, specific mechanisms underlying Yersinia pathogenesis and protective host responses remain poorly understood. Recent studies have shown that Yersinia disrupt cell death pathways, perturb inflammatory processes and exploit immune cells to promote disease. The ensuing host responses following Yersinia infection include coordination of innate and adaptive immune responses in an attempt to control bacterial replication. Here, we highlight current advances in our understanding of the interactions between the pathogenic yersiniae and host cells, as well as the protective host responses mobilized to counteract these pathogens. Together, these studies enhance our understanding of Yersinia pathogenesis and highlight the ongoing battle between host and microbe. PMID:26638030

Genetic and Cellular Mechanisms Regulating Anterior Foregut and Esophageal Development

PubMed Central

Jacobs, Ian J.; Ku, Wei-Yao; Que, Jianwen

2012-01-01

Separation of the single anterior foregut tube into the esophagus and trachea involves cell proliferation and differentiation, as well as dynamic changes in cell-cell adhesion and migration. These biological processes are regulated and coordinated at multiple levels through the interplay of the epithelium and mesenchyme. Genetic studies and in vitro modeling have shed light on relevant regulatory networks that include a number of transcription factors and signaling pathways. These signaling molecules exhibit unique expression patterns and play specific functions in their respective territories before the separation process occurs. Disruption of regulatory networks inevitably leads to defective separation and malformation of the trachea and esophagus and results in the formation of a relatively common birth defect, esophageal atresia with or without tracheoesophageal fistula (EA/TEF). Significantly, some of the signaling pathways and transcription factors involved in anterior foregut separation continue to play important roles in the morphogenesis of the individual organs. In this review, we will focus on new findings related to these different developmental processes and discuss them in the context of developmental disorders (or birth defects) commonly seen in clinics. PMID:22750256

Signaling pathways coordinating the alkaline pH response confer resistance to the hevein-type plant antimicrobial peptide Pn-AMP1 in Saccharomyces cerevisiae.

PubMed

Kwon, Youngho; Chiang, Jennifer; Tran, Grant; Giaever, Guri; Nislow, Corey; Hahn, Bum-Soo; Kwak, Youn-Sig; Koo, Ja-Choon

2016-12-01

Genome-wide screening of Saccharomyces cerevisiae revealed that signaling pathways related to the alkaline pH stress contribute to resistance to plant antimicrobial peptide, Pn-AMP1. Plant antimicrobial peptides (AMPs) are considered to be promising candidates for controlling phytopathogens. Pn-AMP1 is a hevein-type plant AMP that shows potent and broad-spectrum antifungal activity. Genome-wide chemogenomic screening was performed using heterozygous and homozygous diploid deletion pools of Saccharomyces cerevisiae as a chemogenetic model system to identify genes whose deletion conferred enhanced sensitivity to Pn-AMP1. This assay identified 44 deletion strains with fitness defects in the presence of Pn-AMP1. Strong fitness defects were observed in strains with deletions of genes encoding components of several pathways and complex known to participate in the adaptive response to alkaline pH stress, including the cell wall integrity (CWI), calcineurin/Crz1, Rim101, SNF1 pathways and endosomal sorting complex required for transport (ESCRT complex). Gene ontology (GO) enrichment analysis of these genes revealed that the most highly overrepresented GO term was "cellular response to alkaline pH". We found that 32 of the 44 deletion strains tested (72 %) showed significant growth defects compared with their wild type at alkaline pH. Furthermore, 9 deletion strains (20 %) exhibited enhanced sensitivity to Pn-AMP1 at ambient pH compared to acidic pH. Although several hundred plant AMPs have been reported, their modes of action remain largely uncharacterized. This study demonstrates that the signaling pathways that coordinate the adaptive response to alkaline pH also confer resistance to a hevein-type plant AMP in S. cerevisiae. Our findings have broad implications for the design of novel and potent antifungal agents.

Transcriptional Regulation of Pattern-Triggered Immunity in Plants.

PubMed

Li, Bo; Meng, Xiangzong; Shan, Libo; He, Ping

2016-05-11

Perception of microbe-associated molecular patterns (MAMPs) by cell-surface-resident pattern recognition receptors (PRRs) induces rapid, robust, and selective transcriptional reprogramming, which is central for launching effective pattern-triggered immunity (PTI) in plants. Signal relay from PRR complexes to the nuclear transcriptional machinery via intracellular kinase cascades rapidly activates primary immune response genes. The coordinated action of gene-specific transcription factors and the general transcriptional machinery contribute to the selectivity of immune gene activation. In addition, PRR complexes and signaling components are often transcriptionally upregulated upon MAMP perception to ensure the robustness and sustainability of PTI outputs. In this review, we discuss recent advances in deciphering the signaling pathways and regulatory mechanisms that coordinately lead to timely and accurate MAMP-induced gene expression in plants. Copyright © 2016 Elsevier Inc. All rights reserved.

Application of Laser Micro-irradiation for Examination of Single and Double Strand Break Repair in Mammalian Cells.

PubMed

Holton, Nathaniel W; Andrews, Joel F; Gassman, Natalie R

2017-09-05

Highly coordinated DNA repair pathways exist to detect, excise and replace damaged DNA bases, and coordinate repair of DNA strand breaks. While molecular biology techniques have clarified structure, enzymatic functions, and kinetics of repair proteins, there is still a need to understand how repair is coordinated within the nucleus. Laser micro-irradiation offers a powerful tool for inducing DNA damage and monitoring the recruitment of repair proteins. Induction of DNA damage by laser micro-irradiation can occur with a range of wavelengths, and users can reliably induce single strand breaks, base lesions and double strand breaks with a range of doses. Here, laser micro-irradiation is used to examine repair of single and double strand breaks induced by two common confocal laser wavelengths, 355 nm and 405 nm. Further, proper characterization of the applied laser dose for inducing specific damage mixtures is described, so users can reproducibly perform laser micro-irradiation data acquisition and analysis.

Parallel odor processing by mitral and middle tufted cells in the olfactory bulb.

PubMed

Cavarretta, Francesco; Burton, Shawn D; Igarashi, Kei M; Shepherd, Gordon M; Hines, Michael L; Migliore, Michele

2018-05-16

The olfactory bulb (OB) transforms sensory input into spatially and temporally organized patterns of activity in principal mitral (MC) and middle tufted (mTC) cells. Thus far, the mechanisms underlying odor representations in the OB have been mainly investigated in MCs. However, experimental findings suggest that MC and mTC may encode parallel and complementary odor representations. We have analyzed the functional roles of these pathways by using a morphologically and physiologically realistic three-dimensional model to explore the MC and mTC microcircuits in the glomerular layer and deeper plexiform layer. The model makes several predictions. MCs and mTCs are controlled by similar computations in the glomerular layer but are differentially modulated in deeper layers. The intrinsic properties of mTCs promote their synchronization through a common granule cell input. Finally, the MC and mTC pathways can be coordinated through the deep short-axon cells in providing input to the olfactory cortex. The results suggest how these mechanisms can dynamically select the functional network connectivity to create the overall output of the OB and promote the dynamic synchronization of glomerular units for any given odor stimulus.

Putting one step before the other: distinct activation pathways for Cdk1 and Cdk2 bring order to the mammalian cell cycle

PubMed Central

Merrick, Karl A.; Fisher, Robert P.

2010-01-01

Eukaryotic cell division is controlled by the activity of cyclin-dependent kinases (CDKs). Cdk1 and Cdk2, which function at different stages of the mammalian cell cycle, both require cyclin-binding and phosphorylation of the activation (T-) loop for full activity, but differ with respect to the order in which the two steps occur in vivo. To form stable complexes with either of its partners—cyclins A and B—Cdk1 must be phosphorylated on its T-loop, but that phosphorylation in turn depends on the presence of cyclin. Cdk2 can follow a kinetically distinct path to activation in which T-loop phosphorylation precedes cyclin-binding, and thereby out-compete the more abundant Cdk1 for limiting amounts of cyclin A. Mathematical modeling suggests this could be a principal basis for the temporal ordering of CDK activation during S phase, which may dictate the sequence in which replication origins fire. Still to be determined are how: 1) the activation machinery discriminates between closely related CDKs, and 2) coordination of the cell cycle is affected when this mechanism of pathway insulation breaks down. PMID:20139727

Memory and modularity in cell-fate decision making

NASA Astrophysics Data System (ADS)

Norman, Thomas M.; Lord, Nathan D.; Paulsson, Johan; Losick, Richard

2013-11-01

Genetically identical cells sharing an environment can display markedly different phenotypes. It is often unclear how much of this variation derives from chance, external signals, or attempts by individual cells to exert autonomous phenotypic programs. By observing thousands of cells for hundreds of consecutive generations under constant conditions, we dissect the stochastic decision between a solitary, motile state and a chained, sessile state in Bacillus subtilis. We show that the motile state is `memoryless', exhibiting no autonomous control over the time spent in the state. In contrast, the time spent as connected chains of cells is tightly controlled, enforcing coordination among related cells in the multicellular state. We show that the three-protein regulatory circuit governing the decision is modular, as initiation and maintenance of chaining are genetically separable functions. As stimulation of the same initiating pathway triggers biofilm formation, we argue that autonomous timing allows a trial commitment to multicellularity that external signals could extend.

Middle age onset short-term intermittent fasting dietary restriction prevents brain function impairments in male Wistar rats.

PubMed

Singh, Rumani; Manchanda, Shaffi; Kaur, Taranjeet; Kumar, Sushil; Lakhanpal, Dinesh; Lakhman, Sukhwinder S; Kaur, Gurcharan

2015-12-01

Intermittent fasting dietary restriction (IF-DR) is recently reported to be an effective intervention to retard age associated disease load and to promote healthy aging. Since sustaining long term caloric restriction regimen is not practically feasible in humans, so use of alternate approach such as late onset short term IF-DR regimen which is reported to trigger similar biological pathways is gaining scientific interest. The current study was designed to investigate the effect of IF-DR regimen implemented for 12 weeks in middle age rats on their motor coordination skills and protein and DNA damage in different brain regions. Further, the effect of IF-DR regimen was also studied on expression of energy regulators, cell survival pathways and synaptic plasticity marker proteins. Our data demonstrate that there was an improvement in motor coordination and learning response with decline in protein oxidative damage and recovery in expression of energy regulating neuropeptides. We further observed significant downregulation in nuclear factor kappa B (NF-κB) and cytochrome c (Cyt c) levels and moderate upregulation of mortalin and synaptophysin expression. The present data may provide an insight on how a modest level of short term IF-DR, imposed in middle age, can slow down or prevent the age-associated impairment of brain functions and promote healthy aging by involving multiple regulatory pathways aimed at maintaining energy homeostasis.

Control of Organ Growth by Patterning and Hippo Signaling in Drosophila

PubMed Central

Irvine, Kenneth D.; Harvey, Kieran F.

2015-01-01

Control of organ size is of fundamental importance and is controlled by genetic, environmental, and mechanical factors. Studies in many species have pointed to the existence of both organ-extrinsic and -intrinsic size-control mechanisms, which ultimately must coordinate to regulate organ size. Here, we discuss organ size control by organ patterning and the Hippo pathway, which both act in an organ-intrinsic fashion. The influence of morphogens and other patterning molecules couples growth and patterning, whereas emerging evidence suggests that the Hippo pathway controls growth in response to mechanical stimuli and signals emanating from cell–cell interactions. Several points of cross talk have been reported between signaling pathways that control organ patterning and the Hippo pathway, both at the level of membrane receptors and transcriptional regulators. However, despite substantial progress in the past decade, key questions in the growth-control field remain, including precisely how and when organ patterning and the Hippo pathway communicate to control size, and whether these communication mechanisms are organ specific or general. In addition, elucidating mechanisms by which organ-intrinsic cues, such as patterning factors and the Hippo pathway, interface with extrinsic cues, such as hormones to control organ size, remain unresolved. PMID:26032720

Cytoplasmic proliferating cell nuclear antigen connects glycolysis and cell survival in acute myeloid leukemia.

PubMed

Ohayon, Delphine; De Chiara, Alessia; Chapuis, Nicolas; Candalh, Céline; Mocek, Julie; Ribeil, Jean-Antoine; Haddaoui, Lamya; Ifrah, Norbert; Hermine, Olivier; Bouillaud, Frédéric; Frachet, Philippe; Bouscary, Didier; Witko-Sarsat, Véronique

2016-10-19

Cytosolic proliferating cell nuclear antigen (PCNA), a scaffolding protein involved in DNA replication, has been described as a key element in survival of mature neutrophil granulocytes, which are non-proliferating cells. Herein, we demonstrated an active export of PCNA involved in cell survival and chemotherapy resistance. Notably, daunorubicin-resistant HL-60 cells (HL-60R) have a prominent cytosolic PCNA localization due to increased nuclear export compared to daunorubicin-sensitive HL-60 cells (HL-60S). By interacting with nicotinamide phosphoribosyltransferase (NAMPT), a protein involved in NAD biosynthesis, PCNA coordinates glycolysis and survival, especially in HL-60R cells. These cells showed a dramatic increase in intracellular NAD+ concentration as well as glycolysis including increased expression and activity of hexokinase 1 and increased lactate production. Furthermore, this functional activity of cytoplasmic PCNA was also demonstrated in patients with acute myeloid leukemia (AML). Our data uncover a novel pathway of nuclear export of PCNA that drives cell survival by increasing metabolism flux.

Cytoplasmic proliferating cell nuclear antigen connects glycolysis and cell survival in acute myeloid leukemia

PubMed Central

Ohayon, Delphine; De Chiara, Alessia; Chapuis, Nicolas; Candalh, Céline; Mocek, Julie; Ribeil, Jean-Antoine; Haddaoui, Lamya; Ifrah, Norbert; Hermine, Olivier; Bouillaud, Frédéric; Frachet, Philippe; Bouscary, Didier; Witko-Sarsat, Véronique

2016-01-01

Cytosolic proliferating cell nuclear antigen (PCNA), a scaffolding protein involved in DNA replication, has been described as a key element in survival of mature neutrophil granulocytes, which are non-proliferating cells. Herein, we demonstrated an active export of PCNA involved in cell survival and chemotherapy resistance. Notably, daunorubicin-resistant HL-60 cells (HL-60R) have a prominent cytosolic PCNA localization due to increased nuclear export compared to daunorubicin-sensitive HL-60 cells (HL-60S). By interacting with nicotinamide phosphoribosyltransferase (NAMPT), a protein involved in NAD biosynthesis, PCNA coordinates glycolysis and survival, especially in HL-60R cells. These cells showed a dramatic increase in intracellular NAD+ concentration as well as glycolysis including increased expression and activity of hexokinase 1 and increased lactate production. Furthermore, this functional activity of cytoplasmic PCNA was also demonstrated in patients with acute myeloid leukemia (AML). Our data uncover a novel pathway of nuclear export of PCNA that drives cell survival by increasing metabolism flux. PMID:27759041

Interlocked positive and negative feedback network motifs regulate β-catenin activity in the adherens junction pathway

PubMed Central

Klinke, David J.; Horvath, Nicholas; Cuppett, Vanessa; Wu, Yueting; Deng, Wentao; Kanj, Rania

2015-01-01

The integrity of epithelial tissue architecture is maintained through adherens junctions that are created through extracellular homotypic protein–protein interactions between cadherin molecules. Cadherins also provide an intracellular scaffold for the formation of a multiprotein complex that contains signaling proteins, including β-catenin. Environmental factors and controlled tissue reorganization disrupt adherens junctions by cleaving the extracellular binding domain and initiating a series of transcriptional events that aim to restore tissue homeostasis. However, it remains unclear how alterations in cell adhesion coordinate transcriptional events, including those mediated by β-catenin in this pathway. Here were used quantitative single-cell and population-level in vitro assays to quantify the endogenous pathway dynamics after the proteolytic disruption of the adherens junctions. Using prior knowledge of isolated elements of the overall network, we interpreted these data using in silico model-based inference to identify the topology of the regulatory network. Collectively the data suggest that the regulatory network contains interlocked network motifs consisting of a positive feedback loop, which is used to restore the integrity of adherens junctions, and a negative feedback loop, which is used to limit β-catenin–induced gene expression. PMID:26224311

Excitable toxin-antitoxin modules coordinated through intracellular bottlenecks

NASA Astrophysics Data System (ADS)

Mather, William

Chronic infections and pathogenic biofilms present a serious threat to the health of humans by decreasing life expectancy and quality. The resilience of these microbial communities has been attributed to the spontaneous formation of persister cells, which constitute a small fraction of the population capable of surviving a wide range of environmental stressors. Gating of bacterial persistence has recently been linked to toxin-antitoxin (TA) modules, which are operons with an evolutionarily conserved motif that includes a toxin that halts cell growth and a corresponding antitoxin that neutralizes the toxin. While many such modules have been identified and studied in a wide range of organisms, little consideration of the interactions between multiple modules within a single host has been made. Moreover, the multitude of different antitoxin species are degraded by a relatively small number of proteolytic pathways, strongly suggesting competition between antitoxins for degradation machinery, i.e. queueing coupling. Here we present a theoretical understanding of the dynamics of multiple TA modules that are coupled through either proteolytic queueing, a toxic effect on cell growth rate, or both. We conclude that indirect queueing coordination between multiple TA modules may be central to controlling bacterial persistence. NSF Award Number MCB-1330180.

Coordinated regulation of neuronal mRNA steady-state levels through developmentally controlled intron retention

PubMed Central

Yap, Karen; Lim, Zhao Qin; Khandelia, Piyush; Friedman, Brad; Makeyev, Eugene V.

2012-01-01

Differentiated cells acquire unique structural and functional traits through coordinated expression of lineage-specific genes. An extensive battery of genes encoding components of the synaptic transmission machinery and specialized cytoskeletal proteins is activated during neurogenesis, but the underlying regulation is not well understood. Here we show that genes encoding critical presynaptic proteins are transcribed at a detectable level in both neurons and nonneuronal cells. However, in nonneuronal cells, the splicing of 3′-terminal introns within these genes is repressed by the polypyrimidine tract-binding protein (Ptbp1). This inhibits the export of incompletely spliced mRNAs to the cytoplasm and triggers their nuclear degradation. Clearance of these intron-containing transcripts occurs independently of the nonsense-mediated decay (NMD) pathway but requires components of the nuclear RNA surveillance machinery, including the nuclear pore-associated protein Tpr and the exosome complex. When Ptbp1 expression decreases during neuronal differentiation, the regulated introns are spliced out, thus allowing the accumulation of translation-competent mRNAs in the cytoplasm. We propose that this mechanism counters ectopic and precocious expression of functionally linked neuron-specific genes and ensures their coherent activation in the appropriate developmental context. PMID:22661231

Coordinated regulation of neuronal mRNA steady-state levels through developmentally controlled intron retention.

PubMed

Yap, Karen; Lim, Zhao Qin; Khandelia, Piyush; Friedman, Brad; Makeyev, Eugene V

2012-06-01

Differentiated cells acquire unique structural and functional traits through coordinated expression of lineage-specific genes. An extensive battery of genes encoding components of the synaptic transmission machinery and specialized cytoskeletal proteins is activated during neurogenesis, but the underlying regulation is not well understood. Here we show that genes encoding critical presynaptic proteins are transcribed at a detectable level in both neurons and nonneuronal cells. However, in nonneuronal cells, the splicing of 3'-terminal introns within these genes is repressed by the polypyrimidine tract-binding protein (Ptbp1). This inhibits the export of incompletely spliced mRNAs to the cytoplasm and triggers their nuclear degradation. Clearance of these intron-containing transcripts occurs independently of the nonsense-mediated decay (NMD) pathway but requires components of the nuclear RNA surveillance machinery, including the nuclear pore-associated protein Tpr and the exosome complex. When Ptbp1 expression decreases during neuronal differentiation, the regulated introns are spliced out, thus allowing the accumulation of translation-competent mRNAs in the cytoplasm. We propose that this mechanism counters ectopic and precocious expression of functionally linked neuron-specific genes and ensures their coherent activation in the appropriate developmental context.

Differentiation-associated microRNAs antagonize the Rb–E2F pathway to restrict proliferation

PubMed Central

Marzi, Matteo J.; Puggioni, Eleonora M. R.; Dall'Olio, Valentina; Bucci, Gabriele; Bernard, Loris; Bianchi, Fabrizio; Crescenzi, Marco

2012-01-01

The cancer-associated loss of microRNA (miRNA) expression leads to a proliferative advantage and aggressive behavior through largely unknown mechanisms. Here, we exploit a model system that recapitulates physiological terminal differentiation and its reversal upon oncogene expression to analyze coordinated mRNA/miRNA responses. The cell cycle reentry of myotubes, forced by the E1A oncogene, was associated with a pattern of mRNA/miRNA modulation that was largely reciprocal to that induced during the differentiation of myoblasts into myotubes. The E1A-induced mRNA response was preponderantly Retinoblastoma protein (Rb)-dependent. Conversely, the miRNA response was mostly Rb-independent and exerted through tissue-specific factors and Myc. A subset of these miRNAs (miR-1, miR-34, miR-22, miR-365, miR-29, miR-145, and Let-7) was shown to coordinately target Rb-dependent cell cycle and DNA replication mRNAs. Thus, a dual level of regulation—transcriptional regulation via Rb–E2F and posttranscriptional regulation via miRNAs—confers robustness to cell cycle control and provides a molecular basis to understand the role of miRNA subversion in cancer. PMID:23027903

Newt cells secrete extracellular vesicles with therapeutic bioactivity in mammalian cardiomyocytes.

PubMed

Middleton, Ryan C; Rogers, Russell G; De Couto, Geoffrey; Tseliou, Eleni; Luther, Kristin; Holewinski, Ronald; Soetkamp, Daniel; Van Eyk, Jennifer E; Antes, Travis J; Marbán, Eduardo

2018-01-01

Newts can regenerate amputated limbs and cardiac tissue, unlike mammals which lack broad regenerative capacity. Several signaling pathways involved in cell proliferation, differentiation and survival during newt tissue regeneration have been elucidated, however the factors that coordinate signaling between cells, as well as the conservation of these factors in other animals, are not well defined. Here we report that media conditioned by newt limb explant cells (A1 cells) protect mammalian cardiomyocytes from oxidative stress-induced apoptosis. The cytoprotective effect of A1-conditioned media was negated by exposing A1 cells to GW4869, which suppresses the generation of extracellular vesicles (EVs). A1-EVs are similar in diameter (~100-150 nm), structure, and share several membrane surface and cargo proteins with mammalian exosomes. However, isolated A1-EVs contain significantly higher levels of both RNA and protein per particle than mammalian EVs. Additionally, numerous cargo RNAs and proteins are unique to A1-EVs. Of particular note, A1-EVs contain numerous mRNAs encoding nuclear receptors, membrane ligands, as well as transcription factors. Mammalian cardiomyocytes treated with A1-EVs showed increased expression of genes in the PI3K/AKT pathway, a pivotal player in survival signaling. We conclude that newt cells secrete EVs with diverse, distinctive RNA and protein contents. Despite ~300 million years of evolutionary divergence between newts and mammals, newt EVs confer cytoprotective effects on mammalian cardiomyocytes.

Comparative Proteome Analysis between High Lipid-Producing Strain Mucor circinelloides WJ11 and Low Lipid-Producing Strain CBS 277.49.

PubMed

Tang, Xin; Chen, Haiqin; Gu, Zhennan; Zhang, Hao; Chen, Yong Q; Song, Yuanda; Chen, Wei

2017-06-21

Mucor circinelloides is one of few oleaginous fungi that produces a useful oil rich in γ-linolenic acid, but it usually only produces <25% total lipid. Nevertheless, we isolated a new strain WJ11 that can produce up to 36% lipid of cell dry weight. In this study, we have systematically analyzed the global changes in protein levels between the high lipid-producing strain WJ11 and the low lipid-producing strain CBS 277.49 (15%, lipid/cell dry weight) at lipid accumulation phase through comparative proteome analysis. Proteome analysis demonstrated that the branched-chain amino acid and lysine metabolism, glycolytic pathway, and pentose phosphate pathway in WJ11 were up-regulated, while the activities of tricarboxylic acid cycle and branch point enzyme for synthesis of isoprenoids were retarded compared with CBS 277.49. The coordinated regulation at proteome level indicate that more acetyl-CoA and NADPH are provided for fatty acid biosynthesis in WJ11 compared with CBS 277.49.

MRAS: A Close but Understudied Member of the RAS Family.

PubMed

Young, Lucy C; Rodriguez-Viciana, Pablo

2018-01-08

MRAS is the closest relative to the classical RAS oncoproteins and shares most regulatory and effector interactions. However, it also has unique functions, including its ability to function as a phosphatase regulatory subunit when in complex with SHOC2 and protein phosphatase 1 (PP1). This phosphatase complex regulates a crucial step in the activation cycle of RAF kinases and provides a key coordinate input required for efficient ERK pathway activation and transformation by RAS. MRAS mutations rarely occur in cancer but deregulated expression may play a role in tumorigenesis in some settings. Activating mutations in MRAS (as well as SHOC2 and PP1) do occur in the RASopathy Noonan syndrome, underscoring a key role for MRAS within the RAS-ERK pathway. MRAS also has unique roles in cell migration and differentiation and has properties consistent with a key role in the regulation of cell polarity. Further investigations should shed light on what remains a relatively understudied RAS family member. Copyright © 2018 Cold Spring Harbor Laboratory Press; all rights reserved.

The regulation of vascular endothelial growth factor-induced microvascular permeability requires Rac and reactive oxygen species.

PubMed

Monaghan-Benson, Elizabeth; Burridge, Keith

2009-09-18

Vascular permeability is a complex process involving the coordinated regulation of multiple signaling pathways in the endothelial cell. It has long been documented that vascular endothelial growth factor (VEGF) greatly enhances microvascular permeability; however, the molecular mechanisms controlling VEGF-induced permeability remain unknown. Treatment of microvascular endothelial cells with VEGF led to an increase in reactive oxygen species (ROS) production. ROS are required for VEGF-induced permeability as treatment with the free radical scavenger, N-acetylcysteine, inhibited this effect. Additionally, treatment with VEGF caused ROS-dependent tyrosine phosphorylation of both vascular-endothelial (VE)-cadherin and beta-catenin. Rac1 was required for the VEGF-induced increase in permeability and adherens junction protein phosphorylation. Knockdown of Rac1 inhibited VEGF-induced ROS production consistent with Rac lying upstream of ROS in this pathway. Collectively, these data suggest that VEGF leads to a Rac-mediated generation of ROS, which, in turn, elevates the tyrosine phosphorylation of VE-cadherin and beta-catenin, ultimately regulating adherens junction integrity.

The Neurotensin Receptor-1 Pathway Contributes to Human Ductal Breast Cancer Progression

PubMed Central

Dupouy, Sandra; Viardot-Foucault, Véronique; Alifano, Marco; Souazé, Frédérique; Plu-Bureau, Geneviève; Chaouat, Marc; Lavaur, Anne; Hugol, Danielle; Gespach, Christian

2009-01-01

Background The neurotensin (NTS) and its specific high affinity G protein coupled receptor, the NT1 receptor (NTSR1), are considered to be a good candidate for one of the factors implicated in neoplastic progression. In breast cancer cells, functionally expressed NT1 receptor coordinates a series of transforming functions including cellular migration and invasion. Methods and Results we investigated the expression of NTS and NTSR1 in normal human breast tissue and in invasive ductal breast carcinomas (IDCs) by immunohistochemistry and RT-PCR. NTS is expressed and up-regulated by estrogen in normal epithelial breast cells. NTS is also found expressed in the ductal and invasive components of IDCs. The high expression of NTSR1 is associated with the SBR grade, the size of the tumor, and the number of metastatic lymph nodes. Furthermore, the NTSR1 high expression is an independent factor of prognosis associated with the death of patients. Conclusion these data support the activation of neurotensinergic deleterious pathways in breast cancer progression. PMID:19156213

The neurotensin receptor-1 pathway contributes to human ductal breast cancer progression.

PubMed

Dupouy, Sandra; Viardot-Foucault, Véronique; Alifano, Marco; Souazé, Frédérique; Plu-Bureau, Geneviève; Chaouat, Marc; Lavaur, Anne; Hugol, Danielle; Gespach, Christian; Gompel, Anne; Forgez, Patricia

2009-01-01

The neurotensin (NTS) and its specific high affinity G protein coupled receptor, the NT1 receptor (NTSR1), are considered to be a good candidate for one of the factors implicated in neoplastic progression. In breast cancer cells, functionally expressed NT1 receptor coordinates a series of transforming functions including cellular migration and invasion. we investigated the expression of NTS and NTSR1 in normal human breast tissue and in invasive ductal breast carcinomas (IDCs) by immunohistochemistry and RT-PCR. NTS is expressed and up-regulated by estrogen in normal epithelial breast cells. NTS is also found expressed in the ductal and invasive components of IDCs. The high expression of NTSR1 is associated with the SBR grade, the size of the tumor, and the number of metastatic lymph nodes. Furthermore, the NTSR1 high expression is an independent factor of prognosis associated with the death of patients. these data support the activation of neurotensinergic deleterious pathways in breast cancer progression.

Tombusvirus RNA replication depends on the TOR pathway in yeast and plants.

PubMed

Inaba, Jun-Ichi; Nagy, Peter D

2018-06-01

Similar to other (+)RNA viruses, tomato bushy stunt virus (TBSV) utilizes metabolites, lipids, membranes, and co-opted host factors during replication. The coordination of cell metabolism and growth with environmental cues is performed by the target of rapamycin (TOR) kinase in eukaryotic cells. In this paper, we find that TBSV replication partially inhibits TOR activity, likely due to recruitment of glycolytic enzymes to the viral replication compartment, which results in reduced ATP levels in the cytosol. Complete inhibition of TOR activity with rapamycin in yeast or AZD8055 inhibitor in plants reduces tombusvirus replication. We find that high glucose concentration, which stimulates TOR activity, enhanced tombusvirus replication in yeast. Depletion of yeast Sch9 or plant S6K1 kinase, a downstream effector of TOR, also inhibited tombusvirus replication in yeast and plant or the assembly of the viral replicase in vitro. Altogether, the TOR pathway is crucial for TBSV to replicate efficiently in hosts. Copyright © 2018 Elsevier Inc. All rights reserved.

Coordinated gene expression for pheromone biosynthesis in the pine engraver beetle, Ips pini (Coleoptera: Scolytidae)

NASA Astrophysics Data System (ADS)

Keeling, Christopher I.; Blomquist, Gary J.; Tittiger, Claus

In several pine bark beetle species, phloem feeding induces aggregation pheromone production to coordinate a mass attack on the host tree. Male pine engraver beetles, Ips pini (Say) (Coleoptera: Scolytidae), produce the monoterpenoid pheromone component ipsdienol de novo via the mevalonate pathway in the anterior midgut upon feeding. To understand how pheromone production is regulated in this tissue, we used quantitative real-time PCR to examine feeding-induced changes in gene expression of seven mevalonate pathway genes: acetoacetyl-coenzyme A thiolase, 3-hydroxy-3-methylglutaryl coenzyme A synthase, 3-hydroxy-3-methylglutaryl coenzyme A reductase, mevalonate 5-diphosphate decarboxylase, isopentenyl-diphosphate isomerase, geranyl-diphosphate synthase (GPPS), and farnesyl-diphosphate synthase (FPPS). In males, expression of all these genes significantly increased upon feeding. In females, the expression of the early mevalonate pathway genes (up to and including the isomerase) increased significantly, but the expression of the later genes (GPPS and FPPS) was unaffected or decreased upon feeding. Thus, feeding coordinately regulates expression of the mevalonate pathway genes necessary for pheromone biosynthesis in male, but not female, midguts. Furthermore, basal mRNA levels were 5- to 41-fold more abundant in male midguts compared to female midguts. This is the first report of coordinated regulation of mevalonate pathway genes in an invertebrate model consistent with their sex-specific role in de novo pheromone biosynthesis.

Mechanosensation Dynamically Coordinates Polar Growth and Cell Wall Assembly to Promote Cell Survival.

PubMed

Davì, Valeria; Tanimoto, Hirokazu; Ershov, Dmitry; Haupt, Armin; De Belly, Henry; Le Borgne, Rémi; Couturier, Etienne; Boudaoud, Arezki; Minc, Nicolas

2018-04-23

How growing cells cope with size expansion while ensuring mechanical integrity is not known. In walled cells, such as those of microbes and plants, growth and viability are both supported by a thin and rigid encasing cell wall (CW). We deciphered the dynamic mechanisms controlling wall surface assembly during cell growth, using a sub-resolution microscopy approach to monitor CW thickness in live rod-shaped fission yeast cells. We found that polar cell growth yielded wall thinning and that thickness negatively influenced growth. Thickness at growing tips exhibited a fluctuating behavior with thickening phases followed by thinning phases, indicative of a delayed feedback promoting thickness homeostasis. This feedback was mediated by mechanosensing through the CW integrity pathway, which probes strain in the wall to adjust synthase localization and activity to surface growth. Mutants defective in thickness homeostasis lysed by rupturing the wall, demonstrating its pivotal role for walled cell survival. Copyright © 2018 Elsevier Inc. All rights reserved.

PKR is a novel functional direct player that coordinates skeletal muscle differentiation via p38MAPK/AKT pathways.

PubMed

Alisi, A; Spaziani, A; Anticoli, S; Ghidinelli, M; Balsano, C

2008-03-01

Myogenic differentiation is a highly orchestrated multistep process controlled by extracellular growth factors that modulate largely unknown signals into the cell affecting the muscle-transcription program. P38MAPK-dependent signalling, as well as PI3K/Akt pathway, has a key role in the control of muscle gene expression at different stages during the myogenic process. P38MAPK affects the activities of transcription factors, such as MyoD and myogenin, and contributes, together with PI3K/Akt pathway, to control the early and late steps of myogenic differentiation. The aim of our work was to better define the role of PKR, a dsRNA-activated protein kinase, as potential component in the differentiation program of C2C12 murine myogenic cells and to correlate its activity with p38MAPK and PI3K/Akt myogenic regulatory pathways. Here, we demonstrate that PKR is an essential component of the muscle development machinery and forms a functional complex with p38MAPK and/or Akt, contributing to muscle differentiation of committed myogenic cells in vitro. Inhibition of endogenous PKR activity by a specific (si)RNA and a PKR dominant-negative interferes with the myogenic program of C2C12 cells, causing a delay in activation of myogenic specific genes and inducing the formation of thinner myofibers. In addition, the construction of three PKR mutants allowed us to demonstrate that both N and C-terminal regions of PKR are critical for the interaction with p38MAPK and Akt. The novel discovered complex permits PKR to timely regulate the inhibition/activation of p38MAPK and Akt, controlling in this way the different steps characterizing skeletal muscle differentiation.

Organotypic three-dimensional culture model of mesenchymal ...

EPA Pesticide Factsheets

Tissue fusion during early mammalian development requires coordination of multiple cell types, the extracellular matrix, and complex signaling pathways. Fusion events during processes including heart development, neural tube closure, and palatal fusion are dependent on signaling pathways elucidated using gene knockout mouse models. A broad analysis of literature, ToxRefDB, and ToxCast identified 63 chemicals that are related to cleft palate. However,the influence of these putative teratogens on human palatal fusion has not been studied due to the lack of in vitro models. We sought to engineer the stratified mesenchymal and epithelial structure of the developing palate in vitro via organotypic culture of human mesenchymal stem cell (hMSC) spheroids coated with a single layer of human primary epidermalkeratinocytes (hPEKp). hMSC spheroids exhibited uniform size over time (175 ± 21 µm mean diameter) proportional to starting cell density. Further, we developed a novel procedure to coat hMSC spheroids homogeneously with a single layer of hPEKp cells using a seeding ratio of 0.1-0.2 hPEKp per hMSC, and hMSC/hPEKp spheroids expressed mesenchymal markers (vim+, C044+, CD105+, CD34-) and epithelial markers (krt17+, itga6+) via qRT-PCR. Analysis of adverse outcome pathways related to palate fusion points to an EGF/TGFj33 switch that could be a target for cleft palate teratogens, and both egf and egfr were expressed by hMSC/hPEKp spheres. Finally, hMSCs and hPE

Twenty Years of Calcium Imaging: Cell Physiology to Dye For

PubMed Central

Knot, Harm J.; Laher, Ismail; Sobie, Eric A.; Guatimosim, Silvia; Gomez-Viquez, Leticia; Hartmann, Hali; Song, Long-Sheng; Lederer, W.J.; Graier, Wolfgang F.; Malli, Roland; Frieden, Maud; Petersen, Ole H.

2016-01-01

The use of fluorescent dyes over the past two decades has led to a revolution in our understanding of calcium signaling. Given the ubiquitous role of Ca2+ in signal transduction at the most fundamental levels of molecular, cellular, and organismal biology, it has been challenging to understand how the specificity and versatility of Ca2+ signaling is accomplished. In excitable cells, the coordination of changing Ca2+ concentrations at global (cellular) and well-defined subcellular spaces through the course of membrane depolarization can now be conceptualized in the context of disease processes such as cardiac arrhythmogenesis. The spatial and temporal dimensions of Ca2+ signaling are similarly important in non-excitable cells, such as endothelial and epithelial cells, to regulate multiple signaling pathways that participate in organ homeostasis as well as cellular organization and essential secretory processes. PMID:15821159

Extracellular Matrix Signaling from the Cellular Membrane Skeleton to the Nuclear Skeleton: A Model of Gene Regulation

PubMed Central

Lelièvre, Sophie; Weaver, Valerie M.; Bissell, Mina J.

2010-01-01

It is well established that cells must interact with their microenvironment and that such interaction is crucial for coordinated function and homeostasis. However, how cells receive and integrate external signals leading to gene regulation is far from understood. It is now appreciated that two classes of cooperative signals are implicated: a soluble class including hormones and growth factors and a class of insoluble signals emanating from the extracellular matrix (ECM) directly through contact with the cell surface. Using 3-dimensional culture systems and transgenic mice, we have been able to identify some of the elements of this ECM-signaling pathway responsible for gene regulation in rodent mammary gland differentiation and involution. Our major observations are 1) the requirement for a laminin-rich basement membrane; 2) the existence of a cooperative signaling pathway between basement membrane and the lactogenic hormone prolactin (PRL); 3) the importance of β1-integrins and bHLH transcription factor(s) and the presence of DNA response elements (exemplified by BCE-1, located on a milk protein gene, β-casein); and 4) the induction of mammary epithelial cell programmed cell death following degradation of basement membrane. We hypothesize that this cooperative signaling between ECM and PRL may be achieved through integrin- and laminin-directed restructuring of the cytoskeleton leading to profound changes in nuclear architecture and transcription factor localization. We postulate that the latter changes allow the prolactin signal to activate transcription of the β-casein gene. To further understand the molecular mechanisms underlying ECM and hormonal cooperative signaling, we are currently investigating ECM regulation of a “solid-state” signaling pathway including ECM fiber proteins, plasma membrane receptors, cytoskeleton, nuclear matrix and chromatin. We further postulate that disruption of such a pathway may be implicated in cell disorders including transformation and carcinogenesis. PMID:8701089

Coordinated induction of cell survival signaling in the inflamed microenvironment of the prostate.

PubMed

McIlwain, David W; Zoetemelk, Marloes; Myers, Jason D; Edwards, Marshé T; Snider, Brandy M; Jerde, Travis J

2016-06-01

Both prostate cancer and benign prostatic hyperplasia are associated with inflammatory microenvironments. Inflammation is damaging to tissues, but it is unclear how the inflammatory microenvironment protects specialized epithelial cells that function to proliferate and repair the tissue. The objective of this study is to characterize the cell death and cell survival response of the prostatic epithelium in response to inflammation. We assessed induction of cell death (TNF, TRAIL, TWEAK, FasL) and cell survival factors (IGFs, hedgehogs, IL-6, FGFs, and TGFs) in inflamed and control mouse prostates by ELISA. Cell death mechanisms were determined by immunoblotting and immunofluorescence for cleavage of caspases and TUNEL. Survival pathway activation was assessed by immunoblotting and immunofluorescence for Mcl-1, Bcl-2, Bcl-XL, and survivin. Autophagy was determined by immunoblotting and immunofluorescence for free and membrane associated light chain 3 (LC-3). Cleavage of all four caspases was significantly increased during the first 2 days of inflammation, and survival protein expression was substantially increased subsequently, maximizing at 3 days. By 5 days of inflammation, 50% of prostatic epithelial cells expressed survivin. Autophagy was also evident during the recovery phase (3 days). Finally, immunofluorescent staining of human specimens indicates strong activation of survival proteins juxtaposed to inflammation in inflamed prostate specimens. The prostate responds to deleterious inflammation with induction of cell survival mechanisms, most notably survivin and autophagy, demonstrating a coordinated induction of survival factors that protects and expands a specialized set of prostatic epithelial cells as part of the repair and recovery process during inflammation. © 2016 Wiley Periodicals, Inc.

Secretory carrier membrane protein 5 is an autophagy inhibitor that promotes the secretion of α-synuclein via exosome.

PubMed

Yang, Yi; Qin, Meiling; Bao, Puhua; Xu, Wangchao; Xu, Jin

2017-01-01

Autophagy-lysosomal pathway is a cellular protective system to remove aggregated proteins and damaged organelles. Meanwhile, exosome secretion has emerged as a mode to selectively clear the neurotoxic proteins, such as α-synuclein. Mounting evidence suggests that these two cellular processes are coordinated to facilitate the clearance of toxic cellular waste; however the regulators for the transition between these two processes are unclear. Here we show that SCAMP5, a secretory carrier membrane protein significantly induced in the brains of Huntington's disease patients, is quickly and transiently induced by protein stress and autophagic stimulation, and is regulated by the master autophagy transcriptional regulator TFEB. Ironically, SCAMP5 inhibits autophagy flux by blocking the fusion of autophagosomes and lysosomes. Although autophagy is blocked, SCAMP5 does not cause significant protein aggregation in cells. Instead, it promotes the Golgi fragmentation and stimulates the unconventional secretion of the co-localizing α-synuclein via exosome as an exosome component. Therefore, we have identified SCAMP5 as a novel coordinator of autophagy and exosome secretion, which is induced upon protein stress to channel the efficient clearance of toxic proteins via the exosomes rather than autophagy-lysosomal pathway.

Synergistic role of Sprouty2 inactivation and c-Met up-regulation in mouse and human hepatocarcinogenesis.

PubMed

Lee, Susie A; Ladu, Sara; Evert, Matthias; Dombrowski, Frank; De Murtas, Valentina; Chen, Xin; Calvisi, Diego F

2010-08-01

Sprouty2 (Spry2), a negative feedback regulator of the Ras/mitogen-activated protein kinase (MAPK) pathway, is frequently down-regulated in human hepatocellular carcinoma (HCC). We tested the hypothesis that loss of Spry2 cooperates with unconstrained activation of the c-Met protooncogene to induce hepatocarcinogenesis via in vitro and in vivo approaches. We found coordinated down-regulation of Spry2 protein expression and activation of c-Met as well as its downstream effectors extracellular signal-regulated kinase (ERK) and v-akt murine thymoma viral oncogene homolog (AKT) in a subset of human HCC samples with poor outcome. Mechanistic studies revealed that Spry2 function is disrupted in human HCC via multiple mechanisms at both transcriptional and post-transcriptional level, including promoter hypermethylation, loss of heterozygosity, and proteosomal degradation by neural precursor cell expressed, developmentally down-regulated 4 (NEDD4). In HCC cell lines, Spry2 overexpression inhibits c-Met-induced cell proliferation as well as ERK and AKT activation, whereas loss of Spry2 potentiates c-Met signaling. Most importantly, we show that blocking Spry2 activity via a dominant negative form of Spry2 cooperates with c-Met to promote hepatocarcinogenesis in the mouse liver by sustaining proliferation and angiogenesis. The tumors exhibited high levels of activated ERK and AKT, recapitulating the subgroup of human HCC with a clinically aggressive phenotype. The occurrence of frequent genetic, epigenetic, and biochemical events leading to Spry2 inactivation provides solid evidence that Spry2 functions as a tumor suppressor gene in liver cancer. Coordinated deregulation of Spry2 and c-Met signaling may be a pivotal oncogenic mechanism responsible for unrestrained activation of ERK and AKT pathways in human hepatocarcinogenesis.

Enhancing microbial production of biofuels by expanding microbial metabolic pathways.

PubMed

Yu, Ping; Chen, Xingge; Li, Peng

2017-09-01

Fatty acid, isoprenoid, and alcohol pathways have been successfully engineered to produce biofuels. By introducing three genes, atfA, adhE, and pdc, into Escherichia coli to expand fatty acid pathway, up to 1.28 g/L of fatty acid ethyl esters can be achieved. The isoprenoid pathway can be expanded to produce bisabolene with a high titer of 900 mg/L in Saccharomyces cerevisiae. Short- and long-chain alcohols can also be effectively biosynthesized by extending the carbon chain of ketoacids with an engineered "+1" alcohol pathway. Thus, it can be concluded that expanding microbial metabolic pathways has enormous potential for enhancing microbial production of biofuels for future industrial applications. However, some major challenges for microbial production of biofuels should be overcome to compete with traditional fossil fuels: lowering production costs, reducing the time required to construct genetic elements and to increase their predictability and reliability, and creating reusable parts with useful and predictable behavior. To address these challenges, several aspects should be further considered in future: mining and transformation of genetic elements related to metabolic pathways, assembling biofuel elements and coordinating their functions, enhancing the tolerance of host cells to biofuels, and creating modular subpathways that can be easily interconnected. © 2016 International Union of Biochemistry and Molecular Biology, Inc.

Growth and stress response mechanisms underlying post-feeding regenerative organ growth in the Burmese python.

PubMed

Andrew, Audra L; Perry, Blair W; Card, Daren C; Schield, Drew R; Ruggiero, Robert P; McGaugh, Suzanne E; Choudhary, Amit; Secor, Stephen M; Castoe, Todd A

2017-05-02

Previous studies examining post-feeding organ regeneration in the Burmese python (Python molurus bivittatus) have identified thousands of genes that are significantly differentially regulated during this process. However, substantial gaps remain in our understanding of coherent mechanisms and specific growth pathways that underlie these rapid and extensive shifts in organ form and function. Here we addressed these gaps by comparing gene expression in the Burmese python heart, liver, kidney, and small intestine across pre- and post-feeding time points (fasted, one day post-feeding, and four days post-feeding), and by conducting detailed analyses of molecular pathways and predictions of upstream regulatory molecules across these organ systems. Identified enriched canonical pathways and upstream regulators indicate that while downstream transcriptional responses are fairly tissue specific, a suite of core pathways and upstream regulator molecules are shared among responsive tissues. Pathways such as mTOR signaling, PPAR/LXR/RXR signaling, and NRF2-mediated oxidative stress response are significantly differentially regulated in multiple tissues, indicative of cell growth and proliferation along with coordinated cell-protective stress responses. Upstream regulatory molecule analyses identify multiple growth factors, kinase receptors, and transmembrane receptors, both within individual organs and across separate tissues. Downstream transcription factors MYC and SREBF are induced in all tissues. These results suggest that largely divergent patterns of post-feeding gene regulation across tissues are mediated by a core set of higher-level signaling molecules. Consistent enrichment of the NRF2-mediated oxidative stress response indicates this pathway may be particularly important in mediating cellular stress during such extreme regenerative growth.

Time-dependent LXR/RXR pathway modulation characterizes capillary remodeling in inflammatory corneal neovascularization.

PubMed

Mukwaya, Anthony; Lennikov, Anton; Xeroudaki, Maria; Mirabelli, Pierfrancesco; Lachota, Mieszko; Jensen, Lasse; Peebo, Beatrice; Lagali, Neil

2018-05-01

Inflammation in the normally immune-privileged cornea can initiate a pathologic angiogenic response causing vision-threatening corneal neovascularization. Inflammatory pathways, however, are numerous, complex and are activated in a time-dependent manner. Effective resolution of inflammation and associated angiogenesis in the cornea requires knowledge of these pathways and their time dependence, which has, to date, remained largely unexplored. Here, using a model of endogenous resolution of inflammation-induced corneal angiogenesis, we investigate the time dependence of inflammatory genes in effecting capillary regression and the return of corneal transparency. Endogenous capillary regression was characterized by a progressive thinning and remodeling of angiogenic capillaries and inflammatory cell retreat in vivo in the rat cornea. By whole-genome longitudinal microarray analysis, early suppression of VEGF ligand-receptor signaling and inflammatory pathways preceded an unexpected later-phase preferential activation of LXR/RXR, PPARα/RXRα and STAT3 canonical pathways, with a concurrent attenuation of LPS/IL-1 inhibition of RXR function and Wnt/β-catenin signaling pathways. Potent downstream inflammatory cytokines such as Cxcl5, IL-1β, IL-6 and Ccl2 were concomitantly downregulated during the remodeling phase. Upstream regulators of the inflammatory pathways included Socs3, Sparc and ApoE. A complex and coordinated time-dependent interplay between pro- and anti-inflammatory signaling pathways highlights a potential anti-inflammatory role of LXR/RXR, PPARα/RXRα and STAT3 signaling pathways in resolving inflammatory corneal angiogenesis.

Novel prosurvival function of Yip1A in human cervical cancer cells: constitutive activation of the IRE1 and PERK pathways of the unfolded protein response.

PubMed

Taguchi, Yuki; Horiuchi, Yuta; Kano, Fumi; Murata, Masayuki

2017-03-30

Cancer cells are under chronic endoplasmic reticulum (ER) stress due to hypoxia, low levels of nutrients, and a high metabolic demand for proliferation. To survive, they constitutively activate the unfolded protein response (UPR). The inositol-requiring protein 1 (IRE1) and protein kinase RNA-like ER kinase (PERK) signaling branches of the UPR have been shown to have cytoprotective roles in cancer cells. UPR-induced autophagy is another prosurvival strategy of cancer cells, possibly to remove misfolded proteins and supply nutrients. However, the mechanisms by which cancer cells exploit the UPR and autophagy machinery to promote survival and the molecules that are essential for these processes remain to be elucidated. Recently, a multipass membrane protein, Yip1A, was shown to function in the activation of IRE1 and in UPR-induced autophagy. In the present study, we explored the possible role of Yip1A in activation of the UPR by cancer cells for their survival, and found that depletion of Yip1A by RNA interference (RNAi) induced apoptotic cell death in HeLa and CaSki cervical cancer cells. Intriguingly, Yip1A was found to activate the IRE1 and PERK pathways of the UPR constitutively in HeLa and CaSki cells. Yip1A mediated the phosphorylation of IRE1 and also engaged in the transcription of PERK. The activation of these signaling pathways upregulated the expression of anti-apoptotic proteins and autophagy-related proteins. These events might enhance resistance to apoptosis and promote cytoprotective autophagy in HeLa and CaSki cells. The present study is the first to uncover a key prosurvival modulator, Yip1A, which coordinates IRE1 signaling with PERK signaling to support the survival of HeLa and CaSki cervical cancer cells.

Coordinate regulation of the Suf and Isc Fe-S cluster biogenesis pathways by IscR is essential for viability of Escherichia coli.

PubMed

Mettert, Erin L; Kiley, Patricia J

2014-12-01

Fe-S cluster biogenesis is essential for the viability of most organisms. In Escherichia coli, this process requires either the housekeeping Isc or the stress-induced Suf pathway. The global regulator IscR coordinates cluster synthesis by repressing transcription of the isc operon by [2Fe-2S]-IscR and activating expression of the suf operon. We show that either [2Fe-2S]-IscR or apo-IscR can activate suf, making expression sensitive to mainly IscR levels and not the cluster state, unlike isc expression. We also demonstrate that in the absence of isc, IscR-dependent suf activation is essential since strains lacking both the Isc pathway and IscR were not viable unless Suf was expressed ectopically. Similarly, removal of the IscR binding site in the sufA promoter also led to a requirement for isc. Furthermore, suf expression was increased in a Δisc mutant, presumably due to increased IscR levels in this mutant. This was surprising because the iron-dependent repressor Fur, whose higher-affinity binding at the sufA promoter should occlude IscR binding, showed only partial repression. In addition, Fur derepression was not sufficient for viability in the absence of IscR and the Isc pathway, highlighting the importance of direct IscR activation. Finally, a mutant lacking Fur and the Isc pathway increased suf expression to the highest observed levels and nearly restored [2Fe-2S]-IscR activity, providing a mechanism for regulating IscR activity under stress conditions. Together, these findings have enhanced our understanding of the homeostatic mechanism by which cells use one regulator, IscR, to differentially control Fe-S cluster biogenesis pathways to ensure viability. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

The metazoan Mediator co-activator complex as an integrative hub for transcriptional regulation.

PubMed

Malik, Sohail; Roeder, Robert G

2010-11-01

The Mediator is an evolutionarily conserved, multiprotein complex that is a key regulator of protein-coding genes. In metazoan cells, multiple pathways that are responsible for homeostasis, cell growth and differentiation converge on the Mediator through transcriptional activators and repressors that target one or more of the almost 30 subunits of this complex. Besides interacting directly with RNA polymerase II, Mediator has multiple functions and can interact with and coordinate the action of numerous other co-activators and co-repressors, including those acting at the level of chromatin. These interactions ultimately allow the Mediator to deliver outputs that range from maximal activation of genes to modulation of basal transcription to long-term epigenetic silencing.

Stop Stalling: Mus81 Required for Efficient Replication | Center for Cancer Research

Cancer.gov

DNA replication is precisely controlled to ensure that daughter cells receive intact, accurate genetic information. Each segment of DNA must be copied only once, and the rate of replication coordinated genome-wide. Mild replication stress slows DNA synthesis and activates a pathway involving the Mus81 endonuclease, which generates a series of DNA breaks that are rapidly repaired, allowing the cell to avoid activating the S-phase checkpoint and its potentially damaging outcomes of apoptosis or error-prone repair. Mirit Aladjem, Ph.D., of CCR’s Developmental Therapeutics Branch, and her colleagues wondered whether Mus81 also plays a role in regulating the replication rate during growth in the absence of stress.

High-Pressure Behavior of Difluorides: The Case of SrF2

NASA Astrophysics Data System (ADS)

Swadba, K. E.; Stan, C. V.; Dutta, R.; Prakapenka, V.; Duffy, T. S.

2016-12-01

The high-pressure behavior of compounds in the AX2 family has attracted much attention due to their extensive polymorphism, highly coordinated structures, and diverse transformation pathways. The canonical transformation sequence for alkaline earth difluorides is from the fluorite-type structure (8 coordinated) to cotunnite (9 coordinated) to Ni2In (11 coordinated). Lead Fluoride, on the other hand, undergoes an unusual isosymmetric transition from cotunnite to a Co2Si-type structure (10 coordinated) at high pressures, during which it exhibits highly anisotropic lattice parameter trends (Haines et al, 1998; Stan et al 2016). Sr has a similar ionic radius as Pb, and is thus a good candidate for further exploring the compressional anisotropy in alkaline earth fluorides. In this study, we report a detailed examination of the compressional behavior of SrF2 to identify whether an intermediate phase occurs in this system prior to transformation to the Ni2In structure. Raman spectroscopy and x-ray diffraction experiments, performed at Princeton University and the Advanced Photon Source GSECARS beamline, respectively, were carried out on SrF2 up to 63 GPa using a diamond anvil cell. From Raman spectroscopy, we observed evidence for a high-pressure phase transition between 38.9 and 51.0 GPa. The x-ray diffraction data in this region show evidence for highly anisotropic compression, most notably a strong negative compressibility in the b direction, in the pressure region from 45.2 to 51.6 GPa. Comparison of our data with lattice parameter systematics for AX2 phases indicates that our results are consistent with the formation of the Co2Si phase in this region, along with a sluggish transformation to the Ni2In-type structure. Our findings contribute to a broader understanding of AX2 compounds and their phase transition pathways.

Role and regulation of coordinately expressed de novo purine biosynthetic enzymes PPAT and PAICS in lung cancer.

PubMed

Goswami, Moloy T; Chen, Guoan; Chakravarthi, Balabhadrapatruni V S K; Pathi, Satya S; Anand, Sharath K; Carskadon, Shannon L; Giordano, Thomas J; Chinnaiyan, Arul M; Thomas, Dafydd G; Palanisamy, Nallasivam; Beer, David G; Varambally, Sooryanarayana

2015-09-15

Cancer cells exhibit altered metabolism including aerobic glycolysis that channels several glycolytic intermediates into de novo purine biosynthetic pathway. We discovered increased expression of phosphoribosyl amidotransferase (PPAT) and phosphoribosylaminoimidazole carboxylase, phosphoribosylaminoimidazole succinocarboxamide synthetase (PAICS) enzymes of de novo purine biosynthetic pathway in lung adenocarcinomas. Transcript analyses from next-generation RNA sequencing and gene expression profiling studies suggested that PPAT and PAICS can serve as prognostic biomarkers for aggressive lung adenocarcinoma. Immunohistochemical analysis of PAICS performed on tissue microarrays showed increased expression with disease progression and was significantly associated with poor prognosis. Through gene knockdown and over-expression studies we demonstrate that altering PPAT and PAICS expression modulates pyruvate kinase activity, cell proliferation and invasion. Furthermore we identified genomic amplification and aneuploidy of the divergently transcribed PPAT-PAICS genomic region in a subset of lung cancers. We also present evidence for regulation of both PPAT and PAICS and pyruvate kinase activity by L-glutamine, a co-substrate for PPAT. A glutamine antagonist, 6-Diazo-5-oxo-L-norleucine (DON) blocked glutamine mediated induction of PPAT and PAICS as well as reduced pyruvate kinase activity. In summary, this study reveals the regulatory mechanisms by which purine biosynthetic pathway enzymes PPAT and PAICS, and pyruvate kinase activity is increased and exposes an existing metabolic vulnerability in lung cancer cells that can be explored for pharmacological intervention.

The angiopoietin1-Akt pathway regulates barrier function of the cultured spinal cord microvascular endothelial cells through Eps8.

PubMed

Liu, Xinchun; Zhou, Xiaoshu; Yuan, Wei

2014-10-15

In mammalian central nervous system (CNS), the integrity of the blood-spinal cord barrier (BSCB), formed by tight junctions (TJs) between adjacent microvascular endothelial cells near the basement membrane of capillaries and the accessory structures, is important for relatively independent activities of the cellular constituents inside the spinal cord. The barrier function of the BSCB are tightly regulated and coordinated by a variety of physiological or pathological factors, similar with but not quite the same as its counterpart, the blood-brain barrier (BBB). Herein, angiopoietin 1 (Ang1), an identified ligand of the endothelium-specific tyrosine kinase receptor Tie-2, was verified to regulate barrier functions, including permeability, junction protein interactions and F-actin organization, in cultured spinal cord microvascular endothelial cells (SCMEC) of rat through the activity of Akt. Besides, these roles of Ang1 in the BSCB in vitro were found to be accompanied with an increasing expression of epidermal growth factor receptor pathway substrate 8 (Eps8), an F-actin bundling protein. Furthermore, the silencing of Eps8 by lentiviral shRNA resulted in an antagonistic effect vs. Ang1 on the endothelial barrier function of SCMEC. In summary, the Ang1-Akt pathway serves as a regulator in the barrier function modulation of SCMEC via the actin-binding protein Eps8. Copyright © 2014 Elsevier Inc. All rights reserved.

Insights into the Regulation of DMSP Synthesis in the Diatom Thalassiosira pseudonana through APR Activity, Proteomics and Gene Expression Analyses on Cells Acclimating to Changes in Salinity, Light and Nitrogen

PubMed Central

Kettles, Nicola Louise; Kopriva, Stanislav; Malin, Gill

2014-01-01

Despite the importance of dimethylsulphoniopropionate (DMSP) in the global sulphur cycle and climate regulation, the biological pathways underpinning its synthesis in marine phytoplankton remain poorly understood. The intracellular concentration of DMSP increases with increased salinity, increased light intensity and nitrogen starvation in the diatom Thalassiosira pseudonana. We used these conditions to investigate DMSP synthesis at the cellular level via analysis of enzyme activity, gene expression and proteome comparison. The activity of the key sulphur assimilatory enzyme, adenosine 5′-phosphosulphate reductase was not coordinated with increasing intracellular DMSP concentration. Under all three treatments coordination in the expression of sulphur assimilation genes was limited to increases in sulphite reductase transcripts. Similarly, proteomic 2D gel analysis only revealed an increase in phosphoenolpyruvate carboxylase following increases in DMSP concentration. Our findings suggest that increased sulphur assimilation might not be required for increased DMSP synthesis, instead the availability of carbon and nitrogen substrates may be important in the regulation of this pathway. This contrasts with the regulation of sulphur metabolism in higher plants, which generally involves up-regulation of several sulphur assimilatory enzymes. In T. pseudonana changes relating to sulphur metabolism were specific to the individual treatments and, given that little coordination was seen in transcript and protein responses across the three growth conditions, different patterns of regulation might be responsible for the increase in DMSP concentration seen under each treatment. PMID:24733415

Elevation of adenylate energy charge by angiopoietin-like 4 enhances epithelial-mesenchymal transition by inducing 14-3-3γ expression.

PubMed

Teo, Z; Sng, M K; Chan, J S K; Lim, M M K; Li, Y; Li, L; Phua, T; Lee, J Y H; Tan, Z W; Zhu, P; Tan, N S

2017-11-16

Metastatic cancer cells acquire energy-intensive processes including increased invasiveness and chemoresistance. However, how the energy demand is met and the molecular drivers that coordinate an increase in cellular metabolic activity to drive epithelial-mesenchymal transition (EMT), the first step of metastasis, remain unclear. Using different in vitro and in vivo EMT models with clinical patient's samples, we showed that EMT is an energy-demanding process fueled by glucose metabolism-derived adenosine triphosphate (ATP). We identified angiopoietin-like 4 (ANGPTL4) as a key player that coordinates an increase in cellular energy flux crucial for EMT via an ANGPTL4/14-3-3γ signaling axis. This augmented cellular metabolic activity enhanced metastasis. ANGPTL4 knockdown suppresses an adenylate energy charge elevation, delaying EMT. Using an in vivo dual-inducible EMT model, we found that ANGPTL4 deficiency reduces cancer metastasis to the lung and liver. Unbiased kinase inhibitor screens and Ingenuity Pathway Analysis revealed that ANGPTL4 regulates the expression of 14-3-3γ adaptor protein via the phosphatidylinositol-3-kinase/AKT and mitogen-activated protein kinase signaling pathways that culminate to activation of transcription factors, CREB, cFOS and STAT3. Using a different mode of action, as compared with protein kinases, the ANGPTL4/14-3-3γ signaling axis consolidated cellular bioenergetics and stabilized critical EMT proteins to coordinate energy demand and enhanced EMT competency and metastasis, through interaction with specific phosphorylation signals on target proteins.

An origin-deficient yeast artificial chromosome triggers a cell cycle checkpoint.

PubMed

van Brabant, A J; Buchanan, C D; Charboneau, E; Fangman, W L; Brewer, B J

2001-04-01

Checkpoint controls coordinate entry into mitosis with the completion of DNA replication. Depletion of nucleotide precursors by treatment with the drug hydroxyurea triggers such a checkpoint response. However, it is not clear whether the signal for this hydroxyurea-induced checkpoint pathway is the presence of unreplicated DNA, or rather the persistence of single-stranded or damaged DNA. In a yeast artificial chromosome (YAC) we have engineered an approximately 170 kb region lacking efficient replication origins that allows us to explore the specific effects of unreplicated DNA on cell cycle progression. Replication of this YAC extends the length of S phase and causes cells to engage an S/M checkpoint. In the absence of Rad9 the YAC becomes unstable, undergoing deletions within the origin-free region.

Yersinia versus host immunity: how a pathogen evades or triggers a protective response.

PubMed

Chung, Lawton K; Bliska, James B

2016-02-01

The human pathogenic Yersinia species cause diseases that represent a significant source of morbidity and mortality. Despite this, specific mechanisms underlying Yersinia pathogenesis and protective host responses remain poorly understood. Recent studies have shown that Yersinia disrupt cell death pathways, perturb inflammatory processes and exploit immune cells to promote disease. The ensuing host responses following Yersinia infection include coordination of innate and adaptive immune responses in an attempt to control bacterial replication. Here, we highlight current advances in our understanding of the interactions between the pathogenic yersiniae and host cells, as well as the protective host responses mobilized to counteract these pathogens. Together, these studies enhance our understanding of Yersinia pathogenesis and highlight the ongoing battle between host and microbe. Copyright © 2015 Elsevier Ltd. All rights reserved.

Oncogenic Properties of Apoptotic Tumor Cells in Aggressive B Cell Lymphoma

PubMed Central

Ford, Catriona A.; Petrova, Sofia; Pound, John D.; Voss, Jorine J.L.P.; Melville, Lynsey; Paterson, Margaret; Farnworth, Sarah L.; Gallimore, Awen M.; Cuff, Simone; Wheadon, Helen; Dobbin, Edwina; Ogden, Carol Anne; Dumitriu, Ingrid E.; Dunbar, Donald R.; Murray, Paul G.; Ruckerl, Dominik; Allen, Judith E.; Hume, David A.; van Rooijen, Nico; Goodlad, John R.; Freeman, Tom C.; Gregory, Christopher D.

2015-01-01

Summary Background Cells undergoing apoptosis are known to modulate their tissue microenvironments. By acting on phagocytes, notably macrophages, apoptotic cells inhibit immunological and inflammatory responses and promote trophic signaling pathways. Paradoxically, because of their potential to cause death of tumor cells and thereby militate against malignant disease progression, both apoptosis and tumor-associated macrophages (TAMs) are often associated with poor prognosis in cancer. We hypothesized that, in progression of malignant disease, constitutive loss of a fraction of the tumor cell population through apoptosis could yield tumor-promoting effects. Results Here, we demonstrate that apoptotic tumor cells promote coordinated tumor growth, angiogenesis, and accumulation of TAMs in aggressive B cell lymphomas. Through unbiased “in situ transcriptomics” analysis—gene expression profiling of laser-captured TAMs to establish their activation signature in situ—we show that these cells are activated to signal via multiple tumor-promoting reparatory, trophic, angiogenic, tissue remodeling, and anti-inflammatory pathways. Our results also suggest that apoptotic lymphoma cells help drive this signature. Furthermore, we demonstrate that, upon induction of apoptosis, lymphoma cells not only activate expression of the tumor-promoting matrix metalloproteinases MMP2 and MMP12 in macrophages but also express and process these MMPs directly. Finally, using a model of malignant melanoma, we show that the oncogenic potential of apoptotic tumor cells extends beyond lymphoma. Conclusions In addition to its profound tumor-suppressive role, apoptosis can potentiate cancer progression. These results have important implications for understanding the fundamental biology of cell death, its roles in malignant disease, and the broader consequences of apoptosis-inducing anti-cancer therapy. PMID:25702581

Guidance signalling regulates leading edge behaviour during collective cell migration of cardiac cells in Drosophila.

PubMed

Raza, Qanber; Jacobs, J Roger

2016-11-15

Collective cell migration is the coordinated movement of cells, which organize tissues during morphogenesis, repair and some cancers. The motile cell membrane of the advancing front in collective cell migration is termed the Leading Edge. The embryonic development of the vertebrate and Drosophila hearts are both characterized by the coordinated medial migration of a bilateral cluster of mesodermal cells. In Drosophila, the cardioblasts form cohesive bilateral rows that migrate collectively as a unit towards the dorsal midline to form the dorsal vessel. We have characterized the collective cell migration of cardioblasts as an in vivo quantitative model to study the behaviour of the Leading Edge. We investigated whether guidance signalling through Slit and Netrin pathways plays a role in cell migration during heart development. Through time-lapse imaging and quantitative assessment of migratory behaviour of the cardioblasts in loss-of-function mutants, we demonstrate that both Slit and Netrin mediated signals are autonomously and concomitantly required to maximize migration velocity, filopodial and lamellipodial activities. Additionally, we show that another Slit and Netrin receptor, Dscam1, the role of which during heart development was previously unknown, is required for both normal migration of cardioblasts and luminal expansion. Leading edge behaviour analysis revealed a dosage dependent genetic interaction between Slit and Netrin receptors suggesting that downstream signalling through these receptors converge on a common output that increases leading edge activity of the cardioblasts. Finally, we found that guidance signalling maintains the balance between epithelial and mesenchymal characteristics of the migrating cardioblasts. Copyright © 2016 Elsevier Inc. All rights reserved.

CXCR4 Chemokine Receptor Signaling Induces Apoptosis in Acute Myeloid Leukemia Cells via Regulation of the Bcl-2 Family Members Bcl-XL, Noxa, and Bak*

PubMed Central

Kremer, Kimberly N.; Peterson, Kevin L.; Schneider, Paula A.; Meng, X. Wei; Dai, Haiming; Hess, Allan D.; Smith, B. Douglas; Rodriguez-Ramirez, Christie; Karp, Judith E.; Kaufmann, Scott H.; Hedin, Karen E.

2013-01-01

The CXCR4 chemokine receptor promotes survival of many different cell types. Here, we describe a previously unsuspected role for CXCR4 as a potent inducer of apoptosis in acute myeloid leukemia (AML) cell lines and a subset of clinical AML samples. We show that SDF-1, the sole ligand for CXCR4, induces the expected migration and ERK activation in the KG1a AML cell line transiently overexpressing CXCR4, but ERK activation did not lead to survival. Instead, SDF-1 treatment led via a CXCR4-dependent mechanism to apoptosis, as evidenced by increased annexin V staining, condensation of chromatin, and cleavage of both procaspase-3 and PARP. This SDF-1-induced death pathway was partially inhibited by hypoxia, which is often found in the bone marrow of AML patients. SDF-1-induced apoptosis was inhibited by dominant negative procaspase-9 but not by inhibition of caspase-8 activation, implicating the intrinsic apoptotic pathway. Further analysis showed that this pathway was activated by multiple mechanisms, including up-regulation of Bak at the level of mRNA and protein, stabilization of the Bak activator Noxa, and down-regulation of antiapoptotic Bcl-XL. Furthermore, adjusting expression levels of Bak, Bcl-XL, or Noxa individually altered the level of apoptosis in AML cells, suggesting that the combined modulation of these family members by SDF-1 coordinates their interplay to produce apoptosis. Thus, rather than mediating survival, SDF-1 may be a means to induce apoptosis of CXCR4-expressing AML cells directly in the SDF-1-rich bone marrow microenvironment if the survival cues of the bone marrow are disrupted. PMID:23798675

Chemical Inactivation of the Cinnamate 4-Hydroxylase Allows for the Accumulation of Salicylic Acid in Elicited Cells1

PubMed Central

Schoch, Guillaume A.; Nikov, Georgi N.; Alworth, William L.; Werck-Reichhart, Danièle

2002-01-01

The cinnamate (CA) 4-hydroxylase (C4H) is a cytochrome P450 that catalyzes the second step of the main phenylpropanoid pathway, leading to the synthesis of lignin, pigments, and many defense molecules. Salicylic acid (SA) is an essential trigger of plant disease resistance. Some plant species can synthesize SA from CA by a mechanism not yet understood. A set of specific inhibitors of the C4H, including competitive, tight-binding, mechanism-based irreversible, and quasi-irreversible inhibitors have been developed with the main objective to redirect cinnamic acid to the synthesis of SA. Competitive inhibitors such as 2-hydroxy-1-naphthoic acid and the heme-coordinating compound 3-(4-pyridyl)-acrylic acid allowed strong inhibition of C4H activity in a tobacco (Nicotiana tabacum cv Bright Yellow [BY]) cell suspension culture. This inhibition was however rapidly relieved either because of substrate accumulation or because of inhibitor metabolism. Substrate analogs bearing a methylenedioxo function such as piperonylic acid (PIP) or a terminal acetylene such as 4-propynyloxybenzoic acid (4PB), 3-propynyloxybenzoic acid, and 4-propynyloxymethylbenzoic acid are potent mechanism-based inactivators of the C4H. PIP and 4PB, the best inactivators in vitro, were also efficient inhibitors of the enzyme in BY cells. Inhibition was not reversed 46 h after cell treatment. Cotreatment of BY cells with the fungal elicitor β-megaspermin and PIP or 4PB led to a dramatic increase in SA accumulation. PIP and 4PB do not trigger SA accumulation in nonelicited cells in which the SA biosynthetic pathway is not activated. Mechanism-based C4H inactivators, thus, are promising tools for the elucidation of the CA-derived SA biosynthetic pathway and for the potentiation of plant defense. PMID:12376665

Cell Cycle Regulation of Stem Cells by MicroRNAs.

PubMed

Mens, Michelle M J; Ghanbari, Mohsen

2018-06-01

MicroRNAs (miRNAs) are a class of small non-coding RNA molecules involved in the regulation of gene expression. They are involved in the fine-tuning of fundamental biological processes such as proliferation, differentiation, survival and apoptosis in many cell types. Emerging evidence suggests that miRNAs regulate critical pathways involved in stem cell function. Several miRNAs have been suggested to target transcripts that directly or indirectly coordinate the cell cycle progression of stem cells. Moreover, previous studies have shown that altered expression levels of miRNAs can contribute to pathological conditions, such as cancer, due to the loss of cell cycle regulation. However, the precise mechanism underlying miRNA-mediated regulation of cell cycle in stem cells is still incompletely understood. In this review, we discuss current knowledge of miRNAs regulatory role in cell cycle progression of stem cells. We describe how specific miRNAs may control cell cycle associated molecules and checkpoints in embryonic, somatic and cancer stem cells. We further outline how these miRNAs could be regulated to influence cell cycle progression in stem cells as a potential clinical application.

Molecular and physiological evidence of genetic assimilation to high CO2 in the marine nitrogen fixer Trichodesmium.

PubMed

Walworth, Nathan G; Lee, Michael D; Fu, Fei-Xue; Hutchins, David A; Webb, Eric A

2016-11-22

Most investigations of biogeochemically important microbes have focused on plastic (short-term) phenotypic responses in the absence of genetic change, whereas few have investigated adaptive (long-term) responses. However, no studies to date have investigated the molecular progression underlying the transition from plasticity to adaptation under elevated CO 2 for a marine nitrogen-fixer. To address this gap, we cultured the globally important cyanobacterium Trichodesmium at both low and high CO 2 for 4.5 y, followed by reciprocal transplantation experiments to test for adaptation. Intriguingly, fitness actually increased in all high-CO 2 adapted cell lines in the ancestral environment upon reciprocal transplantation. By leveraging coordinated phenotypic and transcriptomic profiles, we identified expression changes and pathway enrichments that rapidly responded to elevated CO 2 and were maintained upon adaptation, providing strong evidence for genetic assimilation. These candidate genes and pathways included those involved in photosystems, transcriptional regulation, cell signaling, carbon/nitrogen storage, and energy metabolism. Conversely, significant changes in specific sigma factor expression were only observed upon adaptation. These data reveal genetic assimilation as a potentially adaptive response of Trichodesmium and importantly elucidate underlying metabolic pathways paralleling the fixation of the plastic phenotype upon adaptation, thereby contributing to the few available data demonstrating genetic assimilation in microbial photoautotrophs. These molecular insights are thus critical for identifying pathways under selection as drivers in plasticity and adaptation.

Suppression of the Escherichia coli dnaA46 mutation by changes in the activities of the pyruvate-acetate node links DNA replication regulation to central carbon metabolism.

PubMed

Tymecka-Mulik, Joanna; Boss, Lidia; Maciąg-Dorszyńska, Monika; Matias Rodrigues, João F; Gaffke, Lidia; Wosinski, Anna; Cech, Grzegorz M; Szalewska-Pałasz, Agnieszka; Węgrzyn, Grzegorz; Glinkowska, Monika

2017-01-01

To ensure faithful transmission of genetic material to progeny cells, DNA replication is tightly regulated, mainly at the initiation step. Escherichia coli cells regulate the frequency of initiation according to growth conditions. Results of the classical, as well as the latest studies, suggest that the DNA replication in E. coli starts at a predefined, constant cell volume per chromosome but the mechanisms coordinating DNA replication with cell growth are still not fully understood. Results of recent investigations have revealed a role of metabolic pathway proteins in the control of cell division and a direct link between metabolism and DNA replication has also been suggested both in Bacillus subtilis and E. coli cells. In this work we show that defects in the acetate overflow pathway suppress the temperature-sensitivity of a defective replication initiator-DnaA under acetogenic growth conditions. Transcriptomic and metabolic analyses imply that this suppression is correlated with pyruvate accumulation, resulting from alterations in the pyruvate dehydrogenase (PDH) activity. Consequently, deletion of genes encoding the pyruvate dehydrogenase subunits likewise resulted in suppression of the thermal-sensitive growth of the dnaA46 strain. We propose that the suppressor effect may be directly related to the PDH complex activity, providing a link between an enzyme of the central carbon metabolism and DNA replication.

Control of epithelial immune-response genes and implications for airway immunity and inflammation.

PubMed

Holtzman, M J; Look, D C; Sampath, D; Castro, M; Koga, T; Walter, M J

1998-01-01

A major goal of our research is to understand how immune cells (especially T cells) infiltrate the pulmonary airway during host defense and inflammatory disease (especially asthma). In that context, we have proposed that epithelial cells lining the airway provide critical biochemical signals for immune-cell influx and activation and that this epithelial-immune cell interaction is a critical feature of airway inflammation and hyperreactivity. In this brief report, we describe our progress in defining a subset of epithelial immune-response genes the expression of which is coordinated for viral defense both directly in response to replicating virus and indirectly under the control of a specific interferon-gamma signal transduction pathway featuring the Stat1 transcription factor as a critical relay signal between cytoplasm and nucleus. Unexpectedly, the same pathway is also activated during asthmatic airway inflammation in a setting where there is no apparent infection and no increase in interferon-gamma levels. The findings provide the first evidence of an overactive Stat1-dependent gene network in asthmatic airways and a novel molecular link between mucosal immunity and inflammation. The findings also offer the possibility that overactivity of Stat1-dependent genes might augment a subsequent T helper cell (Th1)-type response to virus or might combine with a heightened Th2-type response to allergen to account for more severe exacerbations of asthma.

Involvement of complex sphingolipids and phosphatidylserine in endosomal trafficking in yeast Saccharomyces cerevisiae.

PubMed

Tani, Motohiro; Kuge, Osamu

2012-12-01

Sphingolipids play critical roles in many physiologically important events in the yeast Saccharomyces cerevisiae. In this study, we found that csg2Δ mutant cells defective in the synthesis of mannosylinositol phosphorylceramide exhibited abnormal intracellular accumulation of an exocytic v-SNARE, Snc1, under phosphatidylserine synthase gene (PSS1)-repressive conditions, although in wild-type cells, Snc1 was known to cycle between plasma membranes and the late Golgi via post-Golgi endosomes. The mislocalized Snc1 was co-localized with an endocytic marker dye, FM4-64, upon labelling for a short time. The abnormal distribution of Snc1 was suppressed by deletion of GYP2 encoding a GTPase-activating protein that negatively regulates endosomal vesicular trafficking, or expression of GTP-restricted form of Ypt32 GTPase. Furthermore, an endocytosis-deficient mutant of Snc1 was localized to plasma membranes in PSS1-repressed csg2Δ mutant cells as well as wild-type cells. Thus, the PSS1-repressed csg2Δ mutant cells were indicated to be defective in the trafficking of Snc1 from post-Golgi endosomes to the late Golgi. In contrast, the vesicular trafficking pathways via pre-vacuolar endosomes in the PSS1-repressed csg2Δ mutant cells seemed to be normal. These results suggested that specific complex sphingolipids and phosphatidylserine are co-ordinately involved in specific vesicular trafficking pathway. © 2012 Blackwell Publishing Ltd.

Coordinated regulation of intracellular pH by two glucose-sensing pathways in yeast.

PubMed

Isom, Daniel G; Page, Stephani C; Collins, Leonard B; Kapolka, Nicholas J; Taghon, Geoffrey J; Dohlman, Henrik G

2018-02-16

The yeast Saccharomyces cerevisiae employs multiple pathways to coordinate sugar availability and metabolism. Glucose and other sugars are detected by a G protein-coupled receptor, Gpr1, as well as a pair of transporter-like proteins, Rgt2 and Snf3. When glucose is limiting, however, an ATP-driven proton pump (Pma1) is inactivated, leading to a marked decrease in cytoplasmic pH. Here we determine the relative contribution of the two sugar-sensing pathways to pH regulation. Whereas cytoplasmic pH is strongly dependent on glucose abundance and is regulated by both glucose-sensing pathways, ATP is largely unaffected and therefore cannot account for the changes in Pma1 activity. These data suggest that the pH is a second messenger of the glucose-sensing pathways. We show further that different sugars differ in their ability to control cellular acidification, in the manner of inverse agonists. We conclude that the sugar-sensing pathways act via Pma1 to invoke coordinated changes in cellular pH and metabolism. More broadly, our findings support the emerging view that cellular systems have evolved the use of pH signals as a means of adapting to environmental stresses such as those caused by hypoxia, ischemia, and diabetes. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Cellular localization of the activated EGFR determines its effect on cell growth in MDA-MB-468 cells

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

Hyatt, Dustin C.; Ceresa, Brian P.

2008-11-01

The epidermal growth factor (EGF) receptor (EGFR) is a ubiquitously expressed receptor tyrosine kinase that regulates diverse cell functions that are dependent upon cell type, the presence of downstream effectors, and receptor density. In addition to activating biochemical pathways, ligand stimulation causes the EGFR to enter the cell via clathrin-coated pits. Endocytic trafficking influences receptor signaling by controlling the duration of EGFR phosphorylation and coordinating the receptor's association with downstream effectors. To better understand the individual contributions of cell surface and cytosolic EGFRs on cell physiology, we used EGF that was conjugated to 900 nm polystyrene beads (EGF-beads). EGF-beads canmore » stimulate the EGFR and retain the activated receptor at the plasma membrane. In MDA-MB-468 cells, a breast cancer cell line that over-expresses the EGFR, only internalized, activated EGFRs stimulate caspase-3 and induce cell death. Conversely, signaling cascades triggered from activated EGFR retained at the cell surface inhibit caspase-3 and promote cell proliferation. Thus, through endocytosis, the activated EGFR can differentially regulate cell growth in MDA-MB-468 cells.« less

Neural control of the kidney: functionally specific renal sympathetic nerve fibers.

PubMed

DiBona, G F

2000-11-01

The sympathetic nervous system provides differentiated regulation of the functions of various organs. This differentiated regulation occurs via mechanisms that operate at multiple sites within the classic reflex arc: peripherally at the level of afferent input stimuli to various reflex pathways, centrally at the level of interconnections between various central neuron pools, and peripherally at the level of efferent fibers targeted to various effectors within the organ. In the kidney, increased renal sympathetic nerve activity regulates the functions of the intrarenal effectors: the tubules, the blood vessels, and the juxtaglomerular granular cells. This enables a physiologically appropriate coordination between the circulatory, filtration, reabsorptive, excretory, and renin secretory contributions to overall renal function. Anatomically, each of these effectors has a dual pattern of innervation consisting of a specific and selective innervation by unmyelinated slowly conducting C-type renal sympathetic nerve fibers in addition to an innervation that is shared among all the effectors. This arrangement permits the maximum flexibility in the coordination of physiologically appropriate responses of the tubules, the blood vessels, and the juxtaglomerular granular cells to a variety of homeostatic requirements.

Functionally specific renal sympathetic nerve fibers: role in cardiovascular regulation.

PubMed

DiBona, G F

2001-06-01

The sympathetic nervous system provides differentiated regulation of the functions of various organs. This differentiated regulation occurs through mechanisms that operate at multiple sites within the classic reflex arc: peripherally at the level of afferent input stimuli to various reflex pathways, centrally at the level of interconnections between various central neuron pools, and peripherally at the level of efferent fibers targeted to various effectors within the organ. In the kidney, increased renal sympathetic nerve activity regulates the functions of the intrarenal effectors: the tubules, the blood vessels, and the juxtaglomerular granular cells. This enables a physiologically appropriate coordination between the circulatory, filtration, reabsorptive, excretory, and renin secretory contributions to overall renal function. Anatomically, each of these effectors has a dual pattern of innervation consisting of a specific and selective innervation by unmyelinated slowly conducting C-type renal sympathetic nerve fibers and an innervation that is shared among all the effectors. This arrangement facilitates maximum flexibility in the coordination of the tubules, the blood vessels, and the juxtaglomerular granular cells so as to produce physiologically appropriate responses to a variety of homeostatic requirements.

Loss of Dlg-1 in the Mouse Lens Impairs Fibroblast Growth Factor Receptor Signaling

PubMed Central

Lee, SungKyoung; Griep, Anne E.

2014-01-01

Coordination of cell proliferation, differentiation and survival is essential for normal development and maintenance of tissues in the adult organism. Growth factor receptor tyrosine kinase signaling pathways and planar cell polarity pathways are two regulators of many developmental processes. We have previously shown through analysis of mice conditionally null in the lens for the planar cell polarity gene (PCP), Dlg-1, that Dlg-1 is required for fiber differentiation. Herein, we asked if Dlg-1 is a regulator of the Fibroblast growth factor receptor (Fgfr) signaling pathway, which is known to be required for fiber cell differentiation. Western blot analysis of whole fiber cell extracts from control and Dlg-1 deficient lenses showed that levels of the Fgfr signaling intermediates pErk, pAkt, and pFrs2α, the Fgfr target, Erm, and the fiber cell specific protein, Mip26, were reduced in the Dlg-1 deficient fiber cells. The levels of Fgfr2 were decreased in Dlg-1 deficient lenses compared to controls. Conversely, levels of Fgfr1 in Dlg-1 deficient lenses were increased compared to controls. The changes in Fgfr levels were found to be specifically in the triton insoluble, cytoskeletal associated fraction of Dlg-1 deficient lenses. Immunofluorescent staining of lenses from E13.5 embryos showed that expression levels of pErk were reduced in the transition zone, a region of the lens that exhibits PCP, in the Dlg-1 deficient lenses as compared to controls. In control lenses, immunofluorescent staining for Fgfr2 was observed in the epithelium, transition zone and fibers. By E13.5, the intensity of staining for Fgfr2 was reduced in these regions of the Dlg-1 deficient lenses. Thus, loss of Dlg-1 in the lens impairs Fgfr signaling and leads to altered levels of Fgfrs, suggesting that Dlg-1 is a modulator of Fgfr signaling pathway at the level of the receptors and that Dlg-1 regulates fiber cell differentiation through its role in PCP. PMID:24824078

Control of DEMETER DNA demethylase gene transcription in male and female gamete companion cells in Arabidopsis thaliana.

PubMed

Park, Jin-Sup; Frost, Jennifer M; Park, Kyunghyuk; Ohr, Hyonhwa; Park, Guen Tae; Kim, Seohyun; Eom, Hyunjoo; Lee, Ilha; Brooks, Janie S; Fischer, Robert L; Choi, Yeonhee

2017-02-21

The DEMETER (DME) DNA glycosylase initiates active DNA demethylation via the base-excision repair pathway and is vital for reproduction in Arabidopsis thaliana DME-mediated DNA demethylation is preferentially targeted to small, AT-rich, and nucleosome-depleted euchromatic transposable elements, influencing expression of adjacent genes and leading to imprinting in the endosperm. In the female gametophyte, DME expression and subsequent genome-wide DNA demethylation are confined to the companion cell of the egg, the central cell. Here, we show that, in the male gametophyte, DME expression is limited to the companion cell of sperm, the vegetative cell, and to a narrow window of time: immediately after separation of the companion cell lineage from the germline. We define transcriptional regulatory elements of DME using reporter genes, showing that a small region, which surprisingly lies within the DME gene, controls its expression in male and female companion cells. DME expression from this minimal promoter is sufficient to rescue seed abortion and the aberrant DNA methylome associated with the null dme-2 mutation. Within this minimal promoter, we found short, conserved enhancer sequences necessary for the transcriptional activities of DME and combined predicted binding motifs with published transcription factor binding coordinates to produce a list of candidate upstream pathway members in the genetic circuitry controlling DNA demethylation in gamete companion cells. These data show how DNA demethylation is regulated to facilitate endosperm gene imprinting and potential transgenerational epigenetic regulation, without subjecting the germline to potentially deleterious transposable element demethylation.

A conserved signaling network monitors delivery of sphingolipids to the plasma membrane in budding yeast

PubMed Central

Clarke, Jesse; Dephoure, Noah; Horecka, Ira; Gygi, Steven; Kellogg, Douglas

2017-01-01

In budding yeast, cell cycle progression and ribosome biogenesis are dependent on plasma membrane growth, which ensures that events of cell growth are coordinated with each other and with the cell cycle. However, the signals that link the cell cycle and ribosome biogenesis to membrane growth are poorly understood. Here we used proteome-wide mass spectrometry to systematically discover signals associated with membrane growth. The results suggest that membrane trafficking events required for membrane growth generate sphingolipid-dependent signals. A conserved signaling network appears to play an essential role in signaling by responding to delivery of sphingolipids to the plasma membrane. In addition, sphingolipid-dependent signals control phosphorylation of protein kinase C (Pkc1), which plays an essential role in the pathways that link the cell cycle and ribosome biogenesis to membrane growth. Together these discoveries provide new clues as to how growth-­dependent signals control cell growth and the cell cycle. PMID:28794263

A Non-Cell-Autonomous Role of BEC-1/BECN1/Beclin1 in Coordinating Cell-Cycle Progression and Stem Cell Proliferation during Germline Development.

PubMed

Ames, Kristina; Da Cunha, Dayse S; Gonzalez, Brenda; Konta, Marina; Lin, Feng; Shechter, Gabriel; Starikov, Lev; Wong, Sara; Bülow, Hannes E; Meléndez, Alicia

2017-03-20

The decision of stem cells to proliferate and differentiate is finely controlled. The Caenorhabditis elegans germline provides a tractable system for studying the mechanisms that control stem cell proliferation and homeostasis [1-4]. Autophagy is a conserved cellular recycling process crucial for cellular homeostasis in many different contexts [5], but its function in germline stem cell proliferation remains poorly understood. Here, we describe a function for autophagy in germline stem cell proliferation. We found that autophagy genes such as bec-1/BECN1/Beclin1, atg-16.2/ATG16L, atg-18/WIPI1/2, and atg-7/ATG7 are required for the late larval expansion of germline stem cell progenitors in the C. elegans gonad. We further show that BEC-1/BECN1/Beclin1 acts independently of the GLP-1/Notch or DAF-7/TGF-β pathways but together with the DAF-2/insulin IGF-1 receptor (IIR) signaling pathway to promote germline stem cell proliferation. Similar to DAF-2/IIR, BEC-1/BECN1/Beclin1, ATG-18/WIPI1/2, and ATG-16.2/ATG16L all promote cell-cycle progression and are negatively regulated by the phosphatase and tensin homolog DAF-18/PTEN. However, whereas BEC-1/BECN1/Beclin1 acts through the transcriptional regulator SKN-1/Nrf1, ATG-18/WIPI1/2 and ATG-16.2/ATG16L exert their function through the DAF-16/FOXO transcription factor. In contrast, ATG-7 functions in concert with the DAF-7/TGF-β pathway to promote germline proliferation and is not required for cell-cycle progression. Finally, we report that BEC-1/BECN1/Beclin1 functions non-cell-autonomously to facilitate cell-cycle progression and stem cell proliferation. Our findings demonstrate a novel non-autonomous role for BEC-1/BECN1/Beclin1 in the control of stem cell proliferation and cell-cycle progression, which may have implications for the understanding and development of therapies against malignant cell growth in the future. Copyright © 2017 Elsevier Ltd. All rights reserved.

Rupturing the hemi-fission intermediate in membrane fission under tension: Reaction coordinates, kinetic pathways, and free-energy barriers

NASA Astrophysics Data System (ADS)

Zhang, Guojie; Müller, Marcus

2017-08-01

Membrane fission is a fundamental process in cells, involved inter alia in endocytosis, intracellular trafficking, and virus infection. Its underlying molecular mechanism, however, is only incompletely understood. Recently, experiments and computer simulation studies have revealed that dynamin-mediated membrane fission is a two-step process that proceeds via a metastable hemi-fission intermediate (or wormlike micelle) formed by dynamin's constriction. Importantly, this hemi-fission intermediate is remarkably metastable, i.e., its subsequent rupture that completes the fission process does not occur spontaneously but requires additional, external effects, e.g., dynamin's (unknown) conformational changes or membrane tension. Using simulations of a coarse-grained, implicit-solvent model of lipid membranes, we investigate the molecular mechanism of rupturing the hemi-fission intermediate, such as its pathway, the concomitant transition states, and barriers, as well as the role of membrane tension. The membrane tension is controlled by the chemical potential of the lipids, and the free-energy landscape as a function of two reaction coordinates is obtained by grand canonical Wang-Landau sampling. Our results show that, in the course of rupturing, the hemi-fission intermediate undergoes a "thinning → local pinching → rupture/fission" pathway, with a bottle-neck-shaped cylindrical micelle as a transition state. Although an increase of membrane tension facilitates the fission process by reducing the corresponding free-energy barrier, for biologically relevant tensions, the free-energy barriers still significantly exceed the thermal energy scale kBT.

Rupturing the hemi-fission intermediate in membrane fission under tension: Reaction coordinates, kinetic pathways, and free-energy barriers.

PubMed

Zhang, Guojie; Müller, Marcus

2017-08-14

Membrane fission is a fundamental process in cells, involved inter alia in endocytosis, intracellular trafficking, and virus infection. Its underlying molecular mechanism, however, is only incompletely understood. Recently, experiments and computer simulation studies have revealed that dynamin-mediated membrane fission is a two-step process that proceeds via a metastable hemi-fission intermediate (or wormlike micelle) formed by dynamin's constriction. Importantly, this hemi-fission intermediate is remarkably metastable, i.e., its subsequent rupture that completes the fission process does not occur spontaneously but requires additional, external effects, e.g., dynamin's (unknown) conformational changes or membrane tension. Using simulations of a coarse-grained, implicit-solvent model of lipid membranes, we investigate the molecular mechanism of rupturing the hemi-fission intermediate, such as its pathway, the concomitant transition states, and barriers, as well as the role of membrane tension. The membrane tension is controlled by the chemical potential of the lipids, and the free-energy landscape as a function of two reaction coordinates is obtained by grand canonical Wang-Landau sampling. Our results show that, in the course of rupturing, the hemi-fission intermediate undergoes a "thinning → local pinching → rupture/fission" pathway, with a bottle-neck-shaped cylindrical micelle as a transition state. Although an increase of membrane tension facilitates the fission process by reducing the corresponding free-energy barrier, for biologically relevant tensions, the free-energy barriers still significantly exceed the thermal energy scale k B T.

A network of conserved formins, regulated by the guanine exchange factor EXC-5 and the GTPase CDC-42, modulates tubulogenesis in vivo.

PubMed

Shaye, Daniel D; Greenwald, Iva

2016-11-15

The C. elegans excretory cell (EC) is a powerful model for tubulogenesis, a conserved process that requires precise cytoskeletal regulation. EXC-6, an ortholog of the disease-associated formin INF2, coordinates cell outgrowth and lumen formation during EC tubulogenesis by regulating F-actin at the tip of the growing canal and the dynamics of basolateral microtubules. EXC-6 functions in parallel with EXC-5/FGD, a predicted activator of the Rho GTPase Cdc42. Here, we identify the parallel pathway: EXC-5 functions through CDC-42 to regulate two other formins: INFT-2, another INF2 ortholog, and CYK-1, the sole ortholog of the mammalian diaphanous (mDia) family of formins. We show that INFT-2 promotes F-actin accumulation in the EC, and that CYK-1 inhibits INFT-2 to regulate F-actin levels and EXC-6-promoted outgrowth. As INF2 and mDia physically interact and cross-regulate in cultured cells, our work indicates that a conserved EXC-5-CDC-42 pathway modulates this regulatory interaction and that it is functionally important in vivo during tubulogenesis. © 2016. Published by The Company of Biologists Ltd.

Evidence of isotopic fractionation of natural uranium in cultured human cells

NASA Astrophysics Data System (ADS)

Paredes, Eduardo; Avazeri, Emilie; Malard, Véronique; Vidaud, Claude; Reiller, Pascal E.; Ortega, Richard; Nonell, Anthony; Isnard, Hélène; Chartier, Frédéric; Bresson, Carole

2016-12-01

The study of the isotopic fractionation of endogen elements and toxic heavy metals in living organisms for biomedical applications, and for metabolic and toxicological studies, is a cutting-edge research topic. This paper shows that human neuroblastoma cells incorporated small amounts of uranium (U) after exposure to 10 µM natural U, with preferential uptake of the 235U isotope with regard to 238U. Efforts were made to develop and then validate a procedure for highly accurate n(238U)/n(235U) determinations in microsamples of cells. We found that intracellular U is enriched in 235U by 0.38 ± 0.13‰ (2σ, n = 7) relative to the exposure solutions. These in vitro experiments provide clues for the identification of biological processes responsible for uranium isotopic fractionation and link them to potential U incorporation pathways into neuronal cells. Suggested incorporation processes are a kinetically controlled process, such as facilitated transmembrane diffusion, and the uptake through a high-affinity uranium transport protein involving the modification of the uranyl (UO22+) coordination sphere. These findings open perspectives on the use of isotopic fractionation of metals in cellular models, offering a probe to track uptake/transport pathways and to help decipher associated cellular metabolic processes.

Evidence of isotopic fractionation of natural uranium in cultured human cells

PubMed Central

Paredes, Eduardo; Avazeri, Emilie; Malard, Véronique; Vidaud, Claude; Reiller, Pascal E.; Ortega, Richard; Nonell, Anthony; Isnard, Hélène; Chartier, Frédéric; Bresson, Carole

2016-01-01

The study of the isotopic fractionation of endogen elements and toxic heavy metals in living organisms for biomedical applications, and for metabolic and toxicological studies, is a cutting-edge research topic. This paper shows that human neuroblastoma cells incorporated small amounts of uranium (U) after exposure to 10 µM natural U, with preferential uptake of the 235U isotope with regard to 238U. Efforts were made to develop and then validate a procedure for highly accurate n(238U)/n(235U) determinations in microsamples of cells. We found that intracellular U is enriched in 235U by 0.38 ± 0.13‰ (2σ, n = 7) relative to the exposure solutions. These in vitro experiments provide clues for the identification of biological processes responsible for uranium isotopic fractionation and link them to potential U incorporation pathways into neuronal cells. Suggested incorporation processes are a kinetically controlled process, such as facilitated transmembrane diffusion, and the uptake through a high-affinity uranium transport protein involving the modification of the uranyl (UO22+) coordination sphere. These findings open perspectives on the use of isotopic fractionation of metals in cellular models, offering a probe to track uptake/transport pathways and to help decipher associated cellular metabolic processes. PMID:27872304

Evidence of isotopic fractionation of natural uranium in cultured human cells.

PubMed

Paredes, Eduardo; Avazeri, Emilie; Malard, Véronique; Vidaud, Claude; Reiller, Pascal E; Ortega, Richard; Nonell, Anthony; Isnard, Hélène; Chartier, Frédéric; Bresson, Carole

2016-12-06

The study of the isotopic fractionation of endogen elements and toxic heavy metals in living organisms for biomedical applications, and for metabolic and toxicological studies, is a cutting-edge research topic. This paper shows that human neuroblastoma cells incorporated small amounts of uranium (U) after exposure to 10 µM natural U, with preferential uptake of the 235 U isotope with regard to 238 U. Efforts were made to develop and then validate a procedure for highly accurate n( 238 U)/n( 235 U) determinations in microsamples of cells. We found that intracellular U is enriched in 235 U by 0.38 ± 0.13‰ (2σ, n = 7) relative to the exposure solutions. These in vitro experiments provide clues for the identification of biological processes responsible for uranium isotopic fractionation and link them to potential U incorporation pathways into neuronal cells. Suggested incorporation processes are a kinetically controlled process, such as facilitated transmembrane diffusion, and the uptake through a high-affinity uranium transport protein involving the modification of the uranyl (UO 2 2+ ) coordination sphere. These findings open perspectives on the use of isotopic fractionation of metals in cellular models, offering a probe to track uptake/transport pathways and to help decipher associated cellular metabolic processes.

Murine Dishevelled 3 Functions in Redundant Pathways with Dishevelled 1 and 2 in Normal Cardiac Outflow Tract, Cochlea, and Neural Tube Development

PubMed Central

Etheridge, S. Leah; Ray, Saugata; Li, Shuangding; Hamblet, Natasha S.; Lijam, Nardos; Tsang, Michael; Greer, Joy; Kardos, Natalie; Wang, Jianbo; Sussman, Daniel J.; Chen, Ping; Wynshaw-Boris, Anthony

2008-01-01

Dishevelled (Dvl) proteins are important signaling components of both the canonical β-catenin/Wnt pathway, which controls cell proliferation and patterning, and the planar cell polarity (PCP) pathway, which coordinates cell polarity within a sheet of cells and also directs convergent extension cell (CE) movements that produce narrowing and elongation of the tissue. Three mammalian Dvl genes have been identified and the developmental roles of Dvl1 and Dvl2 were previously determined. Here, we identify the functions of Dvl3 in development and provide evidence of functional redundancy among the three murine Dvls. Dvl3 −/− mice died perinatally with cardiac outflow tract abnormalities, including double outlet right ventricle and persistent truncus arteriosis. These mutants also displayed a misorientated stereocilia in the organ of Corti, a phenotype that was enhanced with the additional loss of a single allele of the PCP component Vangl2/Ltap (LtapLp/+). Although neurulation appeared normal in both Dvl3 −/− and LtapLp/+ mutants, Dvl3 +/−;LtapLp/+ combined mutants displayed incomplete neural tube closure. Importantly, we show that many of the roles of Dvl3 are also shared by Dvl1 and Dvl2. More severe phenotypes were observed in Dvl3 mutants with the deficiency of another Dvl, and increasing Dvl dosage genetically with Dvl transgenes demonstrated the ability of Dvls to compensate for each other to enable normal development. Interestingly, global canonical Wnt signaling appeared largely unaffected in the double Dvl mutants, suggesting that low Dvl levels are sufficient for functional canonical Wnt signals. In summary, we demonstrate that Dvl3 is required for cardiac outflow tract development and describe its importance in the PCP pathway during neurulation and cochlea development. Finally, we establish several developmental processes in which the three Dvls are functionally redundant. PMID:19008950

Murine dishevelled 3 functions in redundant pathways with dishevelled 1 and 2 in normal cardiac outflow tract, cochlea, and neural tube development.

PubMed

Etheridge, S Leah; Ray, Saugata; Li, Shuangding; Hamblet, Natasha S; Lijam, Nardos; Tsang, Michael; Greer, Joy; Kardos, Natalie; Wang, Jianbo; Sussman, Daniel J; Chen, Ping; Wynshaw-Boris, Anthony

2008-11-01

Dishevelled (Dvl) proteins are important signaling components of both the canonical beta-catenin/Wnt pathway, which controls cell proliferation and patterning, and the planar cell polarity (PCP) pathway, which coordinates cell polarity within a sheet of cells and also directs convergent extension cell (CE) movements that produce narrowing and elongation of the tissue. Three mammalian Dvl genes have been identified and the developmental roles of Dvl1 and Dvl2 were previously determined. Here, we identify the functions of Dvl3 in development and provide evidence of functional redundancy among the three murine Dvls. Dvl3(-/-) mice died perinatally with cardiac outflow tract abnormalities, including double outlet right ventricle and persistent truncus arteriosis. These mutants also displayed a misorientated stereocilia in the organ of Corti, a phenotype that was enhanced with the additional loss of a single allele of the PCP component Vangl2/Ltap (LtapLp/+). Although neurulation appeared normal in both Dvl3(-/-) and LtapLp/+ mutants, Dvl3(+/-);LtapLp/+ combined mutants displayed incomplete neural tube closure. Importantly, we show that many of the roles of Dvl3 are also shared by Dvl1 and Dvl2. More severe phenotypes were observed in Dvl3 mutants with the deficiency of another Dvl, and increasing Dvl dosage genetically with Dvl transgenes demonstrated the ability of Dvls to compensate for each other to enable normal development. Interestingly, global canonical Wnt signaling appeared largely unaffected in the double Dvl mutants, suggesting that low Dvl levels are sufficient for functional canonical Wnt signals. In summary, we demonstrate that Dvl3 is required for cardiac outflow tract development and describe its importance in the PCP pathway during neurulation and cochlea development. Finally, we establish several developmental processes in which the three Dvls are functionally redundant.

Understanding the cancer cell phenotype beyond the limitations of current omics analyses.

PubMed

Moreno-Sánchez, Rafael; Saavedra, Emma; Gallardo-Pérez, Juan Carlos; Rumjanek, Franklin D; Rodríguez-Enríquez, Sara

2016-01-01

Efforts to understand the mechanistic principles driving cancer metabolism and proliferation have been lately governed by genomic, transcriptomic and proteomic studies. This paper analyzes the caveats of these approaches. As molecular biology's central dogma proposes a unidirectional flux of information from genes to mRNA to proteins, it has frequently been assumed that monitoring the changes in the gene sequences and in mRNA and protein contents is sufficient to explain complex cellular processes. Such a stance commonly disregards that post-translational modifications can alter the protein function/activity and also that regulatory mechanisms enter into action, to coordinate the protein activities of pathways/cellular processes, in order to keep the cellular homeostasis. Hence, the actual protein activities (as enzymes/transporters/receptors) and their regulatory mechanisms ultimately dictate the final outcomes of a pathway/cellular process. In this regard, it is here documented that the mRNA levels of many metabolic enzymes and transcriptional factors have no correlation with the respective protein contents and activities. The validity of current clinical mRNA-based tests and proposed metabolite biomarkers for cancer detection/prognosis is also discussed. Therefore, it is proposed that, to achieve a thorough understanding of the modifications undergone by proliferating cancer cells, it is mandatory to experimentally analyze the cellular processes at the functional level. This could be achieved (a) locally, by examining the actual protein activities in the cell and their kinetic properties (or at least kinetically characterize the most controlling steps of the pathway/cellular process); (b) systemically, by analyzing the main fluxes of the pathway/cellular process, and how they are modulated by metabolites, all which should contribute to comprehending the regulatory mechanisms that have been altered in cancer cells. By adopting a more holistic approach it may become possible to improve the design of therapeutic strategies that would target cancer cells more specifically. © 2015 FEBS.

Spatially Compact Neural Clusters in the Dorsal Striatum Encode Locomotion Relevant Information.

PubMed

Barbera, Giovanni; Liang, Bo; Zhang, Lifeng; Gerfen, Charles R; Culurciello, Eugenio; Chen, Rong; Li, Yun; Lin, Da-Ting

2016-10-05

An influential striatal model postulates that neural activities in the striatal direct and indirect pathways promote and inhibit movement, respectively. Normal behavior requires coordinated activity in the direct pathway to facilitate intended locomotion and indirect pathway to inhibit unwanted locomotion. In this striatal model, neuronal population activity is assumed to encode locomotion relevant information. Here, we propose a novel encoding mechanism for the dorsal striatum. We identified spatially compact neural clusters in both the direct and indirect pathways. Detailed characterization revealed similar cluster organization between the direct and indirect pathways, and cluster activities from both pathways were correlated with mouse locomotion velocities. Using machine-learning algorithms, cluster activities could be used to decode locomotion relevant behavioral states and locomotion velocity. We propose that neural clusters in the dorsal striatum encode locomotion relevant information and that coordinated activities of direct and indirect pathway neural clusters are required for normal striatal controlled behavior. VIDEO ABSTRACT. Published by Elsevier Inc.

Impaired light detection of the circadian clock in a zebrafish melanoma model

PubMed Central

Hamilton, Noémie; Diaz-de-Cerio, Natalia; Whitmore, David

2015-01-01

The circadian clock controls the timing of the cell cycle in healthy tissues and clock disruption is known to increase tumourigenesis. Melanoma is one of the most rapidly increasing forms of cancer and the precise molecular circadian changes that occur in a melanoma tumor are unknown. Using a melanoma zebrafish model, we have explored the molecular changes that occur to the circadian clock within tumors. We have found disruptions in melanoma clock gene expression due to a major impairment to the light input pathway, with a parallel loss of light-dependent activation of DNA repair genes. Furthermore, the timing of mitosis in tumors is perturbed, as well as the regulation of certain key cell cycle regulators, such that cells divide arhythmically. The inability to co-ordinate DNA damage repair and cell division is likely to promote further tumourigenesis and accelerate melanoma development. PMID:25832911

Impaired light detection of the circadian clock in a zebrafish melanoma model.

PubMed

Hamilton, Noémie; Diaz-de-Cerio, Natalia; Whitmore, David

2015-01-01

The circadian clock controls the timing of the cell cycle in healthy tissues and clock disruption is known to increase tumourigenesis. Melanoma is one of the most rapidly increasing forms of cancer and the precise molecular circadian changes that occur in a melanoma tumor are unknown. Using a melanoma zebrafish model, we have explored the molecular changes that occur to the circadian clock within tumors. We have found disruptions in melanoma clock gene expression due to a major impairment to the light input pathway, with a parallel loss of light-dependent activation of DNA repair genes. Furthermore, the timing of mitosis in tumors is perturbed, as well as the regulation of certain key cell cycle regulators, such that cells divide arhythmically. The inability to co-ordinate DNA damage repair and cell division is likely to promote further tumourigenesis and accelerate melanoma development.

Effector-triggered immunity: from pathogen perception to robust defense.

PubMed

Cui, Haitao; Tsuda, Kenichi; Parker, Jane E

2015-01-01

In plant innate immunity, individual cells have the capacity to sense and respond to pathogen attack. Intracellular recognition mechanisms have evolved to intercept perturbations by pathogen virulence factors (effectors) early in host infection and convert it to rapid defense. One key to resistance success is a polymorphic family of intracellular nucleotide-binding/leucine-rich-repeat (NLR) receptors that detect effector interference in different parts of the cell. Effector-activated NLRs connect, in various ways, to a conserved basal resistance network in order to transcriptionally boost defense programs. Effector-triggered immunity displays remarkable robustness against pathogen disturbance, in part by employing compensatory mechanisms within the defense network. Also, the mobility of some NLRs and coordination of resistance pathways across cell compartments provides flexibility to fine-tune immune outputs. Furthermore, a number of NLRs function close to the nuclear chromatin by balancing actions of defense-repressing and defense-activating transcription factors to program cells dynamically for effective disease resistance.

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

Molecular architecture and function of the SEA complex, a modulator of the TORC1 pathway.

PubMed

Algret, Romain; Fernandez-Martinez, Javier; Shi, Yi; Kim, Seung Joong; Pellarin, Riccardo; Cimermancic, Peter; Cochet, Emilie; Sali, Andrej; Chait, Brian T; Rout, Michael P; Dokudovskaya, Svetlana

2014-11-01

The TORC1 signaling pathway plays a major role in the control of cell growth and response to stress. Here we demonstrate that the SEA complex physically interacts with TORC1 and is an important regulator of its activity. During nitrogen starvation, deletions of SEA complex components lead to Tor1 kinase delocalization, defects in autophagy, and vacuolar fragmentation. TORC1 inactivation, via nitrogen deprivation or rapamycin treatment, changes cellular levels of SEA complex members. We used affinity purification and chemical cross-linking to generate the data for an integrative structure modeling approach, which produced a well-defined molecular architecture of the SEA complex and showed that the SEA complex comprises two regions that are structurally and functionally distinct. The SEA complex emerges as a platform that can coordinate both structural and enzymatic activities necessary for the effective functioning of the TORC1 pathway. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

p53 coordinates decidual sestrin 2/AMPK/mTORC1 signaling to govern parturition timing.

PubMed

Deng, Wenbo; Cha, Jeeyeon; Yuan, Jia; Haraguchi, Hirofumi; Bartos, Amanda; Leishman, Emma; Viollet, Benoit; Bradshaw, Heather B; Hirota, Yasushi; Dey, Sudhansu K

2016-08-01

Inflammation and oxidative stress are known risk factors for preterm birth (PTB); however, the mechanisms and pathways that influence this condition are not fully described. Previously, we showed that mTORC1 signaling is increased in mice harboring a uterine-specific deletion of transformation-related protein 53 (p53d/d mice), which exhibit premature decidual senescence that triggers spontaneous and inflammation-induced PTB. Treatment with the mTORC1 inhibitor rapamycin reduced the incidence of PTB in the p53d/d mice. Decidual senescence with heightened mTORC1 signaling is also a signature of human PTB. Here, we have identified an underlying mechanism for PTB and a potential therapeutic strategy for treating the condition. Treatment of pregnant p53d/d mice with either the antidiabetic drug metformin or the antioxidant resveratrol activated AMPK signaling and inhibited mTORC1 signaling in decidual cells. Both metformin and resveratrol protected against spontaneous and inflammation-induced PTB in p53d/d females. Using multiple approaches, we determined that p53 interacts with sestrins to coordinate an inverse relationship between AMPK and mTORC1 signaling that determines parturition timing. This signature was also observed in human decidual cells. Together, these results reveal that p53-dependent coordination of AMPK and mTORC1 signaling controls parturition timing and suggest that metformin and resveratrol have therapeutic potential to prevent PTB.

Investigating the pharmacodynamic and magnetic properties of pyrophosphate-bridged coordination complexes

NASA Astrophysics Data System (ADS)

Ikotun, Oluwatayo (Tayo) F.

The multidentate nature of pyrophosphate makes it an attractive ligand for complexation of metal cations. The participation of pyrophosphate in a variety of biological pathways and its metal catalyzed hydrolysis has driven our investigation into its coordination chemistry. We have successfully synthesized a library of binuclear pyrophosphate bridge coordination complexes. The problem of pyrophosphate hydrolysis to phosphate in the presence of divalent metal ions was overcome by incorporating capping ligands such as 1,10-phenanthroline and 2,2'-bipyridine prior to the addition of the pyrophosphate. The magnetic properties of these complexes was investigated and magneto-structural analysis was conducted. The biological abundance of pyrophosphate and the success of metal based drugs such as cisplatin, prompted our investigation of the cytotoxic properties of M(II) pyrophosphate dimeric complexes (where M(II) is CoII, CuII, and NiII) in adriamycin resistant human ovarian cancer cells. Thess compounds were found to exhibit toxicity in the nanomolar to picomolar range. We conducted in vitro stability studies and the mechanism of cytoxicity was elucidated by performing DNA mobility and binding assays, enzyme inhibition assays, and in vitro oxidative stress studies.

Arabidopsis JAGGED links floral organ patterning to tissue growth by repressing Kip-related cell cycle inhibitors.

PubMed

Schiessl, Katharina; Muiño, Jose M; Sablowski, Robert

2014-02-18

Plant morphogenesis requires coordinated cytoplasmic growth, oriented cell wall extension, and cell cycle progression, but it is debated which of these processes are primary drivers for tissue growth and directly targeted by developmental genes. Here, we used ChIP high-throughput sequencing combined with transcriptome analysis to identify global target genes of the Arabidopsis transcription factor JAGGED (JAG), which promotes growth of the distal region of floral organs. Consistent with the roles of JAG during organ initiation and subsequent distal organ growth, we found that JAG directly repressed genes involved in meristem development, such as CLAVATA1 and HANABA TARANU, and genes involved in the development of the basal region of shoot organs, such as BLADE ON PETIOLE 2 and the GROWTH REGULATORY FACTOR pathway. At the same time, JAG regulated genes involved in tissue polarity, cell wall modification, and cell cycle progression. In particular, JAG directly repressed KIP RELATED PROTEIN 4 (KRP4) and KRP2, which control the transition to the DNA synthesis phase (S-phase) of the cell cycle. The krp2 and krp4 mutations suppressed jag defects in organ growth and in the morphology of petal epidermal cells, showing that the interaction between JAG and KRP genes is functionally relevant. Our work reveals that JAG is a direct mediator between genetic pathways involved in organ patterning and cellular functions required for tissue growth, and it shows that a regulatory gene shapes plant organs by releasing a constraint on S-phase entry.

EBV and Apoptosis: The Viral Master Regulator of Cell Fate?

PubMed Central

Kelly, Gemma L.

2017-01-01

Epstein–Barr virus (EBV) was first discovered in cells from a patient with Burkitt lymphoma (BL), and is now known to be a contributory factor in 1–2% of all cancers, for which there are as yet, no EBV-targeted therapies available. Like other herpesviruses, EBV adopts a persistent latent infection in vivo and only rarely reactivates into replicative lytic cycle. Although latency is associated with restricted patterns of gene expression, genes are never expressed in isolation; always in groups. Here, we discuss (1) the ways in which the latent genes of EBV are known to modulate cell death, (2) how these mechanisms relate to growth transformation and lymphomagenesis, and (3) how EBV genes cooperate to coordinately regulate key cell death pathways in BL and lymphoblastoid cell lines (LCLs). Since manipulation of the cell death machinery is critical in EBV pathogenesis, understanding the mechanisms that underpin EBV regulation of apoptosis therefore provides opportunities for novel therapeutic interventions. PMID:29137176

Extracellular signals that define distinct and coexisting cell fates in Bacillus subtilis.

PubMed

López, Daniel; Kolter, Roberto

2010-03-01

The soil-dwelling bacterium Bacillus subtilis differentiates into distinct subpopulations of specialized cells that coexist within highly structured communities. The coordination and interplay between these cell types requires extensive extracellular communication driven mostly by sensing self-generated secreted signals. These extracellular signals activate a set of sensor kinases, which respond by phosphorylating three major regulatory proteins, Spo0A, DegU and ComA. Each phosphorylated regulator triggers a specific differentiation program while at the same time repressing other differentiation programs. This allows a cell to differentiate in response to a specific cue, even in the presence of other, possibly conflicting, signals. The sensor kinases involved respond to an eclectic group of extracellular signals, such as quorum-sensing molecules, natural products, temperature, pH or scarcity of nutrients. This article reviews the cascades of cell differentiation pathways that are triggered by sensing extracellular signals. We also present a tentative developmental model in which the diverse cell types sequentially differentiate to achieve the proper development of the bacterial community.

Examining the architecture of cellular computing through a comparative study with a computer

PubMed Central

Wang, Degeng; Gribskov, Michael

2005-01-01

The computer and the cell both use information embedded in simple coding, the binary software code and the quadruple genomic code, respectively, to support system operations. A comparative examination of their system architecture as well as their information storage and utilization schemes is performed. On top of the code, both systems display a modular, multi-layered architecture, which, in the case of a computer, arises from human engineering efforts through a combination of hardware implementation and software abstraction. Using the computer as a reference system, a simplistic mapping of the architectural components between the two is easily detected. This comparison also reveals that a cell abolishes the software–hardware barrier through genomic encoding for the constituents of the biochemical network, a cell's ‘hardware’ equivalent to the computer central processing unit (CPU). The information loading (gene expression) process acts as a major determinant of the encoded constituent's abundance, which, in turn, often determines the ‘bandwidth’ of a biochemical pathway. Cellular processes are implemented in biochemical pathways in parallel manners. In a computer, on the other hand, the software provides only instructions and data for the CPU. A process represents just sequentially ordered actions by the CPU and only virtual parallelism can be implemented through CPU time-sharing. Whereas process management in a computer may simply mean job scheduling, coordinating pathway bandwidth through the gene expression machinery represents a major process management scheme in a cell. In summary, a cell can be viewed as a super-parallel computer, which computes through controlled hardware composition. While we have, at best, a very fragmented understanding of cellular operation, we have a thorough understanding of the computer throughout the engineering process. The potential utilization of this knowledge to the benefit of systems biology is discussed. PMID:16849179

Function-specific intracellular signaling pathways downstream of heparin-binding EGF-like growth factor utilized by human trophoblasts.

PubMed

Jessmon, Philip; Kilburn, Brian A; Romero, Roberto; Leach, Richard E; Armant, D Randall

2010-05-01

Heparin-binding EGF-like growth factor (HBEGF) is expressed by trophoblast cells throughout gestation. First-trimester cytotrophoblast cells are protected from hypoxia-induced apoptosis because of the accumulation of HBEGF through a posttranscriptional autocrine mechanism. Exogenous application of HBEGF is cytoprotective in a hypoxia/reoxygenation (H/R) injury model and initiates trophoblast extravillous differentiation to an invasive phenotype. The downstream signaling pathways induced by HBEGF that mediate these various cellular activities were identified using two human first-trimester cytotrophoblast cell lines, HTR-8/SVneo and SW.71, with similar results. Recombinant HBEGF (1 nM) induced transient phosphorylation of MAPK3/1 (ERK), MAPK14 (p38), and AKT within 15 min and JNK after 1-2 h. To determine which downstream pathways regulate the various functions of HBEGF, cells were treated with specific inhibitors of the ERK upstream regulator MEK (U0126), the AKT upstream regulator phosphoinositide-3 (PI3)-kinase (LY294002), MAPK14 (SB203580), and JNK (SP600125), as well as with inactive structural analogues. Only SB203580 specifically prevented HBEGF-mediated rescue during H/R, while each inhibitor attenuated HBEGF-stimulated cell migration. Accumulation of HBEGF at reduced oxygen was blocked only by a combination of U0126, SB203580, and SP600125. We conclude that HBEGF advances trophoblast extravillous differentiation through coordinate activation of PI3 kinase, ERK, MAPK14, and JNK, while only MAPK14 is required for its antiapoptotic activity. Additionally, hypoxia induces an autocrine increase in HBEGF protein levels through MAPK14, JNK or ERK. These experiments reveal a complexity of the intracellular signaling circuitry that regulates trophoblast functions critical for implantation and placentation.

Function-Specific Intracellular Signaling Pathways Downstream of Heparin-Binding EGF-Like Growth Factor Utilized by Human Trophoblasts1

PubMed Central

Jessmon, Philip; Kilburn, Brian A.; Romero, Roberto; Leach, Richard E.; Armant, D. Randall

2010-01-01

Heparin-binding EGF-like growth factor (HBEGF) is expressed by trophoblast cells throughout gestation. First-trimester cytotrophoblast cells are protected from hypoxia-induced apoptosis because of the accumulation of HBEGF through a posttranscriptional autocrine mechanism. Exogenous application of HBEGF is cytoprotective in a hypoxia/reoxygenation (H/R) injury model and initiates trophoblast extravillous differentiation to an invasive phenotype. The downstream signaling pathways induced by HBEGF that mediate these various cellular activities were identified using two human first-trimester cytotrophoblast cell lines, HTR-8/SVneo and SW.71, with similar results. Recombinant HBEGF (1 nM) induced transient phosphorylation of MAPK3/1 (ERK), MAPK14 (p38), and AKT within 15 min and JNK after 1–2 h. To determine which downstream pathways regulate the various functions of HBEGF, cells were treated with specific inhibitors of the ERK upstream regulator MEK (U0126), the AKT upstream regulator phosphoinositide-3 (PI3)-kinase (LY294002), MAPK14 (SB203580), and JNK (SP600125), as well as with inactive structural analogues. Only SB203580 specifically prevented HBEGF-mediated rescue during H/R, while each inhibitor attenuated HBEGF-stimulated cell migration. Accumulation of HBEGF at reduced oxygen was blocked only by a combination of U0126, SB203580, and SP600125. We conclude that HBEGF advances trophoblast extravillous differentiation through coordinate activation of PI3 kinase, ERK, MAPK14, and JNK, while only MAPK14 is required for its antiapoptotic activity. Additionally, hypoxia induces an autocrine increase in HBEGF protein levels through MAPK14, JNK or ERK. These experiments reveal a complexity of the intracellular signaling circuitry that regulates trophoblast functions critical for implantation and placentation. PMID:20130271

Examining the architecture of cellular computing through a comparative study with a computer.

PubMed

Wang, Degeng; Gribskov, Michael

2005-06-22

The computer and the cell both use information embedded in simple coding, the binary software code and the quadruple genomic code, respectively, to support system operations. A comparative examination of their system architecture as well as their information storage and utilization schemes is performed. On top of the code, both systems display a modular, multi-layered architecture, which, in the case of a computer, arises from human engineering efforts through a combination of hardware implementation and software abstraction. Using the computer as a reference system, a simplistic mapping of the architectural components between the two is easily detected. This comparison also reveals that a cell abolishes the software-hardware barrier through genomic encoding for the constituents of the biochemical network, a cell's "hardware" equivalent to the computer central processing unit (CPU). The information loading (gene expression) process acts as a major determinant of the encoded constituent's abundance, which, in turn, often determines the "bandwidth" of a biochemical pathway. Cellular processes are implemented in biochemical pathways in parallel manners. In a computer, on the other hand, the software provides only instructions and data for the CPU. A process represents just sequentially ordered actions by the CPU and only virtual parallelism can be implemented through CPU time-sharing. Whereas process management in a computer may simply mean job scheduling, coordinating pathway bandwidth through the gene expression machinery represents a major process management scheme in a cell. In summary, a cell can be viewed as a super-parallel computer, which computes through controlled hardware composition. While we have, at best, a very fragmented understanding of cellular operation, we have a thorough understanding of the computer throughout the engineering process. The potential utilization of this knowledge to the benefit of systems biology is discussed.

A difference in the pattern of repair in a large genomic region in UV-irradiated normal human and Cockayne syndrome cells.

PubMed

Shanower, G A; Kantor, G J

1997-11-01

Xeroderma pigmentosum group C cells repair DNA damaged by ultraviolet radiation in an unusual pattern throughout the genome. They remove cyclobutane pyrimidine dimers only from the DNA of transcriptionally active chromatin regions and only from the strand that contains the transcribed strand. The repair proceeds in a manner that creates damage-free islands which are in some cases much larger than the active gene associated with them. For example, the small transcriptionally active beta-actin gene (3.5 kb) is repaired as part of a 50 kb single-stranded region. The repair responsible for creating these islands requires active transcription, suggesting that the two activities are coupled. A preferential repair pathway in normal human cells promotes repair of actively transcribed DNA strands and is coupled to transcription. It is not known if similar large islands, referred to as repair domains, are preferentially created as a result of the coupling. Data are presented showing that in normal cells, preferential repair in the beta-actin region is associated with the creation of a large, completely repaired region in the partially repaired genome. Repair at other genomic locations which contain inactive genes (insulin, 754) does not create similar large regions as quickly. In contrast, repair in Cockayne syndrome cells, which are defective in the preferential repair pathway but not in genome-overall repair, proceeds in the beta-actin region by a mechanism which does not create preferentially a large repaired region. Thus a correlation between the activity required to preferentially repair active genes and that required to create repaired domains is detected. We propose an involvement of the transcription-repair coupling factor in a coordinated repair pathway for removing DNA damage from entire transcription units.

A direct GABAergic output from the basal ganglia to frontal cortex

PubMed Central

Saunders, Arpiar; Oldenburg, Ian A.; Berezovskii, Vladimir K.; Johnson, Caroline A.; Kingery, Nathan D.; Elliott, Hunter L.; Xie, Tiao; Gerfen, Charles R.; Sabatini, Bernardo L.

2014-01-01

The basal ganglia (BG) are phylogenetically conserved subcortical nuclei necessary for coordinated motor action and reward learning1. Current models postulate that the BG modulate cerebral cortex indirectly via an inhibitory output to thalamus, bidirectionally controlled by the BG via direct (dSPNs) and indirect (iSPNs) pathway striatal projection neurons2–4. The BG thalamic output sculpts cortical activity by interacting with signals from sensory and motor systems5. Here we describe a direct projection from the globus pallidus externus (GP), a central nucleus of the BG, to frontal regions of the cerebral cortex (FC). Two cell types make up the GP-FC projection, distinguished by their electrophysiological properties, cortical projections and expression of choline acetyltransferase (ChAT), a synthetic enzyme for the neurotransmitter acetylcholine (ACh). Despite these differences, ChAT+ cells, which have been historically identified as an extension of the nucleus basalis (NB), as well as ChAT− cells, release the inhibitory neurotransmitter GABA (γ-aminobutyric acid) and are inhibited by iSPNs and dSPNs of dorsal striatum. Thus GP-FC cells comprise a direct GABAergic/cholinergic projection under the control of striatum that activates frontal cortex in vivo. Furthermore, iSPN inhibition of GP-FC cells is sensitive to dopamine 2 receptor signaling, revealing a pathway by which drugs that target dopamine receptors for the treatment of neuropsychiatric disorders can act in the BG to modulate frontal cortices. PMID:25739505

Drosophila nemo is an essential gene involved in the regulation of programmed cell death.

PubMed

Mirkovic, Ivana; Charish, Kristi; Gorski, Sharon M; McKnight, Kristen; Verheyen, Esther M

2002-11-01

Nemo-like kinases define a novel family of serine/threonine kinases that are involved in integrating multiple signaling pathways. They are conserved regulators of Wnt/Wingless pathways, which may coordinate Wnt with TGFbeta-mediated signaling. Drosophila nemo was identified through its involvement in epithelial planar polarity, a process regulated by a non-canonical Wnt pathway. We have previously found that ectopic expression of Nemo using the Gal4-UAS system resulted in embryonic lethality associated with defects in patterning and head development. In this study we present our analyses of the phenotypes of germline clone-derived embryos. We observe lethality associated with head defects and reduction of programmed cell death and conclude that nmo is an essential gene. We also present data showing that nmo is involved in regulating apoptosis during eye development, based on both loss of function phenotypes and on genetic interactions with the pro-apoptotic gene reaper. Finally, we present genetic data from the adult wing that suggest the activity of ectopically expressed Nemo can be modulated by Jun N-terminal kinase (JNK) signaling. Such an observation supports the model that there is cross-talk between Wnt, TGFbeta and JNK signaling at multiple stages of development. Copyright 2002 Elsevier Science Ireland Ltd.

The REP2 Repeats of the Genome of Neisseria meningitidis Are Associated with Genes Coordinately Regulated during Bacterial Cell Interaction

PubMed Central

Morelle, Sandrine; Carbonnelle, Etienne; Nassif, Xavier

2003-01-01

Interaction with host cells is essential in meningococcal pathogenesis especially at the blood-brain barrier. This step is likely to involve a common regulatory pathway allowing coordinate regulation of genes necessary for the interaction with endothelial cells. The analysis of the genomic sequence of Neisseria meningitidis Z2491 revealed the presence of many repeats. One of these, designated REP2, contains a −24/−12 type promoter and a ribosome binding site 5 to 13 bp before an ATG. In addition most of these REP2 sequences are located immediately upstream of an ORF. Among these REP2-associated genes are pilC1 and crgA, described as being involved in steps essential for the interaction of N. meningitidis with host cells. Furthermore, the REP2 sequences located upstream of pilC1 and crgA correspond to the previously identified promoters known to be induced during the initial localized adhesion of N. meningitidis with human cells. This characteristic led us to hypothesize that at least some of the REP2-associated genes were upregulated under the same circumstances as pilC1 and crgA. Quantitative PCR in real time demonstrated that the expression of 14 out of 16 REP2-associated genes were upregulated during the initial localized adhesion of N. meningitidis. Taken together, these data suggest that these repeats control a set of genes necessary for the efficient interaction of this pathogen with host cells. Subsequent mutational analysis was performed to address the role of these genes during meningococcus-cell interaction. PMID:12670987

Study of ATM Phosphorylation by Cdk5 in Neuronal Cells.

PubMed

She, Hua; Mao, Zixu

2017-01-01

The phosphatidylinositol-3-kinase-like kinase ATM (ataxia-telangiectasia mutated) plays a central role in coordinating the DNA damage responses including cell cycle checkpoint control, DNA repair, and apoptosis. Mutations of ATM cause a spectrum of defects ranging from neurodegeneration to cancer predisposition. We previously showed that Cdk5 (cyclin-dependent kinase 5) is activated by DNA damage and directly phosphorylates ATM at serine 794 in postmitotic neurons. Phosphorylation at serine 794 precedes and is required for ATM autophosphorylation at serine 1981, and activates ATM kinase activity. Cdk5-ATM pathway plays a crucial role in DNA damage-induced neuronal injury. This chapter describes protocols used in analyzing ATM phosphorylation by Cdk5 in CGNs (cerebellar granule neurons) and its effects on neuronal survival.

The Myb-domain protein ULTRAPETALA1 INTERACTING FACTOR 1 controls floral meristem activities in Arabidopsis.

PubMed

Moreau, Fanny; Thévenon, Emmanuel; Blanvillain, Robert; Lopez-Vidriero, Irene; Franco-Zorrilla, Jose Manuel; Dumas, Renaud; Parcy, François; Morel, Patrice; Trehin, Christophe; Carles, Cristel C

2016-04-01

Higher plants continuously and iteratively produce new above-ground organs in the form of leaves, stems and flowers. These organs arise from shoot apical meristems whose homeostasis depends on coordination between self-renewal of stem cells and their differentiation into organ founder cells. This coordination is stringently controlled by the central transcription factor WUSCHEL (WUS), which is both necessary and sufficient for stem cell specification in Arabidopsis thaliana ULTRAPETALA1 (ULT1) was previously identified as a plant-specific, negative regulator of WUS expression. However, molecular mechanisms underlying this regulation remain unknown. ULT1 protein contains a SAND putative DNA-binding domain and a B-box, previously proposed as a protein interaction domain in eukaryotes. Here, we characterise a novel partner of ULT1, named ULT1 INTERACTING FACTOR 1 (UIF1), which contains a Myb domain and an EAR motif. UIF1 and ULT1 function in the same pathway for regulation of organ number in the flower. Moreover, UIF1 displays DNA-binding activity and specifically binds to WUS regulatory elements. We thus provide genetic and molecular evidence that UIF1 and ULT1 work together in floral meristem homeostasis, probably by direct repression of WUS expression. © 2016. Published by The Company of Biologists Ltd.

Three ancient hormonal cues co-ordinate shoot branching in a moss.

PubMed

Coudert, Yoan; Palubicki, Wojtek; Ljung, Karin; Novak, Ondrej; Leyser, Ottoline; Harrison, C Jill

2015-03-25

Shoot branching is a primary contributor to plant architecture, evolving independently in flowering plant sporophytes and moss gametophytes. Mechanistic understanding of branching is largely limited to flowering plants such as Arabidopsis, which have a recent evolutionary origin. We show that in gametophytic shoots of Physcomitrella, lateral branches arise by re-specification of epidermal cells into branch initials. A simple model co-ordinating the activity of leafy shoot tips can account for branching patterns, and three known and ancient hormonal regulators of sporophytic branching interact to generate the branching pattern- auxin, cytokinin and strigolactone. The mode of auxin transport required in branch patterning is a key divergence point from known sporophytic pathways. Although PIN-mediated basipetal auxin transport regulates branching patterns in flowering plants, this is not so in Physcomitrella, where bi-directional transport is required to generate realistic branching patterns. Experiments with callose synthesis inhibitors suggest plasmodesmal connectivity as a potential mechanism for transport.

Neural crest development and craniofacial morphogenesis is coordinated by nitric oxide and histone acetylation

PubMed Central

Kong, Yawei; Grimaldi, Michael; Curtin, Eugene; Dougherty, Max; Kaufman, Charles; White, Richard M.; Zon, Leonard I.; Liao, Eric C.

2015-01-01

Cranial neural crest (CNC) cells are patterned and coalesce to facial prominences that undergo convergence and extension to generate the craniofacial form. We applied a chemical genetics approach to identify pathways that regulate craniofacial development during embryogenesis. Treatment with the nitric oxide synthase inhibitor TRIM abrogated first pharyngeal arch structures and induced ectopic ceratobranchial formation. TRIM promoted a progenitor CNC fate and inhibited chondrogenic differentiation, which were mediated through impaired nitric oxide (NO) production without appreciable effect on global protein S-nitrosylation. Instead, TRIM perturbed hox gene patterning and caused histone hypoacetylation. Rescue of TRIM phenotype was achieved with over-expression of histone acetyltransferase kat6a, inhibition of histone deacetylase, and complimentary NO. These studies demonstrate that NO signaling and histone acetylation are coordinated mechanisms that regulate CNC patterning, differentiation and convergence during craniofacial morphogenesis. PMID:24684905

Phosphoinositide 3-Kinase Regulates Glycolysis through Mobilization of Aldolase from the Actin cytoskeleton

PubMed Central

Hu, Hai; Juvekar, Ashish; Lyssiotis, Costas A.; Lien, Evan C.; Albeck, John G.; Oh, Doogie; Varma, Gopal; Hung, Yin Pun; Ullas, Soumya; Lauring, Josh; Seth, Pankaj; Lundquist, Mark R.; Tolan, Dean R.; Grant, Aaron K.; Needleman, Daniel J.; Asara, John M.; Cantley, Lewis C.

2016-01-01

Summary The Phosphoinositide 3-Kinase (PI3K) pathway regulates multiple steps in glucose metabolism but also cytoskeletal functions, such as cell movement and attachment. Here we show that PI3K directly coordinates glycolysis with cytoskeletal dynamics in an AKT-independent manner. Growth factors or insulin stimulate the PI3K-dependent activation of Rac, leading to disruption of the actin cytoskeleton, release of filamentous actin-bound aldolase A and an increase in aldolase activity. Consistently, PI3K-, but not AKT-, SGK- or mTOR-inhibitors, cause a significant decrease in glycolysis at the step catalyzed by aldolase, while activating PIK3CA mutations have the opposite effect. These results point towards a master regulatory function of PI3K that integrates an epithelial cell’s metabolism and its form, shape and function, coordinating glycolysis with the energy-intensive dynamics of actin remodeling. PMID:26824656

Tissue architecture and breast cancer: the role of extracellular matrix and steroid hormones

PubMed Central

Hansen, R K; Bissell, M J

2010-01-01

The changes in tissue architecture that accompany the development of breast cancer have been the focus of investigations aimed at developing new cancer therapeutics. As we learn more about the normal mammary gland, we have begun to understand the complex signaling pathways underlying the dramatic shifts in the structure and function of breast tissue. Integrin-, growth factor-, and steroid hormone-signaling pathways all play an important part in maintaining tissue architecture; disruption of the delicate balance of signaling results in dramatic changes in the way cells interact with each other and with the extracellular matrix, leading to breast cancer. The extracellular matrix itself plays a central role in coordinating these signaling processes. In this review, we consider the interrelationships between the extracellular matrix, integrins, growth factors, and steroid hormones in mammary gland development and function. PMID:10903527

Coordinate Regulation of Yeast Sterol Regulatory Element-binding Protein (SREBP) and Mga2 Transcription Factors.

PubMed

Burr, Risa; Stewart, Emerson V; Espenshade, Peter J

2017-03-31

The Mga2 and Sre1 transcription factors regulate oxygen-responsive lipid homeostasis in the fission yeast Schizosaccharomyces pombe in a manner analogous to the mammalian sterol regulatory element-binding protein (SREBP)-1 and SREBP-2 transcription factors. Mga2 and SREBP-1 regulate triacylglycerol and glycerophospholipid synthesis, whereas Sre1 and SREBP-2 regulate sterol synthesis. In mammals, a shared activation mechanism allows for coordinate regulation of SREBP-1 and SREBP-2. In contrast, distinct pathways activate fission yeast Mga2 and Sre1. Therefore, it is unclear whether and how these two related pathways are coordinated to maintain lipid balance in fission yeast. Previously, we showed that Sre1 cleavage is defective in the absence of mga2 Here, we report that this defect is due to deficient unsaturated fatty acid synthesis, resulting in aberrant membrane transport. This defect is recapitulated by treatment with the fatty acid synthase inhibitor cerulenin and is rescued by addition of exogenous unsaturated fatty acids. Furthermore, sterol synthesis inhibition blocks Mga2 pathway activation. Together, these data demonstrate that Sre1 and Mga2 are each regulated by the lipid product of the other transcription factor pathway, providing a source of coordination for these two branches of lipid synthesis. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Surfing the wave, cycle, life history, and genes/proteins expressed by testicular germ cells. Part 4: intercellular bridges, mitochondria, nuclear envelope, apoptosis, ubiquitination, membrane/voltage-gated channels, methylation/acetylation, and transcription factors.

PubMed

Hermo, Louis; Pelletier, R-Marc; Cyr, Daniel G; Smith, Charles E

2010-04-01

As germ cells divide and differentiate from spermatogonia to spermatozoa, they share a number of structural and functional features that are common to all generations of germ cells and these features are discussed herein. Germ cells are linked to one another by large intercellular bridges which serve to move molecules and even large organelles from the cytoplasm of one cell to another. Mitochondria take on different shapes and features and topographical arrangements to accommodate their specific needs during spermatogenesis. The nuclear envelope and pore complex also undergo extensive modifications concomitant with the development of germ cell generations. Apoptosis is an event that is normally triggered by germ cells and involves many proteins. It occurs to limit the germ cell pool and acts as a quality control mechanism. The ubiquitin pathway comprises enzymes that ubiquitinate as well as deubiquitinate target proteins and this pathway is present and functional in germ cells. Germ cells express many proteins involved in water balance and pH control as well as voltage-gated ion channel movement. In the nucleus, proteins undergo epigenetic modifications which include methylation, acetylation, and phosphorylation, with each of these modifications signaling changes in chromatin structure. Germ cells contain specialized transcription complexes that coordinate the differentiation program of spermatogenesis, and there are many male germ cell-specific differences in the components of this machinery. All of the above features of germ cells will be discussed along with the specific proteins/genes and abnormalities to fertility related to each topic. Copyright 2009 Wiley-Liss, Inc.

pH-responsive, gluconeogenic renal epithelial LLC-PK1-FBPase+cells: a versatile in vitro model to study renal proximal tubule metabolism and function

PubMed Central

Curthoys, Norman P.

2014-01-01

Ammoniagenesis and gluconeogenesis are prominent metabolic features of the renal proximal convoluted tubule that contribute to maintenance of systemic acid-base homeostasis. Molecular analysis of the mechanisms that mediate the coordinate regulation of the two pathways required development of a cell line that recapitulates these features in vitro. By adapting porcine renal epithelial LLC-PK1 cells to essentially glucose-free medium, a gluconeogenic subline, termed LLC-PK1-FBPase+ cells, was isolated. LLC-PK1-FBPase+ cells grow in the absence of hexoses and pentoses and exhibit enhanced oxidative metabolism and increased levels of phosphate-dependent glutaminase. The cells also express significant levels of the key gluconeogenic enzymes, fructose-1,6-bisphosphatase (FBPase) and phosphoenolpyruvate carboxykinase (PEPCK). Thus the altered phenotype of LLC-PK1-FBPase+ cells is pleiotropic. Most importantly, when transferred to medium that mimics a pronounced metabolic acidosis (9 mM HCO3−, pH 6.9), the LLC-PK1-FBPase+ cells exhibit a gradual increase in NH4+ ion production, accompanied by increases in glutaminase and cytosolic PEPCK mRNA levels and proteins. Therefore, the LLC-PK1-FBPase+ cells retained in culture many of the metabolic pathways and pH-responsive adaptations characteristic of renal proximal tubules. The molecular mechanisms that mediate enhanced expression of the glutaminase and PEPCK in LLC-PK1-FBPase+ cells have been extensively reviewed. The present review describes novel properties of this unique cell line and summarizes the molecular mechanisms that have been defined more recently using LLC-PK1-FBPase+ cells to model the renal proximal tubule. It also identifies future studies that could be performed using these cells. PMID:24808535

Oncogenic properties of apoptotic tumor cells in aggressive B cell lymphoma.

PubMed

Ford, Catriona A; Petrova, Sofia; Pound, John D; Voss, Jorine J L P; Melville, Lynsey; Paterson, Margaret; Farnworth, Sarah L; Gallimore, Awen M; Cuff, Simone; Wheadon, Helen; Dobbin, Edwina; Ogden, Carol Anne; Dumitriu, Ingrid E; Dunbar, Donald R; Murray, Paul G; Ruckerl, Dominik; Allen, Judith E; Hume, David A; van Rooijen, Nico; Goodlad, John R; Freeman, Tom C; Gregory, Christopher D

2015-03-02

Cells undergoing apoptosis are known to modulate their tissue microenvironments. By acting on phagocytes, notably macrophages, apoptotic cells inhibit immunological and inflammatory responses and promote trophic signaling pathways. Paradoxically, because of their potential to cause death of tumor cells and thereby militate against malignant disease progression, both apoptosis and tumor-associated macrophages (TAMs) are often associated with poor prognosis in cancer. We hypothesized that, in progression of malignant disease, constitutive loss of a fraction of the tumor cell population through apoptosis could yield tumor-promoting effects. Here, we demonstrate that apoptotic tumor cells promote coordinated tumor growth, angiogenesis, and accumulation of TAMs in aggressive B cell lymphomas. Through unbiased "in situ transcriptomics" analysis-gene expression profiling of laser-captured TAMs to establish their activation signature in situ-we show that these cells are activated to signal via multiple tumor-promoting reparatory, trophic, angiogenic, tissue remodeling, and anti-inflammatory pathways. Our results also suggest that apoptotic lymphoma cells help drive this signature. Furthermore, we demonstrate that, upon induction of apoptosis, lymphoma cells not only activate expression of the tumor-promoting matrix metalloproteinases MMP2 and MMP12 in macrophages but also express and process these MMPs directly. Finally, using a model of malignant melanoma, we show that the oncogenic potential of apoptotic tumor cells extends beyond lymphoma. In addition to its profound tumor-suppressive role, apoptosis can potentiate cancer progression. These results have important implications for understanding the fundamental biology of cell death, its roles in malignant disease, and the broader consequences of apoptosis-inducing anti-cancer therapy. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Phosphorylation of the Bacillus subtilis Replication Controller YabA Plays a Role in Regulation of Sporulation and Biofilm Formation

PubMed Central

García García, Tránsito; Ventroux, Magali; Derouiche, Abderahmane; Bidnenko, Vladimir; Correia Santos, Sara; Henry, Céline; Mijakovic, Ivan; Noirot-Gros, Marie-Françoise; Poncet, Sandrine

2018-01-01

Bacillus subtilis cells can adopt different life-styles in response to various environmental cues, including planktonic cells during vegetative growth, sessile cells during biofilm formation and sporulation. While switching life-styles, bacteria must coordinate the progression of their cell cycle with their physiological status. Our current understanding of the regulatory pathways controlling the decision-making processes and triggering developmental switches highlights a key role of protein phosphorylation. The regulatory mechanisms that integrate the bacterial chromosome replication status with sporulation involve checkpoint proteins that target the replication initiator DnaA or the kinase phosphorelay controlling the master regulator Spo0A. B. subtilis YabA is known to interact with DnaA to prevent over-initiation of replication during vegetative growth. Here, we report that YabA is phosphorylated by YabT, a Ser/Thr kinase expressed during sporulation and biofilm formation. The phosphorylation of YabA has no effect on replication initiation control but hyper-phosphorylation of YabA leads to an increase in sporulation efficiency and a strong inhibition of biofilm formation. We also provide evidence that YabA phosphorylation affects the level of Spo0A-P in cells. These results indicate that YabA is a multifunctional protein with a dual role in regulating replication initiation and life-style switching, thereby providing a potential mechanism for cross-talk and coordination of cellular processes during adaptation to environmental change. PMID:29619013

Phosphorylation of the Bacillus subtilis Replication Controller YabA Plays a Role in Regulation of Sporulation and Biofilm Formation.

PubMed

García García, Tránsito; Ventroux, Magali; Derouiche, Abderahmane; Bidnenko, Vladimir; Correia Santos, Sara; Henry, Céline; Mijakovic, Ivan; Noirot-Gros, Marie-Françoise; Poncet, Sandrine

2018-01-01

Bacillus subtilis cells can adopt different life-styles in response to various environmental cues, including planktonic cells during vegetative growth, sessile cells during biofilm formation and sporulation. While switching life-styles, bacteria must coordinate the progression of their cell cycle with their physiological status. Our current understanding of the regulatory pathways controlling the decision-making processes and triggering developmental switches highlights a key role of protein phosphorylation. The regulatory mechanisms that integrate the bacterial chromosome replication status with sporulation involve checkpoint proteins that target the replication initiator DnaA or the kinase phosphorelay controlling the master regulator Spo0A. B. subtilis YabA is known to interact with DnaA to prevent over-initiation of replication during vegetative growth. Here, we report that YabA is phosphorylated by YabT, a Ser/Thr kinase expressed during sporulation and biofilm formation. The phosphorylation of YabA has no effect on replication initiation control but hyper-phosphorylation of YabA leads to an increase in sporulation efficiency and a strong inhibition of biofilm formation. We also provide evidence that YabA phosphorylation affects the level of Spo0A-P in cells. These results indicate that YabA is a multifunctional protein with a dual role in regulating replication initiation and life-style switching, thereby providing a potential mechanism for cross-talk and coordination of cellular processes during adaptation to environmental change.

Primary cilium - antenna-like structure on the surface of most mammalian cell types

NASA Astrophysics Data System (ADS)

Dvorak, J.; Sitorova, V.; Hadzi Nikolov, D.; Mokry, J.; Richter, I.; Kasaova, L.; Filip, S.; Ryska, A.; Petera, J.

2011-12-01

The primary cilium is a sensory solitary non-motile microtubule-based organelle protruding in the quiescent phase of the cell cycle from the surface of the majority of human cells, including embryonic cells, stem cells and stromal cells of malignant tumors. The presence of a primary cilium on the surface of a cell is transient, limited to the quiescent G1(G0) phase and the beginning of the S phase of the cell cycle. The primary cilium is formed from the mother centriole. Primary cilia are key coordinators of signaling pathways during development and tissue homeostasis and, when deffective, they are a major cause of human diseases and developmental disorders, now commonly referred to as ciliopathies. Most cancer cells do not possess a primary cilium. The loss of the primary cilium is a regular feature of neoplastic transformation in the majority of solid tumors. The primary cilium could serve as a tumor suppressor organelle. The aim of this paper was to provide a review of the current knowledge of the primary cilium.

The Human Transcriptome During Nontyphoid Salmonella and HIV Coinfection Reveals Attenuated NFκB-Mediated Inflammation and Persistent Cell Cycle Disruption

PubMed Central

Schreiber, Fernanda; Lynn, David J.; Houston, Angela; Peters, Joanna; Mwafulirwa, Gershom; Finlay, Brett B.; Brinkman, Fiona S. L.; Hancock, Robert E. W.; Heyderman, Robert S.; Dougan, Gordon

2011-01-01

Background. Invasive nontyphoid Salmonella (iNTS) disease is common and severe in adults with human immunodeficiency virus (HIV) infection in Africa. We previously observed that ex vivo macrophages from HIV-infected subjects challenged with Salmonella Typhimurium exhibit dysregulated proinflammatory cytokine responses. Methods. We studied the transcriptional response in whole blood from HIV-positive patients during acute and convalescent iNTS disease compared to other invasive bacterial diseases, and to HIV-positive and -negative controls. Results. During iNTS disease, there was a remarkable lack of a coordinated inflammatory or innate immune signaling response. Few interferon γ (IFNγ)--induced genes or Toll-like receptor/transcription factor nuclear factor κB (TLR/NFκB) gene pathways were upregulated in expression. Ex vivo lipopolysacharide (LPS) or flagellin stimulation of whole blood, however, showed that convalescent iNTS subjects and controls were competent to mount prominent TLR/NFκB-associated patterns of mRNA expression. In contrast, HIV-positive patients with other invasive bacterial infections (Escherichia coli and Streptococcus pneumoniae) displayed a pronounced proinflammatory innate immune transcriptional response. There was also upregulated mRNA expression in cell cycle, DNA replication, translation and repair, and viral replication pathways during iNTS. These patterns persisted for up to 2 months into convalescence. Conclusions. Attenuation of NFκB-mediated inflammation and dysregulation of cell cycle and DNA-function gene pathway expression are key features of the interplay between iNTS and HIV. PMID:21917897

REV7 counteracts DNA double-strand break resection and impacts PARP inhibition

PubMed Central

Xu, Guotai; Yuan, Jingsong; Mistrik, Martin; Bouwman, Peter; Bartkova, Jirina; Gogola, Ewa; Warmerdam, Daniël; Barazas, Marco; Jaspers, Janneke E.; Watanabe, Kenji; Pieterse, Mark; Kersbergen, Ariena; Sol, Wendy; Celie, Patrick H. N.; Schouten, Philip C.; van den Broek, Bram; Salman, Ahmed; Nieuwland, Marja; de Rink, Iris; de Ronde, Jorma; Jalink, Kees; Boulton, Simon J.; Chen, Junjie; van Gent, Dik C.; Bartek, Jiri; Jonkers, Jos; Borst, Piet; Rottenberg, Sven

2015-01-01

Summary Error-free repair of DNA double-strand breaks (DSB) is achieved by homologous recombination (HR), and BRCA1 is an important factor for this repair pathway1. In the absence of BRCA1-mediated HR, administration of PARP inhibitors induces synthetic lethality of tumor cells of patients with breast or ovarian cancers2,3. Despite the benefit of this tailored therapy, drug resistance can occur by HR restoration4. Genetic reversion of BRCA1-inactivating mutations can be the underlying mechanism of drug resistance, but this does not explain resistance in all cases5. In particular, little is known about BRCA1-independent restoration of HR. Here, we show that loss of REV7 (also known as MAD2L2) re-establishes CtIP-dependent end resection of DSBs in BRCA1-deficient cells, leading to HR restoration and PARP inhibitor resistance, reversed by ATM kinase inhibition. REV7 is recruited to DSBs in a manner dependent on the H2AX-MDC1-RNF8-RNF168-53BP1 chromatin pathway, and appears to block HR and promote end joining in addition to its regulatory role in DNA damage tolerance6. Finally, we establish that REV7 blocks DSB resection to promote non-homologous end-joining (NHEJ) during immunoglobulin class switch recombination. Our results reveal an unexpected critical function of REV7 downstream of 53BP1 in coordinating pathological DSB repair pathway choices in BRCA1-deficient cells. PMID:25799992

Failure of protein quality control in amyotrophic lateral sclerosis.

PubMed

Kabashi, Edor; Durham, Heather D

2006-01-01

The protein chaperoning and ubiquitin-proteasome systems perform many homeostatic functions within cells involving protein folding, transport and degradation. Of paramount importance is ridding cells of mutant or post-translationally modified proteins that otherwise tend to aggregate into insoluble complexes and form inclusions. Such inclusions are characteristic of many neurodegenerative diseases and implicate protein misfolding and aggregation as common aspects of pathogenesis. In the most common familial form of ALS, mutations in SOD1 promote misfolding of the protein and target it for degradation by proteasomes. Although proteasomes can degrade the mutant proteins efficiently, altered solubility and aggregation of mutant SOD1 are features of the disease and occur most prominently in the most vulnerable cells and tissues. Indeed, lumbar spinal cord of mutant SOD1 transgenic mice show early reduction in their capacity for protein chaperoning and proteasome-mediated hydrolysis of substrates, and motor neurons are particularly vulnerable to aggregation of mutant SOD1. A high threshold for upregulating key pathways in response to the stress of added substrate load may contribute to this vulnerability. The broad spectrum neuroprotective capability and efficacy of some chaperone-based therapies in preclinical models makes these pathways attractive as targets for therapy in ALS, as well as other neurodegenerative diseases. A better understanding of the mechanisms governing the regulation of protein chaperones and UPS components would facilitate development of treatments that upregulate these pathways in a coordinated manner in neural tissue without long term toxicity.

Rubrocerebellar Feedback Loop Isolates the Interposed Nucleus as an Independent Processor of Corollary Discharge Information in Mice

PubMed Central

Beitzel, Christy S.; Houck, Brenda D.; Lewis, Samantha M.

2017-01-01

Understanding cerebellar contributions to motor coordination requires deeper insight into how the output structures of the cerebellum, the cerebellar nuclei, integrate their inputs and influence downstream motor pathways. The magnocellular red nucleus (RNm), a brainstem premotor structure, is a major target of the interposed nucleus (IN), and has also been described in previous studies to send feedback collaterals to the cerebellum. Because such a pathway is in a key position to provide motor efferent information to the cerebellum, satisfying predictions about the use of corollary discharge in cerebellar computations, we studied it in mice of both sexes. Using anterograde viral tracing, we show that innervation of cerebellum by rubrospinal neuron collaterals is remarkably selective for the IN compared with the cerebellar cortex. Optogenetic activation of the pathway in acute mouse brain slices drove IN activity despite small amplitude synaptic currents, suggesting an active role in IN information processing. Monosynaptic transsynaptic rabies tracing indicated the pathway contacts multiple cell types within the IN. By contrast, IN inputs to the RNm targeted a region that lacked inhibitory neurons. Optogenetic drive of IN inputs to the RNm revealed strong, direct excitation but no inhibition of RNm neurons. Together, these data indicate that the cerebellar nuclei are under afferent control independent of the cerebellar cortex, potentially diversifying its roles in motor control. SIGNIFICANCE STATEMENT The common assumption that all cerebellar mossy fibers uniformly collateralize to the cerebellar nuclei and cortex underlies classic models of convergent Purkinje influence on cerebellar output. Specifically, mossy fibers are thought to both directly excite nuclear neurons and drive polysynaptic feedforward inhibition via Purkinje neurons, setting up a fundamental computational unit. Here we present data that challenge this rule. A dedicated cerebellar nuclear afferent comprised of feedback collaterals from premotor rubrospinal neurons can directly modulate IN output independent of Purkinje cell modulation. In contrast to the IN-RNm pathway, the RNm-IN feedback pathway targets multiple cell types, potentially influencing both motor output pathways and nucleo-olivary feedback. PMID:28916520

Rubrocerebellar Feedback Loop Isolates the Interposed Nucleus as an Independent Processor of Corollary Discharge Information in Mice.

PubMed

Beitzel, Christy S; Houck, Brenda D; Lewis, Samantha M; Person, Abigail L

2017-10-18

Understanding cerebellar contributions to motor coordination requires deeper insight into how the output structures of the cerebellum, the cerebellar nuclei, integrate their inputs and influence downstream motor pathways. The magnocellular red nucleus (RNm), a brainstem premotor structure, is a major target of the interposed nucleus (IN), and has also been described in previous studies to send feedback collaterals to the cerebellum. Because such a pathway is in a key position to provide motor efferent information to the cerebellum, satisfying predictions about the use of corollary discharge in cerebellar computations, we studied it in mice of both sexes. Using anterograde viral tracing, we show that innervation of cerebellum by rubrospinal neuron collaterals is remarkably selective for the IN compared with the cerebellar cortex. Optogenetic activation of the pathway in acute mouse brain slices drove IN activity despite small amplitude synaptic currents, suggesting an active role in IN information processing. Monosynaptic transsynaptic rabies tracing indicated the pathway contacts multiple cell types within the IN. By contrast, IN inputs to the RNm targeted a region that lacked inhibitory neurons. Optogenetic drive of IN inputs to the RNm revealed strong, direct excitation but no inhibition of RNm neurons. Together, these data indicate that the cerebellar nuclei are under afferent control independent of the cerebellar cortex, potentially diversifying its roles in motor control. SIGNIFICANCE STATEMENT The common assumption that all cerebellar mossy fibers uniformly collateralize to the cerebellar nuclei and cortex underlies classic models of convergent Purkinje influence on cerebellar output. Specifically, mossy fibers are thought to both directly excite nuclear neurons and drive polysynaptic feedforward inhibition via Purkinje neurons, setting up a fundamental computational unit. Here we present data that challenge this rule. A dedicated cerebellar nuclear afferent comprised of feedback collaterals from premotor rubrospinal neurons can directly modulate IN output independent of Purkinje cell modulation. In contrast to the IN-RNm pathway, the RNm-IN feedback pathway targets multiple cell types, potentially influencing both motor output pathways and nucleo-olivary feedback. Copyright © 2017 the authors 0270-6474/17/3710085-12$15.00/0.

TGF-β induction of FGF-2 expression in stromal cells requires integrated smad3 and MAPK pathways.

PubMed

Strand, Douglas W; Liang, Yao-Yun; Yang, Feng; Barron, David A; Ressler, Steven J; Schauer, Isaiah G; Feng, Xin-Hua; Rowley, David R

2014-01-01

Transforming Growth Factor-β (TGF-β) regulates the reactive stroma microenvironment associated with most carcinomas and mediates expression of many stromal derived factors important for tumor progression, including FGF-2 and CTGF. TGF-β is over-expressed in most carcinomas, and FGF-2 action is important in tumor-induced angiogenesis. The signaling mechanisms of how TGF-β regulates FGF-2 expression in the reactive stroma microenvironment are not understood. Accordingly, we have assessed key signaling pathways that mediate TGF-β1-induced FGF-2 expression in prostate stromal fibroblasts and mouse embryo fibroblasts (MEFs) null for Smad2 and Smad3. TGF-β1 induced phosphorylation of Smad2, Smad3, p38 and ERK1/2 proteins in both control MEFs and prostate fibroblasts. Of these, Smad3, but not Smad2 was found to be required for TGF-β1 induction of FGF-2 expression in stromal cells. ChIP analysis revealed a Smad3/Smad4 complex was associated with the -1.9 to -2.3 kb upstream proximal promoter of the FGF-2 gene, further suggesting a Smad3-specific regulation. In addition, chemical inhibition of p38 or ERK1/2 MAPK activity also blocked TGF-β1-induced FGF-2 expression in a Smad3-independent manner. Conversely, inhibition of JNK signaling enhanced FGF-2 expression. Together, these data indicate that expression of FGF-2 in fibroblasts in the tumor stromal cell microenvironment is coordinately dependent on both intact Smad3 and MAP kinase signaling pathways. These pathways and key downstream mediators of TGF-β action in the tumor reactive stroma microenvironment, may evolve as putative targets for therapeutic intervention.

The Transcription Factor EB Links Cellular Stress to the Immune Response



PubMed Central

Nabar, Neel R.; Kehrl, John H.

2017-01-01

The transcription factor EB (TFEB) is the master transcriptional regulator of autophagy and lysosome biogenesis. Recent advances have led to a paradigm shift in our understanding of lysosomes from a housekeeping cellular waste bin to a dynamically regulated pathway that is efficiently turned up or down based on cellular needs. TFEB coordinates the cellular response to nutrient deprivation and other forms of cell stress through the lysosome system, and regulates a myriad of cellular processes associated with this system including endocytosis, phagocytosis, autophagy, and lysosomal exocytosis. Autophagy and the endolysosomal system are critical to both the innate and adaptive arms of the immune system, with functions in effector cell priming and direct pathogen clearance. Recent studies have linked TFEB to the regulation of the immune response through the endolysosmal pathway and by direct transcriptional activation of immune related genes. In this review, we discuss the current understanding of TFEB’s function and the molecular mechanisms behind TFEB activation. Finally, we discuss recent advances linking TFEB to the immune response that positions lysosomal signaling as a potential target for immune modulation. PMID:28656016

A transcription factor links growth rate and metabolism in the hypersaline adapted archaeon Halobacterium salinarum.

PubMed

Todor, Horia; Dulmage, Keely; Gillum, Nicholas; Bain, James R; Muehlbauer, Michael J; Schmid, Amy K

2014-09-01

Co-ordinating metabolism and growth is a key challenge for all organisms. Despite fluctuating environments, cells must produce the same metabolic outputs to thrive. The mechanisms underlying this 'growth homeostasis' are known in bacteria and eukaryotes, but remain unexplored in archaea. In the model archaeon Halobacterium salinarum, the transcription factor TrmB regulates enzyme-coding genes in diverse metabolic pathways in response to glucose. However, H. salinarum is thought not to catabolize glucose. To resolve this discrepancy, we demonstrate that TrmB regulates the gluconeogenic production of sugars incorporated into the cell surface S-layer glycoprotein. Additionally, we show that TrmB-DNA binding correlates with instantaneous growth rate, likely because S-layer glycosylation is proportional to growth. This suggests that TrmB transduces a growth rate signal to co-regulated metabolic pathways including amino acid, purine, and cobalamin biosynthesis. Remarkably, the topology and function of this growth homeostatic network appear conserved across domains despite extensive alterations in protein components. © 2014 The Authors. Molecular Microbiology published by John Wiley & Sons Ltd.

The Transcription Factor EB Links Cellular Stress to the Immune Response

.

PubMed

Nabar, Neel R; Kehrl, John H

2017-06-01

The transcription factor EB (TFEB) is the master transcriptional regulator of autophagy and lysosome biogenesis. Recent advances have led to a paradigm shift in our understanding of lysosomes from a housekeeping cellular waste bin to a dynamically regulated pathway that is efficiently turned up or down based on cellular needs. TFEB coordinates the cellular response to nutrient deprivation and other forms of cell stress through the lysosome system, and regulates a myriad of cellular processes associated with this system including endocytosis, phagocytosis, autophagy, and lysosomal exocytosis. Autophagy and the endolysosomal system are critical to both the innate and adaptive arms of the immune system, with functions in effector cell priming and direct pathogen clearance. Recent studies have linked TFEB to the regulation of the immune response through the endolysosmal pathway and by direct transcriptional activation of immune related genes. In this review, we discuss the current understanding of TFEB's function and the molecular mechanisms behind TFEB activation. Finally, we discuss recent advances linking TFEB to the immune response that positions lysosomal signaling as a potential target for immune modulation.

YAP/TAZ regulates sprouting angiogenesis and vascular barrier maturation

PubMed Central

Kim, Yoo Hyung; Kim, Jaeryung; Park, Do Young; Bae, Hosung; Lee, Da-Hye; Kim, Kyun Hoo; Hong, Seon Pyo; Jang, Seung Pil; Kwon, Young-Guen; Lim, Dae-Sik

2017-01-01

Angiogenesis is a multistep process that requires coordinated migration, proliferation, and junction formation of vascular endothelial cells (ECs) to form new vessel branches in response to growth stimuli. Major intracellular signaling pathways that regulate angiogenesis have been well elucidated, but key transcriptional regulators that mediate these signaling pathways and control EC behaviors are only beginning to be understood. Here, we show that YAP/TAZ, a transcriptional coactivator that acts as an end effector of Hippo signaling, is critical for sprouting angiogenesis and vascular barrier formation and maturation. In mice, endothelial-specific deletion of Yap/Taz led to blunted-end, aneurysm-like tip ECs with fewer and dysmorphic filopodia at the vascular front, a hyper-pruned vascular network, reduced and disarranged distributions of tight and adherens junction proteins, disrupted barrier integrity, subsequent hemorrhage in growing retina and brain vessels, and reduced pathological choroidal neovascularization. Mechanistically, YAP/TAZ activates actin cytoskeleton remodeling, an important component of filopodia formation and junction assembly. Moreover, YAP/TAZ coordinates EC proliferation and metabolic activity by upregulating MYC signaling. Overall, these results show that YAP/TAZ plays multifaceted roles for EC behaviors, proliferation, junction assembly, and metabolism in sprouting angiogenesis and barrier formation and maturation and could be a potential therapeutic target for treating neovascular diseases. PMID:28805663

Endosperm and Nucellus Develop Antagonistically in Arabidopsis Seeds

PubMed Central

Xu, Wenjia; Coen, Olivier; Pechoux, Christine; Magnani, Enrico

2016-01-01

In angiosperms, seed architecture is shaped by the coordinated development of three genetically different components: embryo, endosperm, and maternal tissues. The relative contribution of these tissues to seed mass and nutrient storage varies considerably among species. The development of embryo, endosperm, or nucellus maternal tissue as primary storage compartments defines three main typologies of seed architecture. It is still debated whether the ancestral angiosperm seed accumulated nutrients in the endosperm or the nucellus. During evolution, plants shifted repeatedly between these two storage strategies through molecular mechanisms that are largely unknown. Here, we characterize the regulatory pathway underlying nucellus and endosperm tissue partitioning in Arabidopsis thaliana. We show that Polycomb-group proteins repress nucellus degeneration before fertilization. A signal initiated in the endosperm by the AGAMOUS-LIKE62 MADS box transcription factor relieves this Polycomb-mediated repression and therefore allows nucellus degeneration. Further downstream in the pathway, the TRANSPARENT TESTA16 (TT16) and GORDITA MADS box transcription factors promote nucellus degeneration. Moreover, we demonstrate that TT16 mediates the crosstalk between nucellus and seed coat maternal tissues. Finally, we characterize the nucellus cell death program and its feedback role in timing endosperm development. Altogether, our data reveal the antagonistic development of nucellus and endosperm, in coordination with seed coat differentiation. PMID:27233529

G protein-coupled receptors Flop1 and Flop2 inhibit Wnt/β-catenin signaling and are essential for head formation in Xenopus.

PubMed

Miyagi, Asuka; Negishi, Takefumi; Yamamoto, Takamasa S; Ueno, Naoto

2015-11-01

Patterning of the vertebrate anterior-posterior axis is regulated by the coordinated action of growth factors whose effects can be further modulated by upstream and downstream mediators and the cross-talk of different intracellular pathways. In particular, the inhibition of the Wnt/β-catenin signaling pathway by various factors is critically required for anterior specification. Here, we report that Flop1 and Flop2 (Flop1/2), G protein-coupled receptors related to Gpr4, contribute to the regulation of head formation by inhibiting Wnt/β-catenin signaling in Xenopus embryos. Using whole-mount in situ hybridization, we showed that flop1 and flop2 mRNAs were expressed in the neural ectoderm during early gastrulation. Both the overexpression and knockdown of Flop1/2 resulted in altered embryonic head phenotypes, while the overexpression of either Flop1/2 or the small GTPase RhoA in the absence of bone morphogenetic protein (BMP) signaling resulted in ectopic head induction. Examination of the Flops' function in Xenopus embryo animal cap cells showed that they inhibited Wnt/β-catenin signaling by promoting β-catenin degradation through both RhoA-dependent and -independent pathways in a cell-autonomous manner. These results suggest that Flop1 and Flop2 are essential regulators of Xenopus head formation that act as novel inhibitory components of the Wnt/β-catenin signaling pathway. Copyright © 2015 Elsevier Inc. All rights reserved.

Somatic polyploidy is associated with the upregulation of c-MYC interacting genes and EMT-like signature

PubMed Central

Vazquez-Martin, Alejandro; Anatskaya, Olga V.; Giuliani, Alessandro; Erenpreisa, Jekaterina; Huang, Sui; Salmina, Kristine; Inashkina, Inna; Huna, Anda; Nikolsky, Nikolai N.; Vinogradov, Alexander E.

2016-01-01

The dependence of cancer on overexpressed c-MYC and its predisposition for polyploidy represents a double puzzle. We address this conundrum by cross-species transcription analysis of c-MYC interacting genes in polyploid vs. diploid tissues and cells, including human vs. mouse heart, mouse vs. human liver and purified 4n vs. 2n mouse decidua cells. Gene-by-gene transcriptome comparison and principal component analysis indicated that c-MYC interactants are significantly overrepresented among ploidy-associated genes. Protein interaction networks and gene module analysis revealed that the most upregulated genes relate to growth, stress response, proliferation, stemness and unicellularity, as well as to the pathways of cancer supported by MAPK and RAS coordinated pathways. A surprising feature was the up-regulation of epithelial-mesenchymal transition (EMT) modules embodied by the N-cadherin pathway and EMT regulators from SNAIL and TWIST families. Metabolic pathway analysis also revealed the EMT-linked features, such as global proteome remodeling, oxidative stress, DNA repair and Warburg-like energy metabolism. Genes associated with apoptosis, immunity, energy demand and tumour suppression were mostly down-regulated. Noteworthy, despite the association between polyploidy and ample features of cancer, polyploidy does not trigger it. Possibly it occurs because normal polyploidy does not go that far in embryonalisation and linked genome destabilisation. In general, the analysis of polyploid transcriptome explained the evolutionary relation of c-MYC and polyploidy to cancer. PMID:27655693

Transition Pathway and Its Free-Energy Profile: A Protocol for Protein Folding Simulations

PubMed Central

Lee, In-Ho; Kim, Seung-Yeon; Lee, Jooyoung

2013-01-01

We propose a protocol that provides a systematic definition of reaction coordinate and related free-energy profile as the function of temperature for the protein-folding simulation. First, using action-derived molecular dynamics (ADMD), we investigate the dynamic folding pathway model of a protein between a fixed extended conformation and a compact conformation. We choose the pathway model to be the reaction coordinate, and the folding and unfolding processes are characterized by the ADMD step index, in contrast to the common a priori reaction coordinate as used in conventional studies. Second, we calculate free-energy profile as the function of temperature, by employing the replica-exchange molecular dynamics (REMD) method. The current method provides efficient exploration of conformational space and proper characterization of protein folding/unfolding dynamics from/to an arbitrary extended conformation. We demonstrate that combination of the two simulation methods, ADMD and REMD, provides understanding on molecular conformational changes in proteins. The protocol is tested on a small protein, penta-peptide of met-enkephalin. For the neuropeptide met-enkephalin system, folded, extended, and intermediate sates are well-defined through the free-energy profile over the reaction coordinate. Results are consistent with those in the literature. PMID:23917881

Arabidopsis EDS1 connects pathogen effector recognition to cell compartment-specific immune responses.

PubMed

Heidrich, Katharina; Wirthmueller, Lennart; Tasset, Céline; Pouzet, Cécile; Deslandes, Laurent; Parker, Jane E

2011-12-09

Pathogen effectors are intercepted by plant intracellular nucleotide binding-leucine-rich repeat (NB-LRR) receptors. However, processes linking receptor activation to downstream defenses remain obscure. Nucleo-cytoplasmic basal resistance regulator EDS1 (ENHANCED DISEASE SUSCEPTIBILITY1) is indispensible for immunity mediated by TIR (Toll-interleukin-1 receptor)-NB-LRR receptors. We show that Arabidopsis EDS1 molecularly connects TIR-NB-LRR disease resistance protein RPS4 recognition of bacterial effector AvrRps4 to defense pathways. RPS4-EDS1 and AvrRps4-EDS1 complexes are detected inside nuclei of living tobacco cells after transient coexpression and in Arabidopsis soluble leaf extracts after resistance activation. Forced AvrRps4 localization to the host cytoplasm or nucleus reveals cell compartment-specific RPS4-EDS1 defense branches. Although nuclear processes restrict bacterial growth, programmed cell death and transcriptional resistance reinforcement require nucleo-cytoplasmic coordination. Thus, EDS1 behaves as an effector target and activated TIR-NB-LRR signal transducer for defenses across cell compartments.

Abscisic Acid–Responsive Guard Cell Metabolomes of Arabidopsis Wild-Type and gpa1 G-Protein Mutants[C][W

PubMed Central

Jin, Xiaofen; Wang, Rui-Sheng; Zhu, Mengmeng; Jeon, Byeong Wook; Albert, Reka; Chen, Sixue; Assmann, Sarah M.

2013-01-01

Individual metabolites have been implicated in abscisic acid (ABA) signaling in guard cells, but a metabolite profile of this specialized cell type is lacking. We used liquid chromatography–multiple reaction monitoring mass spectrometry for targeted analysis of 85 signaling-related metabolites in Arabidopsis thaliana guard cell protoplasts over a time course of ABA treatment. The analysis utilized ∼350 million guard cell protoplasts from ∼30,000 plants of the Arabidopsis Columbia accession (Col) wild type and the heterotrimeric G-protein α subunit mutant, gpa1, which has ABA-hyposensitive stomata. These metabolomes revealed coordinated regulation of signaling metabolites in unrelated biochemical pathways. Metabolites clustered into different temporal modules in Col versus gpa1, with fewer metabolites showing ABA-altered profiles in gpa1. Ca2+-mobilizing agents sphingosine-1-phosphate and cyclic adenosine diphosphate ribose exhibited weaker ABA-stimulated increases in gpa1. Hormone metabolites were responsive to ABA, with generally greater responsiveness in Col than in gpa1. Most hormones also showed different ABA responses in guard cell versus mesophyll cell metabolomes. These findings suggest that ABA functions upstream to regulate other hormones, and are also consistent with G proteins modulating multiple hormonal signaling pathways. In particular, indole-3-acetic acid levels declined after ABA treatment in Col but not gpa1 guard cells. Consistent with this observation, the auxin antagonist α-(phenyl ethyl-2-one)-indole-3-acetic acid enhanced ABA-regulated stomatal movement and restored partial ABA sensitivity to gpa1. PMID:24368793

ONC201 induces cell death in pediatric non-Hodgkin's lymphoma cells

PubMed Central

Talekar, Mala K; Allen, Joshua E; Dicker, David T; El-Deiry, Wafik S

2015-01-01

ONC201/TIC10 is a small molecule initially discovered by its ability to coordinately induce and activate the TRAIL pathway selectively in tumor cells and has recently entered clinical trials in adult advanced cancers. The anti-tumor activity of ONC201 has previously been demonstrated in several preclinical models of cancer, including refractory solid tumors and a transgenic lymphoma mouse model. Based on the need for new safe and effective therapies in pediatric non-Hodgkin's lymphoma (NHL) and the non-toxic preclinical profile of ONC201, we investigated the in vitro efficacy of ONC201 in non-Hodgkin's lymphoma (NHL) cell lines to evaluate its therapeutic potential for this disease. ONC201 caused a dose-dependent reduction in the cell viability of NHL cell lines that resulted from induction of apoptosis. As expected from prior observations, induction of TRAIL and its receptor DR5 was also observed in these cell lines. Furthermore, dual induction of TRAIL and DR5 appeared to drive the observed apoptosis and TRAIL expression was correlated linearly with sub-G1 DNA content, suggesting its potential role as a biomarker of tumor response to ONC201-treated lymphoma cells. We further investigated combinations of ONC201 with approved chemotherapeutic agents used to treat lymphoma. ONC201 exhibited synergy in combination with the anti-metabolic agent cytarabine in vitro, in addition to cooperating with other therapies. Together these findings indicate that ONC201 is an effective TRAIL pathway-inducer as a monoagent that can be combined with chemotherapy to enhance therapeutic responses in pediatric NHL. PMID:26030065

ONC201 induces cell death in pediatric non-Hodgkin's lymphoma cells.

PubMed

Talekar, Mala K; Allen, Joshua E; Dicker, David T; El-Deiry, Wafik S

2015-08-03

ONC201/TIC10 is a small molecule initially discovered by its ability to coordinately induce and activate the TRAIL pathway selectively in tumor cells and has recently entered clinical trials in adult advanced cancers. The anti-tumor activity of ONC201 has previously been demonstrated in several preclinical models of cancer, including refractory solid tumors and a transgenic lymphoma mouse model. Based on the need for new safe and effective therapies in pediatric non-Hodgkin's lymphoma (NHL) and the non-toxic preclinical profile of ONC201, we investigated the in vitro efficacy of ONC201 in non-Hodgkin's lymphoma (NHL) cell lines to evaluate its therapeutic potential for this disease. ONC201 caused a dose-dependent reduction in the cell viability of NHL cell lines that resulted from induction of apoptosis. As expected from prior observations, induction of TRAIL and its receptor DR5 was also observed in these cell lines. Furthermore, dual induction of TRAIL and DR5 appeared to drive the observed apoptosis and TRAIL expression was correlated linearly with sub-G1 DNA content, suggesting its potential role as a biomarker of tumor response to ONC201-treated lymphoma cells. We further investigated combinations of ONC201 with approved chemotherapeutic agents used to treat lymphoma. ONC201 exhibited synergy in combination with the anti-metabolic agent cytarabine in vitro, in addition to cooperating with other therapies. Together these findings indicate that ONC201 is an effective TRAIL pathway-inducer as a monoagent that can be combined with chemotherapy to enhance therapeutic responses in pediatric NHL.

Control of DEMETER DNA demethylase gene transcription in male and female gamete companion cells in Arabidopsis thaliana

PubMed Central

Park, Jin-Sup; Frost, Jennifer M.; Park, Kyunghyuk; Ohr, Hyonhwa; Park, Guen Tae; Kim, Seohyun; Eom, Hyunjoo; Lee, Ilha; Brooks, Janie S.; Fischer, Robert L.; Choi, Yeonhee

2017-01-01

The DEMETER (DME) DNA glycosylase initiates active DNA demethylation via the base-excision repair pathway and is vital for reproduction in Arabidopsis thaliana. DME-mediated DNA demethylation is preferentially targeted to small, AT-rich, and nucleosome-depleted euchromatic transposable elements, influencing expression of adjacent genes and leading to imprinting in the endosperm. In the female gametophyte, DME expression and subsequent genome-wide DNA demethylation are confined to the companion cell of the egg, the central cell. Here, we show that, in the male gametophyte, DME expression is limited to the companion cell of sperm, the vegetative cell, and to a narrow window of time: immediately after separation of the companion cell lineage from the germline. We define transcriptional regulatory elements of DME using reporter genes, showing that a small region, which surprisingly lies within the DME gene, controls its expression in male and female companion cells. DME expression from this minimal promoter is sufficient to rescue seed abortion and the aberrant DNA methylome associated with the null dme-2 mutation. Within this minimal promoter, we found short, conserved enhancer sequences necessary for the transcriptional activities of DME and combined predicted binding motifs with published transcription factor binding coordinates to produce a list of candidate upstream pathway members in the genetic circuitry controlling DNA demethylation in gamete companion cells. These data show how DNA demethylation is regulated to facilitate endosperm gene imprinting and potential transgenerational epigenetic regulation, without subjecting the germline to potentially deleterious transposable element demethylation. PMID:28130550

Coordinated induction of GST and MRP2 by cAMP in Caco-2 cells: Role of protein kinase A signaling pathway and toxicological relevance

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

Arana, Maite Rocío, E-mail: arana@ifise-conicet.gov.ar; Tocchetti, Guillermo Nicolás, E-mail: gtocchetti@live.com.ar; Domizi, Pablo, E-mail: domizi@ibr-conicet.gov.ar

2015-09-01

The cAMP pathway is a universal signaling pathway regulating many cellular processes including metabolic routes, growth and differentiation. However, its effects on xenobiotic biotransformation and transport systems are poorly characterized. The effect of cAMP on expression and activity of GST and MRP2 was evaluated in Caco-2 cells, a model of intestinal epithelium. Cells incubated with the cAMP permeable analog dibutyryl cyclic AMP (db-cAMP: 1,10,100 μM) for 48 h exhibited a dose–response increase in GST class α and MRP2 protein expression. Incubation with forskolin, an activator of adenylyl cyclase, confirmed the association between intracellular cAMP and upregulation of MRP2. Consistent withmore » increased expression of GSTα and MRP2, db-cAMP enhanced their activities, as well as cytoprotection against the common substrate 1-chloro-2,4-dinitrobenzene. Pretreatment with protein kinase A (PKA) inhibitors totally abolished upregulation of MRP2 and GSTα induced by db-cAMP. In silico analysis together with experiments consisting of treatment with db-cAMP of Caco-2 cells transfected with a reporter construct containing CRE and AP-1 sites evidenced participation of these sites in MRP2 upregulation. Further studies involving the transcription factors CREB and AP-1 (c-JUN, c-FOS and ATF2) demonstrated increased levels of total c-JUN and phosphorylation of c-JUN and ATF2 by db-cAMP, which were suppressed by a PKA inhibitor. Co-immunoprecipitation and ChIP assay studies demonstrated that db-cAMP increased c-JUN/ATF2 interaction, with further recruitment to the region of the MRP2 promoter containing CRE and AP-1 sites. We conclude that cAMP induces GSTα and MRP2 expression and activity in Caco-2 cells via the PKA pathway, thus regulating detoxification of specific xenobiotics. - Highlights: • cAMP positively modulates the expression and activity of GST and MRP2 in Caco-2 cells. • Such induction resulted in increased cytoprotection against chemical injury. • PKA signaling pathway is involved downstream of cAMP. • Transcriptional MRP2 regulation ultimately involved participation of c-JUN and ATF2.« less

IP3R-mediated Ca2+ release regulates protein metabolism in Drosophila neuroendocrine cells: implications for development under nutrient stress.

PubMed

Megha; Hasan, Gaiti

2017-04-15

Successful completion of animal development is fundamentally reliant on nutritional cues. Surviving periods of nutritional insufficiency requires adaptations that are coordinated, in part, by neural circuits. As neuropeptides secreted by neuroendocrine (NE) cells modulate neural circuits, we investigated NE cell function during development under nutrient stress. Starved Drosophila larvae exhibited reduced pupariation if either insulin signaling or IP 3 /Ca 2+ signaling were downregulated in NE cells. Moreover, an IP 3 R (inositol 1,4,5-trisphosphate receptor) loss-of-function mutant displayed reduced protein synthesis, which was rescued by overexpression of either InR (insulin receptor) or IP 3 R in NE cells of the mutant, suggesting that the two signaling pathways might be functionally compensatory. Furthermore, cultured IP 3 R mutant NE cells, but not neurons, exhibited reduced protein translation. Thus cell-specific regulation of protein synthesis by IP 3 R in NE cells influences protein metabolism. We propose that this regulation helps developing animals survive in poor nutritional conditions. © 2017. Published by The Company of Biologists Ltd.

Metabolic mysteries of the inflammatory response: T cell polarization and plasticity.

PubMed

Fracchia, Kelley M; Walsh, Craig M

2015-01-01

While simultaneously maintaining homeostasis and reducing further harm to the host, the immune system is equipped to eliminate both tumors and pathogenic microorganisms. Bifurcated into cell-mediated and humoral immunity, the adaptive immune system requires a series of complex and coordinated signals to drive the proliferation and differentiation of appropriate subsets. These include signals that modulate cellular metabolism. When first published in the 1920s, "the Warburg effect" was used to describe a phenomenon in which most cancer cells relied on aerobic glycolysis to meet their biosynthetic demands. Despite the early observations of Warburg and his colleagues, targeting cancer cell metabolism for therapeutic purposes still remains theoretical. Notably, many T cells exhibit the same Warburg metabolism as cancer cells and the therapeutic benefit of targeting their metabolic pathways has since been reexamined. Emerging evidence suggests that specific metabolic alterations associated with T cells may be ancillary to their subset differentiation and influential in their inflammatory response. Thus, T cell lymphocyte activation leads to skewing in metabolic plasticity, and issue that will be the subject of this review.

Remodeling the zonula adherens in response to tension and the role of afadin in this response

PubMed Central

Acharya, Bipul R.; Peyret, Grégoire; Fardin, Marc-Antoine; Mège, René-Marc; Ladoux, Benoit; Yap, Alpha S.; Fanning, Alan S.

2016-01-01

Morphogenesis requires dynamic coordination between cell–cell adhesion and the cytoskeleton to allow cells to change shape and move without losing tissue integrity. We used genetic tools and superresolution microscopy in a simple model epithelial cell line to define how the molecular architecture of cell–cell zonula adherens (ZA) is modified in response to elevated contractility, and how these cells maintain tissue integrity. We previously found that depleting zonula occludens 1 (ZO-1) family proteins in MDCK cells induces a highly organized contractile actomyosin array at the ZA. We find that ZO knockdown elevates contractility via a Shroom3/Rho-associated, coiled-coil containing protein kinase (ROCK) pathway. Our data suggest that each bicellular border is an independent contractile unit, with actin cables anchored end-on to cadherin complexes at tricellular junctions. Cells respond to elevated contractility by increasing junctional afadin. Although ZO/afadin knockdown did not prevent contractile array assembly, it dramatically altered cell shape and barrier function in response to elevated contractility. We propose that afadin acts as a robust protein scaffold that maintains ZA architecture at tricellular junctions. PMID:27114502

Rab coupling protein mediated endosomal recycling of N-cadherin influences cell motility.

PubMed

Lindsay, Andrew J; McCaffrey, Mary W

2017-12-01

Rab coupling protein (RCP) is a Rab GTPase effector that functions in endosomal recycling. The RCP gene is frequently amplified in breast cancer, leading to increased cancer aggressiveness. Furthermore, RCP enhances the motility of ovarian cancer cells by coordinating the recycling of α5β1 integrin and EGF receptor to the leading edge of migrating cells. Here we report that RCP also influences the motility of lung adenocarcinoma cells. Knockdown of RCP inhibits the motility of A549 cells in 2D and 3D migration assays, while its overexpression enhances migration in these assays. Depletion of RCP leads to a reduction in N-cadherin protein levels, which could be restored with lysosomal inhibitors. Trafficking assays revealed that RCP knockdown inhibits the return of endocytosed N-cadherin to the cell surface. We propose that RCP regulates the endosomal recycling of N-cadherin, and in its absence N-cadherin is diverted to the degradative pathway. The increased aggressiveness of tumour cells that overexpress RCP may be due to biased recycling of N-cadherin in metastatic cancer cells.

Genome-wide pathway analysis of memory impairment in the Alzheimer's Disease Neuroimaging Initiative (ADNI) cohort implicates gene candidates, canonical pathways, and networks.

PubMed

Ramanan, Vijay K; Kim, Sungeun; Holohan, Kelly; Shen, Li; Nho, Kwangsik; Risacher, Shannon L; Foroud, Tatiana M; Mukherjee, Shubhabrata; Crane, Paul K; Aisen, Paul S; Petersen, Ronald C; Weiner, Michael W; Saykin, Andrew J

2012-12-01

Memory deficits are prominent features of mild cognitive impairment (MCI) and Alzheimer's disease (AD). The genetic architecture underlying these memory deficits likely involves the combined effects of multiple genetic variants operative within numerous biological pathways. In order to identify functional pathways associated with memory impairment, we performed a pathway enrichment analysis on genome-wide association data from 742 Alzheimer's Disease Neuroimaging Initiative (ADNI) participants. A composite measure of memory was generated as the phenotype for this analysis by applying modern psychometric theory to item-level data from the ADNI neuropsychological test battery. Using the GSA-SNP software tool, we identified 27 canonical, expertly-curated pathways with enrichment (FDR-corrected p-value < 0.05) against this composite memory score. Processes classically understood to be involved in memory consolidation, such as neurotransmitter receptor-mediated calcium signaling and long-term potentiation, were highly represented among the enriched pathways. In addition, pathways related to cell adhesion, neuronal differentiation and guided outgrowth, and glucose- and inflammation-related signaling were also enriched. Among genes that were highly-represented in these enriched pathways, we found indications of coordinated relationships, including one large gene set that is subject to regulation by the SP1 transcription factor, and another set that displays co-localized expression in normal brain tissue along with known AD risk genes. These results 1) demonstrate that psychometrically-derived composite memory scores are an effective phenotype for genetic investigations of memory impairment and 2) highlight the promise of pathway analysis in elucidating key mechanistic targets for future studies and for therapeutic interventions.

α7 Nicotinic Acetylcholine Receptor (α7nAChR) Expression in Bone Marrow–Derived Non–T Cells Is Required for the Inflammatory Reflex

PubMed Central

Olofsson, Peder S; Katz, David A; Rosas-Ballina, Mauricio; Levine, Yaakov A; Ochani, Mahendar; Valdés-Ferrer, Sergio I; Pavlov, Valentin A; Tracey, Kevin J; Chavan, Sangeeta S

2012-01-01

The immune response to infection or injury coordinates host defense and tissue repair, but also has the capacity to damage host tissues. Recent advances in understanding protective mechanisms have found neural circuits that suppress release of damaging cytokines. Stimulation of the vagus nerve protects from excessive cytokine production and ameliorates experimental inflammatory disease. This mechanism, the inflammatory reflex, requires the α7 nicotinic acetylcholine receptor (α7nAChR), a ligand-gated ion channel expressed on macrophages, lymphocytes, neurons and other cells. To investigate cell-specific function of α7nAChR in the inflammatory reflex, we created chimeric mice by cross-transferring bone marrow between wild-type (WT) and α7nAChR-deficient mice. Deficiency of α7nAChR in bone marrow–derived cells significantly impaired vagus nerve–mediated regulation of tumor necrosis factor (TNF), whereas α7nAChR deficiency in neurons and other cells had no significant effect. In agreement with recent work, the inflammatory reflex was not functional in nude mice, because functional T cells are required for the integrity of the pathway. To investigate the role of T-cell α7nAChR, we adoptively transferred α7nAChR-deficient or WT T cells to nude mice. Transfer of WT and α7nAChR-deficient T cells restored function, indicating that α7nAChR expression on T cells is not necessary for this pathway. Together, these results indicate that α7nAChR expression in bone marrow–derived non–T cells is required for the integrity of the inflammatory reflex. PMID:22183893

Pathways for external alkalinization in intact and in microwounded Chara cells are differentially sensitive to wortmannin

PubMed Central

Bulychev, Alexander A.; Foissner, Ilse

2017-01-01

ABSTRACT Proton flows across the plant cell membranes play a major role in electrogenesis and regulation of photosynthesis and ion balance. The profiles of external pH along the illuminated internodal cells of characean algae consist of alternating high- and low-pH zones that are spatially coordinated with the distribution of photosynthetic activity of chloroplasts underlying these zones. The results based on confocal laser scanning fluorescence microscopy, pH microsensors, and pulse-amplitude-modulated chlorophyll microfluorometry revealed that the coordination of H+ transport and photosynthesis is disrupted by the 2 different environmental cues (low light and wounding) and by a chemical, wortmannin interfering with the inositol phospholipid metabolism. On the one hand, the transition from moderate to low irradiance diminished the peaks in the profiles of photosystem II (PSII) quantum efficiency but did not remove the pH bands. On the other hand, the microwounding of the internode with a glass micropipette, impacting primarily the cell wall, resulted in a rapid local alkalinization of the external medium (by 2–2.5 pH units) near the cell surface, thus mimicking the appearance of natural pH bands. Despite their seeming similarity, the alkaline bands of intact cells were eliminated by wortmannin, whereas the wound-induced alkalinization was insensitive to this drug. Furthermore, the attenuation of natural pH bands in wortmannin-treated cells was accompanied by the enhancement in spatial heterogeneity of PSII efficiency and electron transport rates, which indicates the complexity of chloroplast–plasma membrane interactions. The results suggest that the light- and wound-induced alkaline areas on the cell surface are associated with different ion-transport systems. PMID:28805493

Anodal Cerebellar Direct Current Stimulation Reduces Facilitation of Propriospinal Neurons in Healthy Humans.

PubMed

Chothia, Muhammed; Doeltgen, Sebastian; Bradnam, Lynley V

2016-01-01

Coordinated muscle synergies in the human upper limb are controlled, in part, by a neural distribution network located in the cervical spinal cord, known as the cervical propriospinal system. Studies in the cat and non-human primate indicate the cerebellum is indirectly connected to this system via output pathways to the brainstem. Therefore, the cerebellum may indirectly modulate excitability of putative propriospinal neurons (PNs) in humans during upper limb coordination tasks. This study aimed to test whether anodal direct current stimulation (DCS) of the cerebellum modulates PNs and upper limb coordination in healthy adults. The hypothesis was that cerebellar anodal DCS would reduce descending facilitation of PNs and improve upper limb coordination. Transcranial magnetic stimulation (TMS), paired with peripheral nerve stimulation, probed activity in facilitatory and inhibitory descending projections to PNs following an established protocol. Coordination was tested using a pursuit rotor task performed by the non-dominant (ipsilateral) hand. Anodal and sham DCS were delivered over the cerebellum ipsilateral to the non-dominant hand in separate experimental sessions. Anodal DCS was applied to a control site lateral to the vertex in a third session. Twelve right-handed healthy adults participated. Pairing TMS with sub-threshold peripheral nerve stimulation facilitated motor evoked potentials at intensities just above threshold in accordance with the protocol. Anodal cerebellar DCS reduced facilitation without influencing inhibition, but the reduction in facilitation was not associated with performance of the pursuit rotor task. The results of this study indicate dissociated indirect control over cervical PNs by the cerebellum in humans. Anodal DCS of the cerebellum reduced excitability in the facilitatory descending pathway with no effect on the inhibitory pathway to cervical PNs. The reduction in PN excitability is likely secondary to modulation of primary motor cortex or brainstem nuclei, and identifies a neuroanatomical pathway for the cerebellum to assist in coordination of upper limb muscle synergies in humans. Copyright © 2016 Elsevier Inc. All rights reserved.

Mitochondrial quality control: Easy come, easy go

PubMed Central

Stotland, Aleksandr; Gottlieb, Roberta A.

2015-01-01

“Friends come and go but enemies accumulate.”Arthur Bloch Mitochondrial networks in eukaryotic cells are maintained via regular cycles of degradation and biogenesis. These complex processes function in concert with one another to eliminate dysfunctional mitochondria in a specific and targeted manner and coordinate the biogenesis of new organelles. This review covers the two aspects of mitochondrial turnover, focusing on the main pathways and mechanisms involved. The review also summarizes the current methods and techniques for analyzing mitochondrial turnover in vivo and in vitro, from the whole animal proteome level to the level of single organelle. PMID:25596427

Reprogramming of gene expression during compression wood formation in pine: Coordinated modulation of S-adenosylmethionine, lignin and lignan related genes

PubMed Central

2012-01-01

Background Transcript profiling of differentiating secondary xylem has allowed us to draw a general picture of the genes involved in wood formation. However, our knowledge is still limited about the regulatory mechanisms that coordinate and modulate the different pathways providing substrates during xylogenesis. The development of compression wood in conifers constitutes an exceptional model for these studies. Although differential expression of a few genes in differentiating compression wood compared to normal or opposite wood has been reported, the broad range of features that distinguish this reaction wood suggest that the expression of a larger set of genes would be modified. Results By combining the construction of different cDNA libraries with microarray analyses we have identified a total of 496 genes in maritime pine (Pinus pinaster, Ait.) that change in expression during differentiation of compression wood (331 up-regulated and 165 down-regulated compared to opposite wood). Samples from different provenances collected in different years and geographic locations were integrated into the analyses to mitigate the effects of multiple sources of variability. This strategy allowed us to define a group of genes that are consistently associated with compression wood formation. Correlating with the deposition of a thicker secondary cell wall that characterizes compression wood development, the expression of a number of genes involved in synthesis of cellulose, hemicellulose, lignin and lignans was up-regulated. Further analysis of a set of these genes involved in S-adenosylmethionine metabolism, ammonium recycling, and lignin and lignans biosynthesis showed changes in expression levels in parallel to the levels of lignin accumulation in cells undergoing xylogenesis in vivo and in vitro. Conclusions The comparative transcriptomic analysis reported here have revealed a broad spectrum of coordinated transcriptional modulation of genes involved in biosynthesis of different cell wall polymers associated with within-tree variations in pine wood structure and composition. In particular, we demonstrate the coordinated modulation at transcriptional level of a gene set involved in S-adenosylmethionine synthesis and ammonium assimilation with increased demand for coniferyl alcohol for lignin and lignan synthesis, enabling a better understanding of the metabolic requirements in cells undergoing lignification. PMID:22747794

Global functional analyses of cellular responses to pore-forming toxins.

PubMed

Kao, Cheng-Yuan; Los, Ferdinand C O; Huffman, Danielle L; Wachi, Shinichiro; Kloft, Nicole; Husmann, Matthias; Karabrahimi, Valbona; Schwartz, Jean-Louis; Bellier, Audrey; Ha, Christine; Sagong, Youn; Fan, Hui; Ghosh, Partho; Hsieh, Mindy; Hsu, Chih-Shen; Chen, Li; Aroian, Raffi V

2011-03-01

Here we present the first global functional analysis of cellular responses to pore-forming toxins (PFTs). PFTs are uniquely important bacterial virulence factors, comprising the single largest class of bacterial protein toxins and being important for the pathogenesis in humans of many Gram positive and Gram negative bacteria. Their mode of action is deceptively simple, poking holes in the plasma membrane of cells. The scattered studies to date of PFT-host cell interactions indicate a handful of genes are involved in cellular defenses to PFTs. How many genes are involved in cellular defenses against PFTs and how cellular defenses are coordinated are unknown. To address these questions, we performed the first genome-wide RNA interference (RNAi) screen for genes that, when knocked down, result in hypersensitivity to a PFT. This screen identifies 106 genes (∼0.5% of genome) in seven functional groups that protect Caenorhabditis elegans from PFT attack. Interactome analyses of these 106 genes suggest that two previously identified mitogen-activated protein kinase (MAPK) pathways, one (p38) studied in detail and the other (JNK) not, form a core PFT defense network. Additional microarray, real-time PCR, and functional studies reveal that the JNK MAPK pathway, but not the p38 MAPK pathway, is a key central regulator of PFT-induced transcriptional and functional responses. We find C. elegans activator protein 1 (AP-1; c-jun, c-fos) is a downstream target of the JNK-mediated PFT protection pathway, protects C. elegans against both small-pore and large-pore PFTs and protects human cells against a large-pore PFT. This in vivo RNAi genomic study of PFT responses proves that cellular commitment to PFT defenses is enormous, demonstrates the JNK MAPK pathway as a key regulator of transcriptionally-induced PFT defenses, and identifies AP-1 as the first cellular component broadly important for defense against large- and small-pore PFTs.

Spatial and Temporal Eye–Hand Coordination Relies on the Parietal Reach Region

PubMed Central

Hauschild, Markus; Wilke, Melanie; Andersen, Richard A.

2014-01-01

Coordinated eye movements are crucial for precision control of our hands. A commonly believed neural mechanism underlying eye–hand coordination is interaction between the neural networks controlling each effector, exchanging, and matching information, such as movement target location and onset time. Alternatively, eye–hand coordination may result simply from common inputs to independent eye and hand control pathways. Thus far, it remains unknown whether and where either of these two possible mechanisms exists. A candidate location for the former mechanism, interpathway communication, includes the posterior parietal cortex (PPC) where distinct effector-specific areas reside. If the PPC were within the network for eye–hand coordination, perturbing it would affect both eye and hand movements that are concurrently planned. In contrast, if eye–hand coordination arises solely from common inputs, perturbing one effector pathway, e.g., the parietal reach region (PRR), would not affect the other effector. To test these hypotheses, we inactivated part of PRR in the macaque, located in the medial bank of the intraparietal sulcus encompassing the medial intraparietal area and area 5V. When each effector moved alone, PRR inactivation shortened reach but not saccade amplitudes, compatible with the known reach-selective activity of PRR. However, when both effectors moved concurrently, PRR inactivation shortened both reach and saccade amplitudes, and decoupled their reaction times. Therefore, consistent with the interpathway communication hypothesis, we propose that the planning of concurrent eye and hand movements causes the spatial information in PRR to influence the otherwise independent eye control pathways, and that their temporal coupling requires an intact PRR. PMID:25232123

A topological coordinate system for the diamond cubic grid.

PubMed

Čomić, Lidija; Nagy, Benedek

2016-09-01

Topological coordinate systems are used to address all cells of abstract cell complexes. In this paper, a topological coordinate system for cells in the diamond cubic grid is presented and some of its properties are detailed. Four dependent coordinates are used to address the voxels (triakis truncated tetrahedra), their faces (hexagons and triangles), their edges and the points at their corners. Boundary and co-boundary relations, as well as adjacency relations between the cells, can easily be captured by the coordinate values. Thus, this coordinate system is apt for implementation in various applications, such as visualizations, morphological and topological operations and shape analysis.

Coordinating structural and functional synapse development: postsynaptic p21-activated kinase independently specifies glutamate receptor abundance and postsynaptic morphology.

PubMed

Albin, Stephanie D; Davis, Graeme W

2004-08-04

Here, we show that postsynaptic p21-activated kinase (Pak) signaling diverges into two genetically separable pathways at the Drosophila neuromuscular junction. One pathway controls glutamate receptor abundance. Pak signaling within this pathway is specified by a required interaction with the adaptor protein Dreadlocks (Dock). We demonstrate that Dock is localized to the synapse via an Src homology 2-mediated protein interaction. Dock is not necessary for Pak localization but is necessary to restrict Pak signaling to control glutamate receptor abundance. A second genetically separable function of Pak kinase signaling controls muscle membrane specialization through the regulation of synaptic Discs-large. In this pathway, Dock is dispensable. We present a model in which divergent Pak signaling is able to coordinate two different features of postsynaptic maturation, receptor abundance, and muscle membrane specialization.

Bone Marrow Derived Myeloid Cells Orchestrate Antiangiogenic Resistance in Glioblastoma through Coordinated Molecular Networks

PubMed Central

Achyut, B.R.; Shankar, Adarsh; Iskander, ASM; Ara, Roxan; Angara, Kartik; Zeng, Peng; Knight, Robert A.; Scicli, Alfonso G; Arbab, Ali S.

2015-01-01

Glioblastoma (GBM) is a hypervascular and malignant form of brain tumors. Anti-angiogenic therapies (AAT) were used as an adjuvant against VEGF-VEGFR pathway to normalize blood vessels in clinical and preclinical studies, which resulted into marked hypoxia and recruited bone marrow derived cells (BMDCs) to the tumor microenvironment (TME). In vivo animal models to track BMDCs and investigate molecular mechanisms in AAT resistance are rare. We exploited recently established chimeric mouse to develop orthotopic U251 tumor, which uses as low as 5×106 GFP+ BM cells in athymic nude mice and engrafted >70% GFP+ cells within 14 days. Our unpublished data and published studies have indicated the involvement of immunosuppressive myeloid cells in therapeutic resistance in glioma. Similarly, in the present study, vatalanib significantly increased CD68+ myeloid cells, and CD133+, CD34+ and Tie2+ endothelial cell signatures. Therefore, we tested inhibition of CSF1R+ myeloid cells using GW2580 that reduced tumor growth by decreasing myeloid (Gr1+ CD11b+ and F4/80+) and angiogenic (CD202b+ and VEGFR2+) cell signatures in TME. CSF1R blockade significantly decreased inflammatory, proangiogenic and immunosuppressive molecular signatures compared to vehicle, vatalanib or combination. TCK1 or CXCL7, a potent chemoattractant and activator of neutrophils, was observed as most significantly decreased cytokine in CSF1R blockade. ERK MAPK pathway was involved in cytokine network regulation. In conclusion, present study confirmed the contribution of myeloid cells in GBM development and therapeutic resistance using chimeric mouse model. We identified novel molecular networks including CXCL7 chemokine as a promising target for future studies. Nonetheless, survival studies are required to assess the beneficial effect of CSF1R blockade. PMID:26404753

The FANC pathway is activated by adenovirus infection and promotes viral replication-dependent recombination

PubMed Central

Cherubini, Gioia; Naim, Valeria; Caruso, Paola; Burla, Romina; Bogliolo, Massimo; Cundari, Enrico; Benihoud, Karim; Saggio, Isabella; Rosselli, Filippo

2011-01-01

Deciphering the crosstalk between a host cell and a virus during infection is important not only to better define viral biology but also to improve our understanding of cellular processes. We identified the FANC pathway as a helper of viral replication and recombination by searching for cellular targets that are modified by adenovirus (Ad) infection and are involved in its outcome. This pathway, which is involved in the DNA damage response and checkpoint control, is altered in Fanconi anaemia, a rare cancer predisposition syndrome. We show here that Ad5 infection activates the FANC pathway independent of the classical DNA damage response. Infection with a non-replicating Ad shows that the presence of viral DNA is not sufficient to induce the monoubiquitination of FANCD2 but still activates the DNA damage response coordinated by phospho-NBS1 and phospho-CHK1. E1A expression alone fails to induce FANCD2 monoubiquitination, indicating that a productive viral infection and/or replication is required for FANC pathway activation. Our data indicate that Ad5 infection induces FANCD2 activation to promote its own replication. Specifically, we show that FANCD2 is involved in the recombination process that accompanies viral DNA replication. This study provides evidence of a DNA damage-independent function of the FANC pathway and identifies a cellular system involved in Ad5 recombination. PMID:21421559

Activation of YUCCA5 by the Transcription Factor TCP4 Integrates Developmental and Environmental Signals to Promote Hypocotyl Elongation in Arabidopsis.

PubMed

Challa, Krishna Reddy; Aggarwal, Pooja; Nath, Utpal

2016-09-05

Cell expansion is an essential process in plant morphogenesis and is regulated by the coordinated action of environmental stimuli and endogenous factors, such as the phytohormones auxin and brassinosteroid. Although the biosynthetic pathways that generate these hormones and their downstream signaling mechanisms have been extensively studied, the upstream transcriptional network that modulates their levels and connects their action to cell morphogenesis is less clear. Here we show that the miR319-regulated TCP (TEOSINTE BRANCHED 1, CYCLODEA, PROLIFERATING CELL FACTORS) transcription factors, notably TCP4, directly activate YUCCA5 transcription and integrate the auxin response to a brassinosteroid-dependent molecular circuit that promotes cell elongation in Arabidopsis hypocotyls. Further, TCP4 modulates the common transcriptional network downstream to auxin-BR signaling, which is also triggered by environmental cues, such as light, to promote cell expansion. Our study links TCP function with the hormone response during cell morphogenesis and shows that developmental and environmental signals converge on a common transcriptional network to promote cell elongation. {copyright, serif} 2016 American Society of Plant Biologists. All rights reserved.

Regulation of podocalyxin trafficking by Rab small GTPases in epithelial cells

PubMed Central

Mrozowska, Paulina S.; Fukuda, Mitsunori

2016-01-01

ABSTRACT The characteristic feature of polarity establishment in MDCK II cells is transcytosis of apical glycoprotein podocalyxin (PCX) from the outer plasma membrane to the newly formed apical domain. This transcytotic event consists of multiple steps, including internalization from the plasma membrane, transport through early endosomes and Rab11-positive recycling endosomes, and delivery to the apical membrane. These steps are known to be tightly coordinated by Rab small GTPases, which act as molecular switches cycling between active GTP-bound and inactive GDP-bound states. However, our knowledge regarding which sets of Rabs regulate particular steps of PCX trafficking was rather limited. Recently, we have performed a comprehensive analysis of Rab GTPase engagement in the transcytotic pathway of PCX during polarity establishment in 2-dimensional (2D) and 3-dimensional (3D) MDCK II cell cultures. In this Commentary we summarize our findings and set them in the context of previous reports. PMID:27463697

Specificity, cross-talk and adaptation in Interferon signaling

NASA Astrophysics Data System (ADS)

Zilman, Anton

Innate immune system is the first line of defense of higher organisms against pathogens. It coordinates the behavior of millions of cells of multiple types, achieved through numerous signaling molecules. This talk focuses on the signaling specificity of a major class of signaling molecules - Type I Interferons - which are also used therapeutically in the treatment of a number of diseases, such as Hepatitis C, multiple sclerosis and some cancers. Puzzlingly, different Interferons act through the same cell surface receptor but have different effects on the target cells. They also exhibit a strange pattern of temporal cross-talk resulting in a serious clinical problem - loss of response to Interferon therapy. We combined mathematical modeling with quantitative experiments to develop a quantitative model of specificity and adaptation in the Interferon signaling pathway. The model resolves several outstanding experimental puzzles and directly affects the clinical use of Type I Interferons in treatment of viral hepatitis and other diseases.

Rho and Ras GTPases in Axon Growth, Guidance, and Branching

PubMed Central

Hall, Alan; Lalli, Giovanna

2010-01-01

The establishment of precise neuronal cell morphology provides the foundation for all aspects of neurobiology. During development, axons emerge from cell bodies after an initial polarization stage, elongate, and navigate towards target regions guided by a range of environmental cues. The Rho and Ras families of small GTPases have emerged as critical players at all stages of axonogenesis. Their ability to coordinately direct multiple signal transduction pathways with precise spatial control drives many of the activities that underlie this morphogenetic program: the dynamic assembly, disassembly, and reorganization of the actin and microtubule cytoskeletons, the interaction of the growing axon with other cells and extracellular matrix, the delivery of lipids and proteins to the axon through the exocytic machinery, and the internalization of membrane and proteins at the leading edge of the growth cone through endocytosis. This article highlights the contribution of Rho and Ras GTPases to axonogenesis. PMID:20182621

Cell-autonomous mechanisms of chronological aging in the yeast Saccharomyces cerevisiae.

PubMed

Arlia-Ciommo, Anthony; Leonov, Anna; Piano, Amanda; Svistkova, Veronika; Titorenko, Vladimir I

2014-05-27

A body of evidence supports the view that the signaling pathways governing cellular aging - as well as mechanisms of their modulation by longevity-extending genetic, dietary and pharmacological interventions - are conserved across species. The scope of this review is to critically analyze recent advances in our understanding of cell-autonomous mechanisms of chronological aging in the budding yeast Saccharomyces cerevisiae . Based on our analysis, we propose a concept of a biomolecular network underlying the chronology of cellular aging in yeast. The concept posits that such network progresses through a series of lifespan checkpoints. At each of these checkpoints, the intracellular concentrations of some key intermediates and products of certain metabolic pathways - as well as the rates of coordinated flow of such metabolites within an intricate network of intercompartmental communications - are monitored by some checkpoint-specific "master regulator" proteins. The concept envisions that a synergistic action of these master regulator proteins at certain early-life and late-life checkpoints modulates the rates and efficiencies of progression of such processes as cell metabolism, growth, proliferation, stress resistance, macromolecular homeostasis, survival and death. The concept predicts that, by modulating these vital cellular processes throughout lifespan (i.e., prior to an arrest of cell growth and division, and following such arrest), the checkpoint-specific master regulator proteins orchestrate the development and maintenance of a pro- or anti-aging cellular pattern and, thus, define longevity of chronologically aging yeast.

Antiviral Type I and Type III Interferon Responses in the Central Nervous System

PubMed Central

Sorgeloos, Frédéric; Kreit, Marguerite; Hermant, Pascale; Lardinois, Cécile; Michiels, Thomas

2013-01-01

The central nervous system (CNS) harbors highly differentiated cells, such as neurons that are essential to coordinate the functions of complex organisms. This organ is partly protected by the blood-brain barrier (BBB) from toxic substances and pathogens carried in the bloodstream. Yet, neurotropic viruses can reach the CNS either by crossing the BBB after viremia, or by exploiting motile infected cells as Trojan horses, or by using axonal transport. Type I and type III interferons (IFNs) are cytokines that are critical to control early steps of viral infections. Deficiencies in the IFN pathway have been associated with fatal viral encephalitis both in humans and mice. Therefore, the IFN system provides an essential protection of the CNS against viral infections. Yet, basal activity of the IFN system appears to be low within the CNS, likely owing to the toxicity of IFN to this organ. Moreover, after viral infection, neurons and oligodendrocytes were reported to be relatively poor IFN producers and appear to keep some susceptibility to neurotropic viruses, even in the presence of IFN. This review addresses some trends and recent developments concerning the role of type I and type III IFNs in: i) preventing neuroinvasion and infection of CNS cells; ii) the identity of IFN-producing cells in the CNS; iii) the antiviral activity of ISGs; and iv) the activity of viral proteins of neurotropic viruses that target the IFN pathway. PMID:23503326

Antiviral type I and type III interferon responses in the central nervous system.

PubMed

Sorgeloos, Frédéric; Kreit, Marguerite; Hermant, Pascale; Lardinois, Cécile; Michiels, Thomas

2013-03-15

The central nervous system (CNS) harbors highly differentiated cells, such as neurons that are essential to coordinate the functions of complex organisms. This organ is partly protected by the blood-brain barrier (BBB) from toxic substances and pathogens carried in the bloodstream. Yet, neurotropic viruses can reach the CNS either by crossing the BBB after viremia, or by exploiting motile infected cells as Trojan horses, or by using axonal transport. Type I and type III interferons (IFNs) are cytokines that are critical to control early steps of viral infections. Deficiencies in the IFN pathway have been associated with fatal viral encephalitis both in humans and mice. Therefore, the IFN system provides an essential protection of the CNS against viral infections. Yet, basal activity of the IFN system appears to be low within the CNS, likely owing to the toxicity of IFN to this organ. Moreover, after viral infection, neurons and oligodendrocytes were reported to be relatively poor IFN producers and appear to keep some susceptibility to neurotropic viruses, even in the presence of IFN. This review addresses some trends and recent developments concerning the role of type I and type III IFNs in: i) preventing neuroinvasion and infection of CNS cells; ii) the identity of IFN-producing cells in the CNS; iii) the antiviral activity of ISGs; and iv) the activity of viral proteins of neurotropic viruses that target the IFN pathway.

Pancreas lineage allocation and specification are regulated by sphingosine-1-phosphate signalling.

PubMed

Serafimidis, Ioannis; Rodriguez-Aznar, Eva; Lesche, Mathias; Yoshioka, Kazuaki; Takuwa, Yoh; Dahl, Andreas; Pan, Duojia; Gavalas, Anthony

2017-03-01

During development, progenitor expansion, lineage allocation, and implementation of differentiation programs need to be tightly coordinated so that different cell types are generated in the correct numbers for appropriate tissue size and function. Pancreatic dysfunction results in some of the most debilitating and fatal diseases, including pancreatic cancer and diabetes. Several transcription factors regulating pancreas lineage specification have been identified, and Notch signalling has been implicated in lineage allocation, but it remains unclear how these processes are coordinated. Using a combination of genetic approaches, organotypic cultures of embryonic pancreata, and genomics, we found that sphingosine-1-phosphate (S1p), signalling through the G protein coupled receptor (GPCR) S1pr2, plays a key role in pancreas development linking lineage allocation and specification. S1pr2 signalling promotes progenitor survival as well as acinar and endocrine specification. S1pr2-mediated stabilisation of the yes-associated protein (YAP) is essential for endocrine specification, thus linking a regulator of progenitor growth with specification. YAP stabilisation and endocrine cell specification rely on Gαi subunits, revealing an unexpected specificity of selected GPCR intracellular signalling components. Finally, we found that S1pr2 signalling posttranscriptionally attenuates Notch signalling levels, thus regulating lineage allocation. Both S1pr2-mediated YAP stabilisation and Notch attenuation are necessary for the specification of the endocrine lineage. These findings identify S1p signalling as a novel key pathway coordinating cell survival, lineage allocation, and specification and linking these processes by regulating YAP levels and Notch signalling. Understanding lineage allocation and specification in the pancreas will shed light in the origins of pancreatic diseases and may suggest novel therapeutic approaches.

Pancreas lineage allocation and specification are regulated by sphingosine-1-phosphate signalling

PubMed Central

Serafimidis, Ioannis; Rodriguez-Aznar, Eva; Lesche, Mathias; Yoshioka, Kazuaki; Takuwa, Yoh; Dahl, Andreas; Pan, Duojia; Gavalas, Anthony

2017-01-01

During development, progenitor expansion, lineage allocation, and implementation of differentiation programs need to be tightly coordinated so that different cell types are generated in the correct numbers for appropriate tissue size and function. Pancreatic dysfunction results in some of the most debilitating and fatal diseases, including pancreatic cancer and diabetes. Several transcription factors regulating pancreas lineage specification have been identified, and Notch signalling has been implicated in lineage allocation, but it remains unclear how these processes are coordinated. Using a combination of genetic approaches, organotypic cultures of embryonic pancreata, and genomics, we found that sphingosine-1-phosphate (S1p), signalling through the G protein coupled receptor (GPCR) S1pr2, plays a key role in pancreas development linking lineage allocation and specification. S1pr2 signalling promotes progenitor survival as well as acinar and endocrine specification. S1pr2-mediated stabilisation of the yes-associated protein (YAP) is essential for endocrine specification, thus linking a regulator of progenitor growth with specification. YAP stabilisation and endocrine cell specification rely on Gαi subunits, revealing an unexpected specificity of selected GPCR intracellular signalling components. Finally, we found that S1pr2 signalling posttranscriptionally attenuates Notch signalling levels, thus regulating lineage allocation. Both S1pr2-mediated YAP stabilisation and Notch attenuation are necessary for the specification of the endocrine lineage. These findings identify S1p signalling as a novel key pathway coordinating cell survival, lineage allocation, and specification and linking these processes by regulating YAP levels and Notch signalling. Understanding lineage allocation and specification in the pancreas will shed light in the origins of pancreatic diseases and may suggest novel therapeutic approaches. PMID:28248965

Targeting SHP-1, 2 and SHIP Pathways: A Novel Strategy for Cancer Treatment?

PubMed

Dempke, Wolfram C M; Uciechowski, Peter; Fenchel, Klaus; Chevassut, Timothy

2018-06-20

Well-balanced levels of tyrosine phosphorylation, maintained by the reversible and coordinated actions of protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs), are critical for a wide range of cellular processes including growth, differentiation, metabolism, migration, and survival. Aberrant tyrosine phosphorylation, as a result of a perturbed balance between the activities of PTKs and PTPs, is linked to the pathogenesis of numerous human diseases, including cancer, suggesting that PTPs may be innovative molecular targets for cancer treatment. Two PTPs that have an important inhibitory role in haematopoietic cells are SHP-1 and SHP-2. SHP-1, 2 promote cell growth and act by both upregulating positive signaling pathways and by downregulating negative signaling pathways. SHIP is another inhibitory phosphatase that is specific for the inositol phospholipid phosphatidylinositol-3,4,5-trisphosphate (PIP3). SHIP acts as a negative regulator of immune response by hydrolysing PIP3, and SHIP deficiency results in myeloproliferation and B-cell lymphoma in mice. The validation of SHP-1, 2 and SHIP as oncology targets has generated interest in the development of inhibitors as potential therapeutic agents for cancers; however, SHP-1, 2 and SHIP have proven to be an extremely difficult target for drug discovery, primarily due to the highly conserved and positively charged nature of their PTP active site, and many PTP inhibitors lack either appro-priate selectivity or membrane permeability. To overcome these caveats, novel techniques have been employed to synthesise new inhibitors that specifically attenuate the PTP-dependent signaling inside the cell and amongst them; some are already in clinical development which are discussed in this review. © 2018 S. Karger AG, Basel.

Human T-cell leukemia virus type 1 Tax interacts with Chk1 and attenuates DNA-damage induced G2 arrest mediated by Chk1.

PubMed

Park, Hyeon Ung; Jeong, Jae-Hoon; Chung, Jay H; Brady, John N

2004-06-24

Checkpoint kinase 1 (Chk1) mediates diverse cellular responses to genotoxic stress, regulating the network of genome-surveillance pathways that coordinate cell cycle progression with DNA repair. Chk1 is essential for mammalian development and viability, and has been shown to be important for both S and G(2) checkpoints. We now present evidence that the HTLV-1 Tax protein interacts directly with Chk1 and impairs its kinase activities in vitro and in vivo. The direct and physical interaction of Chk1 and Tax was observed in HTLV-1-infected T cells (C81, HuT 102 and MT-2) and transfected fibroblasts (293 T) by coimmunoprecipitation and by in vitro GST pull-down assays. Interestingly, Tax inhibited the kinase activity of Chk1 protein in in vitro and in vivo kinase assays. Consistent with these results, Tax inhibited the phosphorylation-dependent degradation of Cdc25A and G(2) arrest in response to gamma-irradiation (IR) in a dose-dependent manner in vivo. The G(2) arrest did not require Chk2 or p53. These studies provide the first example of a viral transforming protein targeting Chk1 and provide important insights into checkpoint pathway regulation.

Ca2+/Calmodulin-dependent kinase II signaling causes skeletal overgrowth and premature chondrocyte maturation.

PubMed

Taschner, Michael J; Rafigh, Mehran; Lampert, Fabienne; Schnaiter, Simon; Hartmann, Christine

2008-05-01

The long bones of vertebrate limbs originate from cartilage templates and are formed by the process of endochondral ossification. This process requires that chondrocytes undergo a progressive maturation from proliferating to postmitotic prehypertrophic to mature, hypertrophic chondrocytes. Coordinated control of proliferation and maturation regulates growth of the skeletal elements. Various signals and pathways have been implicated in orchestrating these processes, but the underlying intracellular molecular mechanisms are often not entirely known. Here we demonstrated in the chick using replication-competent retroviruses that constitutive activation of Calcium/Calmodulin-dependent kinase II (CaMKII) in the developing wing resulted in elongation of skeletal elements associated with premature differentiation of chondrocytes. The premature maturation of chondrocytes was a cell-autonomous effect of constitutive CaMKII signaling associated with down-regulation of cell-cycle regulators and up-regulation of chondrocyte maturation markers. In contrast, the elongation of the skeletal elements resulted from a non-cell autonomous up-regulation of the Indian hedgehog responsive gene encoding Parathyroid-hormone-related peptide. Reduction of endogenous CaMKII activity by overexpressing an inhibitory peptide resulted in shortening of the skeletal elements associated with a delay in chondrocyte maturation. Thus, CaMKII is an essential component of intracellular signaling pathways regulating chondrocyte maturation.

Axon Termination, Pruning, and Synaptogenesis in the Giant Fiber System of Drosophila melanogaster Is Promoted by Highwire.

PubMed

Borgen, Melissa; Rowland, Kimberly; Boerner, Jana; Lloyd, Brandon; Khan, Aruna; Murphey, Rodney

2017-03-01

The ubiquitin ligase Highwire has a conserved role in synapse formation. Here, we show that Highwire coordinates several facets of central synapse formation in the Drosophila melanogaster giant fiber system, including axon termination, axon pruning, and synaptic function. Despite the similarities to the fly neuromuscular junction, the role of Highwire and the underlying signaling pathways are distinct in the fly's giant fiber system. During development, branching of the giant fiber presynaptic terminal occurs and, normally, the transient branches are pruned away. However, in highwire mutants these ectopic branches persist, indicating that Highwire promotes axon pruning. highwire mutants also exhibit defects in synaptic function. Highwire promotes axon pruning and synaptic function cell-autonomously by attenuating a mitogen-activated protein kinase pathway including Wallenda, c-Jun N-terminal kinase/Basket, and the transcription factor Jun. We also show a novel role for Highwire in non-cell autonomous promotion of synaptic function from the midline glia. Highwire also regulates axon termination in the giant fibers, as highwire mutant axons exhibit severe overgrowth beyond the pruning defect. This excessive axon growth is increased by manipulating Fos expression in the cells surrounding the giant fiber terminal, suggesting that Fos regulates a trans -synaptic signal that promotes giant fiber axon growth. Copyright © 2017 by the Genetics Society of America.

p53 coordinates decidual sestrin 2/AMPK/mTORC1 signaling to govern parturition timing

PubMed Central

Cha, Jeeyeon; Yuan, Jia; Haraguchi, Hirofumi; Bartos, Amanda; Bradshaw, Heather B.; Hirota, Yasushi; Dey, Sudhansu K.

2016-01-01

Inflammation and oxidative stress are known risk factors for preterm birth (PTB); however, the mechanisms and pathways that influence this condition are not fully described. Previously, we showed that mTORC1 signaling is increased in mice harboring a uterine-specific deletion of transformation-related protein 53 (p53d/d mice), which exhibit premature decidual senescence that triggers spontaneous and inflammation-induced PTB. Treatment with the mTORC1 inhibitor rapamycin reduced the incidence of PTB in the p53d/d mice. Decidual senescence with heightened mTORC1 signaling is also a signature of human PTB. Here, we have identified an underlying mechanism for PTB and a potential therapeutic strategy for treating the condition. Treatment of pregnant p53d/d mice with either the antidiabetic drug metformin or the antioxidant resveratrol activated AMPK signaling and inhibited mTORC1 signaling in decidual cells. Both metformin and resveratrol protected against spontaneous and inflammation-induced PTB in p53d/d females. Using multiple approaches, we determined that p53 interacts with sestrins to coordinate an inverse relationship between AMPK and mTORC1 signaling that determines parturition timing. This signature was also observed in human decidual cells. Together, these results reveal that p53-dependent coordination of AMPK and mTORC1 signaling controls parturition timing and suggest that metformin and resveratrol have therapeutic potential to prevent PTB. PMID:27454290

GPCR-mediated PLCβγ/PKCβ/PKD signaling pathway regulates the cofilin phosphatase slingshot 2 in neutrophil chemotaxis

PubMed Central

Xu, Xuehua; Gera, Nidhi; Li, Hongyan; Yun, Michelle; Zhang, Liyong; Wang, Youhong; Wang, Q. Jane; Jin, Tian

2015-01-01

Chemotaxis requires precisely coordinated polymerization and depolymerization of the actin cytoskeleton at leading fronts of migrating cells. However, GPCR activation-controlled F-actin depolymerization remains largely elusive. Here, we reveal a novel signaling pathway, including Gαi, PLC, PKCβ, protein kinase D (PKD), and SSH2, in control of cofilin phosphorylation and actin cytoskeletal reorganization, which is essential for neutrophil chemotaxis. We show that PKD is essential for neutrophil chemotaxis and that GPCR-mediated PKD activation depends on PLC/PKC signaling. More importantly, we discover that GPCR activation recruits/activates PLCγ2 in a PI3K-dependent manner. We further verify that PKCβ specifically interacts with PKD1 and is required for chemotaxis. Finally, we identify slingshot 2 (SSH2), a phosphatase of cofilin (actin depolymerization factor), as a target of PKD1 that regulates cofilin phosphorylation and remodeling of the actin cytoskeleton during neutrophil chemotaxis. PMID:25568344

Tissue architecture and breast cancer: the role of extracellular matrix and steroid hormones

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

Hansen, R K; Bissell, M J

The changes in tissue architecture that accompany the development of breast cancer have been the focus of investigations aimed at developing new cancer therapeutics. As we learn more about the normal mammary gland, we have begun to understand the complex signaling pathways underlying the dramatic shifts in the structure and function of breast tissue. Integrin-, growth factor-, and steroid hormone-signaling pathways all play an important part in maintaining tissue architecture; disruption of the delicate balance of signaling results in dramatic changes in the way cells interact with each other and with the extracellular matrix, leading to breast cancer. The extracellularmore » matrix itself plays a central role in coordinating these signaling processes. In this review, we consider the interrelationships between the extracellular matrix, integrins, growth factors, and steroid hormones in mammary gland development and function.« less

The formin DIAPH1 (mDia1) regulates megakaryocyte proplatelet formation by remodeling the actin and microtubule cytoskeletons.

PubMed

Pan, Jiajia; Lordier, Larissa; Meyran, Deborah; Rameau, Philippe; Lecluse, Yann; Kitchen-Goosen, Susan; Badirou, Idinath; Mokrani, Hayat; Narumiya, Shuh; Alberts, Arthur S; Vainchenker, William; Chang, Yunhua

2014-12-18

Megakaryocytes are highly specialized precursor cells that produce platelets via cytoplasmic extensions called proplatelets. Proplatelet formation (PPF) requires profound changes in microtubule and actin organization. In this work, we demonstrated that DIAPH1 (mDia1), a mammalian homolog of Drosophila diaphanous that works as an effector of the small GTPase Rho, negatively regulates PPF by controlling the dynamics of the actin and microtubule cytoskeletons. Moreover, we showed that inhibition of both DIAPH1 and the Rho-associated protein kinase (Rock)/myosin pathway increased PPF via coordination of both cytoskeletons. We provide evidence that 2 major effectors of the Rho GTPase pathway (DIAPH1 and Rock/myosin II) are involved not only in Rho-mediated stress fibers assembly, but also in the regulation of microtubule stability and dynamics during PPF. © 2014 by The American Society of Hematology.

Plasticity in reflex pathways to the lower urinary tract following spinal cord injury

PubMed Central

de Groat, William C.; Yoshimura, Naoki

2013-01-01

The lower urinary tract has two main functions, storage and periodic expulsion of urine, that are regulated by a complex neural control system in the brain and lumbosacral spinal cord. This neural system coordinates the activity of two functional units in the lower urinary tract: (1) a reservoir (the urinary bladder) and (2) an outlet (consisting of bladder neck, urethra and striated muscles of the external urethra sphincter). During urine storage the outlet is closed and the bladder is quiescent to maintain a low intravesical pressure. During micturition the outlet relaxes and the bladder contracts to promote efficient release of urine. This reciprocal relationship between bladder and outlet is generated by reflex circuits some of which are under voluntary control. Experimental studies in animals indicate that the micturition reflex is mediated by a spinobulbospinal pathway passing through a coordination center (the pontine micturition center) located in the rostral brainstem. This reflex pathway is in turn modulated by higher centers in the cerebral cortex that are involved in the voluntary control of micturition. Spinal cord injury at cervical or thoracic levels disrupts voluntary control of voiding as well as the normal reflex pathways that coordinate bladder and sphincter function. Following spinal cord injury the bladder is initially areflexic but then becomes hyperreflexic due to the emergence of a spinal micturition reflex pathway. However the bladder does not empty efficiently because coordination between the bladder and urethral outlet is lost. Studies in animals indicate that dysfunction of the lower urinary tract after spinal cord injury is dependent in part on plasticity of bladder afferent pathways as well as reorganization of synaptic connections in the spinal cord. Reflex plasticity is associated with changes in the properties of ion channels and electrical excitability of afferent neurons and appears to be mediated in part by neurotrophic factors released in the spinal cord and/or the peripheral target organs. PMID:21596038

PHABULOSA Controls the Quiescent Center-Independent Root Meristem Activities in Arabidopsis thaliana

PubMed Central

Sebastian, Jose; Ryu, Kook Hui; Zhou, Jing; Tarkowská, Danuše; Tarkowski, Petr; Cho, Young-Hee; Yoo, Sang-Dong; Kim, Eun-Sol; Lee, Ji-Young

2015-01-01

Plant growth depends on stem cell niches in meristems. In the root apical meristem, the quiescent center (QC) cells form a niche together with the surrounding stem cells. Stem cells produce daughter cells that are displaced into a transit-amplifying (TA) domain of the root meristem. TA cells divide several times to provide cells for growth. SHORTROOT (SHR) and SCARECROW (SCR) are key regulators of the stem cell niche. Cytokinin controls TA cell activities in a dose-dependent manner. Although the regulatory programs in each compartment of the root meristem have been identified, it is still unclear how they coordinate one another. Here, we investigate how PHABULOSA (PHB), under the posttranscriptional control of SHR and SCR, regulates TA cell activities. The root meristem and growth defects in shr or scr mutants were significantly recovered in the shr phb or scr phb double mutant, respectively. This rescue in root growth occurs in the absence of a QC. Conversely, when the modified PHB, which is highly resistant to microRNA, was expressed throughout the stele of the wild-type root meristem, root growth became very similar to that observed in the shr; however, the identity of the QC was unaffected. Interestingly, a moderate increase in PHB resulted in a root meristem phenotype similar to that observed following the application of high levels of cytokinin. Our protoplast assay and transgenic approach using ARR10 suggest that the depletion of TA cells by high PHB in the stele occurs via the repression of B-ARR activities. This regulatory mechanism seems to help to maintain the cytokinin homeostasis in the meristem. Taken together, our study suggests that PHB can dynamically regulate TA cell activities in a QC-independent manner, and that the SHR-PHB pathway enables a robust root growth system by coordinating the stem cell niche and TA domain. PMID:25730098

Model-Based Analysis of Cell Cycle Responses to Dynamically Changing Environments

PubMed Central

Seaton, Daniel D; Krishnan, J

2016-01-01

Cell cycle progression is carefully coordinated with a cell’s intra- and extracellular environment. While some pathways have been identified that communicate information from the environment to the cell cycle, a systematic understanding of how this information is dynamically processed is lacking. We address this by performing dynamic sensitivity analysis of three mathematical models of the cell cycle in Saccharomyces cerevisiae. We demonstrate that these models make broadly consistent qualitative predictions about cell cycle progression under dynamically changing conditions. For example, it is shown that the models predict anticorrelated changes in cell size and cell cycle duration under different environments independently of the growth rate. This prediction is validated by comparison to available literature data. Other consistent patterns emerge, such as widespread nonmonotonic changes in cell size down generations in response to parameter changes. We extend our analysis by investigating glucose signalling to the cell cycle, showing that known regulation of Cln3 translation and Cln1,2 transcription by glucose is sufficient to explain the experimentally observed changes in cell cycle dynamics at different glucose concentrations. Together, these results provide a framework for understanding the complex responses the cell cycle is capable of producing in response to dynamic environments. PMID:26741131

Resistance to Hemi-Biotrophic F. graminearum Infection Is Associated with Coordinated and Ordered Expression of Diverse Defense Signaling Pathways

PubMed Central

Yi, Hongying; Yang, Liming; Kong, Zhongxin; Zhang, Lixia; Xue, Shulin; Jia, Haiyan; Ma, Zhengqiang

2011-01-01

Fusarium species cause serious diseases in cereal staple food crops such as wheat and maize. Currently, the mechanisms underlying resistance to Fusarium-caused diseases are still largely unknown. In the present study, we employed a combined proteomic and transcriptomic approach to investigate wheat genes responding to F. graminearum infection that causes Fusarium head blight (FHB). We found a total of 163 genes and 37 proteins that were induced by infection. These genes and proteins were associated with signaling pathways mediated by salicylic acid (SA), jasmonic acid (JA), ethylene (ET), calcium ions, phosphatidic acid (PA), as well as with reactive oxygen species (ROS) production and scavenging, antimicrobial compound synthesis, detoxification, and cell wall fortification. We compared the time-course expression profiles between FHB-resistant Wangshuibai plants and susceptible Meh0106 mutant plants of a selected set of genes that are critical to the plants' resistance and defense reactions. A biphasic phenomenon was observed during the first 24 h after inoculation (hai) in the resistant plants. The SA and Ca2+ signaling pathways were activated within 6 hai followed by the JA mediated defense signaling activated around 12 hai. ET signaling was activated between these two phases. Genes for PA and ROS synthesis were induced during the SA and JA phases, respectively. The delayed activation of the SA defense pathway in the mutant was associated with its susceptibility. After F. graminearum infection, the endogenous contents of SA and JA in Wangshuibai and the mutant changed in a manner similar to the investigated genes corresponding to the individual pathways. A few genes for resistance-related cell modification and phytoalexin production were also identified. This study provided important clues for designing strategies to curb diseases caused by Fusarium. PMID:21533105

Examining the intersection between splicing, nuclear export and small RNA pathways.

PubMed

Nabih, Amena; Sobotka, Julia A; Wu, Monica Z; Wedeles, Christopher J; Claycomb, Julie M

2017-11-01

Nuclear Argonaute/small RNA pathways in a variety of eukaryotic species are generally known to regulate gene expression via chromatin modulation and transcription attenuation in a process known as transcriptional gene silencing (TGS). However, recent data, including genetic screens, phylogenetic profiling, and molecular mechanistic studies, also point to a novel and emerging intersection between the splicing and nuclear export machinery with nuclear Argonaute/small RNA pathways in many organisms. In this review, we summarize the field's current understanding regarding the relationship between splicing, export and small RNA pathways, and consider the biological implications for coordinated regulation of transcripts by these pathways. We also address the importance and available approaches for understanding the RNA regulatory logic generated by the intersection of these particular pathways in the context of synthetic biology. The interactions between various eukaryotic RNA regulatory pathways, particularly splicing, nuclear export and small RNA pathways provide a type of combinatorial code that informs the identity ("self" versus "non-self") and dictates the fate of each transcript in a cell. Although the molecular mechanisms for how splicing and nuclear export impact small RNA pathways are not entirely clear at this early stage, the links between these pathways are widespread across eukaryotic phyla. The link between splicing, nuclear export, and small RNA pathways is emerging and establishes a new frontier for understanding the combinatorial logic of gene regulation across species that could someday be harnessed for therapeutic, biotechnology and agricultural applications. This article is part of a Special Issue entitled "Biochemistry of Synthetic Biology - Recent Developments" Guest Editor: Dr. Ilka Heinemann and Dr. Patrick O'Donoghue. Copyright © 2017 Elsevier B.V. All rights reserved.

Strategies of biochemical adaptation for hibernation in a South American marsupial Dromiciops gliroides: 1. Mitogen-activated protein kinases and the cell stress response.

PubMed

Wijenayake, Sanoji; Luu, Bryan E; Zhang, Jing; Tessier, Shannon N; Quintero-Galvis, Julian F; Gaitán-Espitia, Juan Diego; Nespolo, Roberto F; Storey, Kenneth B

2017-12-14

Hibernation is a period of torpor and heterothermy that is typically associated with a strong reduction in metabolic rate, global suppression of transcription and translation, and upregulation of various genes/proteins that are central to the cellular stress response such as protein kinases, antioxidants, and heat shock proteins. The current study examined cell signaling cascades in hibernating monito del monte, Dromiciops gliroides, a South American marsupial of the Order Microbiotheria. Responses to hibernation by members of the mitogen-activated protein kinase (MAPK) pathways, and their roles in coordinating hibernator metabolism were examined in liver, kidney, heart and brain of control and versus hibernating (4days continuous torpor) D. gliroides. The targets evaluated included key protein kinases in their activated phosphorylated forms (p-ERK/MAPK 1/2, p-MEK1, p-MSK1, p-p38, p-JNK) and related target proteins (p-CREB 2, p-ATF2, p-c-Jun and p-p53). Liver exhibited a strong coordinated response by MAPK members to hibernation with significant increases in protein phosphorylation levels of p-MEK1, p-ERK/MAPK1/2, p-MSK1, p-JNK and target proteins c-Jun, and p-ATF2, all combining to signify a strong activation of MAPK signaling during hibernation. Kidney also showed activation of MAPK cascades with significant increases in p-MEK1, p-ERK/MAPK1/2, p-p38, and p-c-Jun levels in hibernating animals. By contrast, responses by heart and brain indicated reduced MAPK pathway function during torpor with reduced phosphorylation of targets including p-ERK/MAPK 1/2 in both tissues as well as lower p-p38 and p-JNK content in heart. Overall, the data indicate a vital role for MAPK signaling in regulating the cell stress response during marsupial hibernation. Copyright © 2017 Elsevier Inc. All rights reserved.

Glycogen synthase kinase-3 inhibition sensitizes human induced pluripotent stem cells to thiol-containing antioxidants induced apoptosis.

PubMed

Tu, Chengyi; Xu, Robert; Koleti, Meghana; Zoldan, Janet

2017-08-01

Inhibition of glycogen synthase kinase 3 (GSK3) is an extensively used strategy to activate Wnt pathway for pluripotent stem cell (PSC) differentiation. However, the effects of such inhibition on PSCs, besides upregulating the Wnt pathway, have rarely been investigated despite that GSK3 is broadly involved in other cellular activities such as insulin signaling and cell growth/survival regulation. Here we describe a previously unknown synergistic effect between GSK3 inhibition (e.g., Chir99021 and LY2090314) and various normally non-toxic thiol-containing antioxidants (e.g., N-acetylcysteine, NAC) on the induction of apoptosis in human induced pluripotent stem cells (iPSCs). Neither Chir99021 nor the antioxidants individually induced significant apoptosis, whereas their combined treatment resulted in rapid and extensive apoptosis, with substantial caspase 3 activity observed within 3h and over 90% decrease in cell viability after 24h. We confirmed the generality of this phenomenon with multiple independent iPSCs lines, various thiol-based antioxidants and distinct GSK3 inhibitors. Mechanistically, we demonstrated that rapamycin treatment could substantially reduce cell death, suggesting the critical role of mammalian target of rapamycin (mTOR). Akt dysregulation was also found to partially contribute to cell apoptosis but was not the primary cause. Further, this coordinated proapoptotic effect was not detected in mouse ESCs but was present in another human cells line: a breast cancer cell line (MDA-MB-231). Given the wide use of GSK3 inhibition in biomedical research: from iPSC differentiation to cancer intervention and the treatment of neuronal diseases, researchers can potentially take advantage of or avoid this synergistic effect for improved experimental or clinical outcome. Copyright © 2017. Published by Elsevier B.V.

Stereochemistry of complexes with double and triple metal-ligand bonds: a continuous shape measures analysis.

PubMed

Alvarez, Santiago; Menjón, Babil; Falceto, Andrés; Casanova, David; Alemany, Pere

2014-11-17

To each coordination polyhedron we can associate a normalized coordination polyhedron that retains the angular orientation of the central atom-ligand bonds but has all the vertices at the same distance from the center. The use of shape measures of these normalized coordination polyhedra provides a simple and efficient way of discriminating angular and bond distance distortions from an ideal polyhedron. In this paper we explore the applications of such an approach to analyses of several stereochemical problems. Among others, we discuss how to discern the off-center displacement of the metal from metal-ligand bond shortening distortions in families of square planar biscarbene and octahedral dioxo complexes. The normalized polyhedron approach is also shown to be very useful to understand stereochemical trends with the help of shape maps, minimal distortion pathways, and ligand association/dissociation pathways, illustrated by the Berry and anti Berry distortions of triple-bonded [X≡ML4] complexes, the square pyramidal geometries of Mo coordination polyhedra in oxido-reductases, the coordination geometries of actinyl complexes, and the tetrahedricity of heavy atom-substituted carbon centers.

Methodological Caveats in the Detection of Coordinated Replay between Place Cells and Grid Cells.

PubMed

Trimper, John B; Trettel, Sean G; Hwaun, Ernie; Colgin, Laura Lee

2017-01-01

At rest, hippocampal "place cells," neurons with receptive fields corresponding to specific spatial locations, reactivate in a manner that reflects recently traveled trajectories. These "replay" events have been proposed as a mechanism underlying memory consolidation, or the transfer of a memory representation from the hippocampus to neocortical regions associated with the original sensory experience. Accordingly, it has been hypothesized that hippocampal replay of a particular experience should be accompanied by simultaneous reactivation of corresponding representations in the neocortex and in the entorhinal cortex, the primary interface between the hippocampus and the neocortex. Recent studies have reported that coordinated replay may occur between hippocampal place cells and medial entorhinal cortex grid cells, cells with multiple spatial receptive fields. Assessing replay in grid cells is problematic, however, as the cells exhibit regularly spaced spatial receptive fields in all environments and, therefore, coordinated replay between place cells and grid cells may be detected by chance. In the present report, we adapted analytical approaches utilized in recent studies of grid cell and place cell replay to determine the extent to which coordinated replay is spuriously detected between grid cells and place cells recorded from separate rats. For a subset of the employed analytical methods, coordinated replay was detected spuriously in a significant proportion of cases in which place cell replay events were randomly matched with grid cell firing epochs of equal duration. More rigorous replay evaluation procedures and minimum spike count requirements greatly reduced the amount of spurious findings. These results provide insights into aspects of place cell and grid cell activity during rest that contribute to false detection of coordinated replay. The results further emphasize the need for careful controls and rigorous methods when testing the hypothesis that place cells and grid cells exhibit coordinated replay.

Metabolic Control of Virulence Genes in Brucella abortus: HutC Coordinates virB Expression and the Histidine Utilization Pathway by Direct Binding to Both Promoters ▿ †

PubMed Central

Sieira, Rodrigo; Arocena, Gastón M.; Bukata, Lucas; Comerci, Diego J.; Ugalde, Rodolfo A.

2010-01-01

Type IV secretion systems (T4SS) are multicomponent machineries involved in the translocation of effector molecules across the bacterial cell envelope. The virB operon of Brucella abortus codes for a T4SS that is essential for virulence and intracellular multiplication of the bacterium in the host. Previous studies showed that the virB operon of B. abortus is tightly regulated within the host cells. In order to identify factors implicated in the control of virB expression, we searched for proteins of Brucella that directly bind to the virB promoter (PvirB). Using different procedures, we isolated a 27-kDa protein that binds specifically to PvirB. This protein was identified as HutC, the transcriptional repressor of the histidine utilization (hut) genes. Analyses of virB and hut promoter activity revealed that HutC exerts two different roles: it acts as a coactivator of transcription of the virB operon, whereas it represses the hut genes. Such activities were observed both intracellularly and in bacteria incubated under conditions that resemble the intracellular environment. Electrophoresis mobility shift assays (EMSA) and DNase I footprinting experiments revealed the structure, affinity, and localization of the HutC-binding sites and supported the regulatory role of HutC in both hut and virB promoters. Taken together, these results indicate that Brucella coopted the function of HutC to coordinate the Hut pathway with transcriptional regulation of the virB genes, probably as a way to sense its own metabolic state and develop adaptive responses to overcome intracellular host defenses. PMID:19854911

WNK1 Promotes PIP2 Synthesis to Coordinate Growth Factor and GPCR-Gq Signaling

PubMed Central

An, Sung-Wan; Cha, Seung-Kuy; Yoon, Joonho; Chang, Seungwoo; Ross, Elliott M.; Huang, Chou-Long

2011-01-01

Summary Background PLC-β signaling is generally thought to be mediated by allosteric activation by G proteins and Ca2+. While availability of the PIP2 substrate is limiting in some cases, its production has not been shown to be independently regulated as a signaling mechanism. WNK1 protein kinase is known to regulate ion homeostasis and cause hypertension when expression is increased by gene mutations. However, its signaling functions remain largely elusive. Results Using diacylglycerol-stimulated TRPC6 and inositol trisphosphate-mediated Ca2+ transients as cellular biosensors, we show that WNK1 stimulates PLC-β signaling in cells by promoting the synthesis of PIP2 via stimulation of phosphatidylinositol 4-kinase IIIα. WNK1 kinase activity is not required. Stimulation of PLC-β by WNK1 and by Gαq are synergistic; WNK1 activity is essential for regulation of PLC-β signaling by Gq-coupled receptors and basal input from Gq is necessary for WNK1 signaling via PLC-β. WNK1 further amplifies PLC-β signaling when it is phosphorylated by Akt kinase in response to insulin-like growth factor. Conclusions WNK1 is a novel regulator of PLC-β that acts by controlling substrate availability. WNK1 thereby coordinates signaling between G protein and Akt kinase pathways. Because PIP2 is itself a signaling molecule, regulation of PIP2 synthesis by WNK1 also allows the cell to initiate PLC signaling while independently controlling the effects of PIP2 on other targets. These findings describe a new signaling pathway for Akt-activating growth factors, a mechanism for G protein-growth factor crosstalk and a means to independently control PLC signaling and PIP2 availability. PMID:22119528

Ablation of PGC1 beta prevents mTOR dependent endoplasmic reticulum stress response

PubMed Central

Camacho, Alberto; Rodriguez-Cuenca, Sergio; Blount, Margaret; Prieur, Xavier; Barbarroja, Nuria; Fuller, Maria; Hardingham, Giles E.; Vidal-Puig, Antonio

2012-01-01

Mitochondria dysfunction contributes to the pathophysiology of obesity, diabetes, neurodegeneration and ageing. The peroxisome proliferator-activated receptor-gamma coactivator-1β (PGC-1β) coordinates mitochondrial biogenesis and function as well as fatty acid metabolism. It has been suggested that endoplasmic reticulum (ER) stress may be one of the mechanisms linking mitochondrial dysfunction and these pathologies. Here we investigate whether PGC-1β ablation affects the ER stress response induced by specific nutritional and pharmacological challenges in the CNS. By using flow cytometry, western blot, real time PCR and several pharmacological and nutritional interventions in PGC-1β knock out and WT mice, we confirmed that PGC-1β coordinates mitochondria function in brain and reported for the first time that a) ablation of PGC-1β is associated with constitutive activation of mTORC1 pathway associated with increased basal GRP78 protein levels in hypothalamus and cortex of animals fed chow diet; and b) in animals fed chronically with high fat diet (HFD) or high protein diet (HPD), we observed a failure to appropriately induce ER stress response in the absence of PGC-1β, associated with an increase in mTOR pathway phosphorylation. This contrasted with the appropriate upregulation of ER stress response observed in wild type littermates. Additionally, inefficient in vitro induction of ER stress by thapsigargin seems result in apoptotic neuronal cell death in PGC-1β KO. Our data indicate that PGC-1β is required for a neuronal ER response to nutritional stress imposed by HFD and HPD diets and that genetic ablation of PGC-1β might increase the susceptibility to neuronal damage and cell death. PMID:22771762

Light-Induced Expression of a MYB Gene Regulates Anthocyanin Biosynthesis in Red Apples1

PubMed Central

Takos, Adam M.; Jaffé, Felix W.; Jacob, Steele R.; Bogs, Jochen; Robinson, Simon P.; Walker, Amanda R.

2006-01-01

Anthocyanins are secondary metabolites found in higher plants that contribute to the colors of flowers and fruits. In apples (Malus domestica Borkh.), several steps of the anthocyanin pathway are coordinately regulated, suggesting control by common transcription factors. A gene encoding an R2R3 MYB transcription factor was isolated from apple (cv Cripps' Pink) and designated MdMYB1. Analysis of the deduced amino acid sequence suggests that this gene encodes an ortholog of anthocyanin regulators in other plants. The expression of MdMYB1 in both Arabidopsis (Arabidopsis thaliana) plants and cultured grape cells induced the ectopic synthesis of anthocyanin. In the grape (Vitis vinifera) cells MdMYB1 stimulated transcription from the promoters of two apple genes encoding anthocyanin biosynthetic enzymes. In ripening apple fruit the transcription of MdMYB1 was correlated with anthocyanin synthesis in red skin sectors of fruit. When dark-grown fruit were exposed to sunlight, MdMYB1 transcript levels increased over several days, correlating with anthocyanin synthesis in the skin. MdMYB1 gene transcripts were more abundant in red skin apple cultivars compared to non-red skin cultivars. Several polymorphisms were identified in the promoter of MdMYB1. A derived cleaved amplified polymorphic sequence marker designed to one of these polymorphisms segregated with the inheritance of skin color in progeny from a cross of an unnamed red skin selection (a sibling of Cripps' Pink) and the non-red skin cultivar Golden Delicious. We conclude that MdMYB1 coordinately regulates genes in the anthocyanin pathway and the expression level of this regulator is the genetic basis for apple skin color. PMID:17012405

Convergent microRNA actions coordinate neocortical development.

PubMed

Barca-Mayo, Olga; De Pietri Tonelli, Davide

2014-08-01

Neocortical development is a complex process that, at the cellular level, involves tight control of self-renewal, cell fate commitment, survival, differentiation and delamination/migration. These processes require, at the molecular level, the precise regulation of intrinsic signaling pathways and extrinsic factors with coordinated action in a spatially and temporally specific manner. Transcriptional regulation plays an important role during corticogenesis; however, microRNAs (miRNAs) are emerging as important post-transcriptional regulators of various aspects of central nervous system development. miRNAs are a class of small, single-stranded noncoding RNA molecules that control the expression of the majority of protein coding genes (i.e., targets). How do different miRNAs achieve precise control of gene networks during neocortical development? Here, we critically review all the miRNA-target interactions validated in vivo, with relevance to the generation and migration of pyramidal-projection glutamatergic neurons, and for the initial formation of cortical layers in the embryonic development of rodent neocortex. In particular, we focus on convergent miRNA actions, which are still a poorly understood layer of complexity in miRNA signaling, but potentially one of the keys to disclosing how miRNAs achieve the precise coordination of complex biological processes such as neocortical development.

The cellular response to vascular endothelial growth factors requires co-ordinated signal transduction, trafficking and proteolysis

PubMed Central

Smith, Gina A.; Fearnley, Gareth W.; Tomlinson, Darren C.; Harrison, Michael A.; Ponnambalam, Sreenivasan

2015-01-01

VEGFs (vascular endothelial growth factors) are a family of conserved disulfide-linked soluble secretory glycoproteins found in higher eukaryotes. VEGFs mediate a wide range of responses in different tissues including metabolic homoeostasis, cell proliferation, migration and tubulogenesis. Such responses are initiated by VEGF binding to soluble and membrane-bound VEGFRs (VEGF receptor tyrosine kinases) and co-receptors. VEGF and receptor splice isoform diversity further enhances complexity of membrane protein assembly and function in signal transduction pathways that control multiple cellular responses. Different signal transduction pathways are simultaneously activated by VEGFR–VEGF complexes with membrane trafficking along the endosome–lysosome network further modulating signal output from multiple enzymatic events associated with such pathways. Balancing VEGFR–VEGF signal transduction with trafficking and proteolysis is essential in controlling the intensity and duration of different intracellular signalling events. Dysfunction in VEGF-regulated signal transduction is important in chronic disease states including cancer, atherosclerosis and blindness. This family of growth factors and receptors is an important model system for understanding human disease pathology and developing new therapeutics for treating such ailments. PMID:26285805

The PINK1-PARKIN Mitochondrial Ubiquitylation Pathway Drives a Program of OPTN/NDP52 Recruitment and TBK1 Activation to Promote Mitophagy.

PubMed

Heo, Jin-Mi; Ordureau, Alban; Paulo, Joao A; Rinehart, Jesse; Harper, J Wade

2015-10-01

Damaged mitochondria are detrimental to cellular homeostasis. One mechanism for removal of damaged mitochondria involves the PINK1-PARKIN pathway, which poly-ubiquitylates damaged mitochondria to promote mitophagy. We report that assembly of ubiquitin chains on mitochondria triggers autophagy adaptor recruitment concomitantly with activation of the TBK1 kinase, which physically associates with OPTN, NDP52, and SQSTM1. TBK1 activation in HeLa cells requires OPTN and NDP52 and OPTN ubiquitin chain binding. In addition to the known role of S177 phosphorylation in OPTN on ATG8 recruitment, TBK1-dependent phosphorylation on S473 and S513 promotes ubiquitin chain binding in vitro as well as TBK1 activation, OPTN mitochondrial retention, and efficient mitophagy in vivo. These data reveal a self-reinforcing positive feedback mechanism that coordinates TBK1-dependent autophagy adaptor phosphorylation with the assembly of ubiquitin chains on mitochondria to facilitate efficient mitophagy, and mechanistically links genes mutated in Parkinson's disease and amyotrophic lateral sclerosis in a common selective autophagy pathway. Copyright © 2015 Elsevier Inc. All rights reserved.

Preface: cardiac control pathways: signaling and transport phenomena.

PubMed

Sideman, Samuel

2008-03-01

Signaling is part of a complex system of communication that governs basic cellular functions and coordinates cellular activity. Transfer of ions and signaling molecules and their interactions with appropriate receptors, transmembrane transport, and the consequent intracellular interactions and functional cellular response represent a complex system of interwoven phenomena of transport, signaling, conformational changes, chemical activation, and/or genetic expression. The well-being of the cell thus depends on a harmonic orchestration of all these events and the existence of control mechanisms that assure the normal behavior of the various parameters involved and their orderly expression. The ability of cells to sustain life by perceiving and responding correctly to their microenvironment is the basis for development, tissue repair, and immunity, as well as normal tissue homeostasis. Natural deviations, or human-induced interference in the signaling pathways and/or inter- and intracellular transport and information transfer, are responsible for the generation, modulation, and control of diseases. The present overview aims to highlight some major topics of the highly complex cellular information transfer processes and their control mechanisms. Our goal is to contribute to the understanding of the normal and pathophysiological phenomena associated with cardiac functions so that more efficient therapeutic modalities can be developed. Our objective in this volume is to identify and enhance the study of some basic passive and active physical and chemical transport phenomena, physiological signaling pathways, and their biological consequences.

Functional connectivity in the neuromuscular system underlying bimanual coordination

PubMed Central

de Vries, Ingmar E. J.; Daffertshofer, Andreas; Stegeman, Dick F.

2016-01-01

Neural synchrony has been suggested as a mechanism for integrating distributed sensorimotor systems involved in coordinated movement. To test the role of corticomuscular and intermuscular coherence in bimanual coordination, we experimentally manipulated the degree of coordination between hand muscles by varying the sensitivity of the visual feedback to differences in bilateral force. In 16 healthy participants, cortical activity was measured using EEG and muscle activity of the flexor pollicis brevis of both hands using high-density electromyography (HDsEMG). Using the uncontrolled manifold framework, coordination between bilateral forces was quantified by the synergy index RV in the time and frequency domain. Functional connectivity was assessed using corticomuscular coherence between muscle activity and cortical source activity and intermuscular coherence between bilateral EMG activity. The synergy index increased in the high coordination condition. RV was higher in the high coordination condition in frequencies between 0 and 0.5 Hz; for the 0.5- to 2-Hz frequency band, this pattern was inverted. Corticomuscular coherence in the beta band (16–30 Hz) was maximal in the contralateral motor cortex and was reduced in the high coordination condition. In contrast, intermuscular coherence was observed at 5–12 Hz and increased with bimanual coordination. Within-subject comparisons revealed a negative correlation between RV and corticomuscular coherence and a positive correlation between RV and intermuscular coherence. Our findings suggest two distinct neural pathways: 1) corticomuscular coherence reflects direct corticospinal projections involved in controlling individual muscles; and 2) intermuscular coherence reflects diverging pathways involved in the coordination of multiple muscles. PMID:27628205

The patient-breast cancer care pathway: how could it be optimized?

PubMed

Baffert, Sandrine; Hoang, Huong Ly; Brédart, Anne; Asselain, Bernard; Alran, Séverine; Berseneff, Hélène; Huchon, Cyrille; Trichot, Caroline; Combes, Aline; Alves, Karine; Koskas, Martin; Nguyen, Thuy; Roulot, Aurélie; Rouzier, Roman; Héquet, Delphine

2015-05-12

A care pathway is defined as patient-focused global care that addresses temporal (effective and coordinated management throughout the illness) and spatial issues (treatment is provided near the health territory in or around the patient's home). Heterogeneity of the care pathways in breast cancer (BC) is presumed but not well evaluated. The OPTISOINS01 study aims to assess every aspect of the care pathway for early BC patients using a temporal and spatial scope. An observational, prospective, multicenter study in a regional health territory (Ile-de-France, France) in different types of structures: university or local hospitals and comprehensive cancer centers. We will include and follow during 1 year 1,000 patients. The study consists of 3 work-packages: - Cost of pathway The aim of this WP is to calculate the overall costs of the early BC pathway at 1 year from different perspectives (society, health insurance and patient) using a cost-of-illness analysis. Using a bottom-up method, we will assess direct costs, including medical direct costs and nonmedical direct costs (transportation, home modifications, home care services, and social services), and indirect costs (loss of production). - Patient satisfaction and work reintegration Three questionnaires will assess the patients' satisfaction and possible return to work: the occupational questionnaire for employed women; the questionnaire on the need for supportive care, SCNS-SF34 ('breast cancer' module, SCNS-BR8); and the OUTPASSAT-35 questionnaire. - Quality, coordination and access to innovation Quality will be evaluated based on visits and treatment within a set period, whether the setting offers a multidisciplinary consultative framework, the management by nurse coordinators, the use of a personalized care plan, the provision of information via documents about treatments and the provision of supportive care. The coordination between structures and caregivers will be evaluated at several levels. Day surgery, home hospitalization and one-stop breast clinic visits will be recorded to assess the patient's access to innovation. The assessment of care pathways encourages the implementation of new payment models. Our approach could help health care professionals and policymakers to establish other cost-of-illness studies and plan the allocation of resources on a patient basis rather than a visit basis.

Automated placement of interfaces in conformational kinetics calculations using machine learning

NASA Astrophysics Data System (ADS)

Grazioli, Gianmarc; Butts, Carter T.; Andricioaei, Ioan

2017-10-01

Several recent implementations of algorithms for sampling reaction pathways employ a strategy for placing interfaces or milestones across the reaction coordinate manifold. Interfaces can be introduced such that the full feature space describing the dynamics of a macromolecule is divided into Voronoi (or other) cells, and the global kinetics of the molecular motions can be calculated from the set of fluxes through the interfaces between the cells. Although some methods of this type are exact for an arbitrary set of cells, in practice, the calculations will converge fastest when the interfaces are placed in regions where they can best capture transitions between configurations corresponding to local minima. The aim of this paper is to introduce a fully automated machine-learning algorithm for defining a set of cells for use in kinetic sampling methodologies based on subdividing the dynamical feature space; the algorithm requires no intuition about the system or input from the user and scales to high-dimensional systems.

Tissue oxygen demand in regulation of the behavior of the cells in the vasculature.

PubMed

Barvitenko, Nadezhda N; Aslam, Muhammad; Filosa, Jessica; Matteucci, Elena; Nikinmaa, Mikko; Pantaleo, Antonella; Saldanha, Carlota; Baskurt, Oguz K

2013-08-01

The control of arteriolar diameters in microvasculature has been in the focus of studies on mechanisms matching oxygen demand and supply at the tissue level. Functionally, important vascular elements include EC, VSMC, and RBC. Integration of these different cell types into functional units aimed at matching tissue oxygen supply with tissue oxygen demand is only achieved when all these cells can respond to the signals of tissue oxygen demand. Many vasoactive agents that serve as signals of tissue oxygen demand have their receptors on all these types of cells (VSMC, EC, and RBC) implying that there can be a coordinated regulation of their behavior by the tissue oxygen demand. Such functions of RBC as oxygen carrying by Hb, rheology, and release of vasoactive agents are considered. Several common extra- and intracellular signaling pathways that link tissue oxygen demand with control of VSMC contractility, EC permeability, and RBC functioning are discussed. © 2013 John Wiley & Sons Ltd.

The budding yeast RSC complex maintains ploidy by promoting spindle pole body insertion.

PubMed

Sing, Tina L; Hung, Minnie P; Ohnuki, Shinsuke; Suzuki, Godai; San Luis, Bryan-Joseph; McClain, Melainia; Unruh, Jay R; Yu, Zulin; Ou, Jiongwen; Marshall-Sheppard, Jesse; Huh, Won-Ki; Costanzo, Michael; Boone, Charles; Ohya, Yoshikazu; Jaspersen, Sue L; Brown, Grant W

2018-06-06

Ploidy is tightly regulated in eukaryotic cells and is critical for cell function and survival. Cells coordinate multiple pathways to ensure replicated DNA is segregated accurately to prevent abnormal changes in chromosome number. In this study, we characterize an unanticipated role for the Saccharomyces cerevisiae "remodels the structure of chromatin" (RSC) complex in ploidy maintenance. We show that deletion of any of six nonessential RSC genes causes a rapid transition from haploid to diploid DNA content because of nondisjunction events. Diploidization is accompanied by diagnostic changes in cell morphology and is stably maintained without further ploidy increases. We find that RSC promotes chromosome segregation by facilitating spindle pole body (SPB) duplication. More specifically, RSC plays a role in distributing two SPB insertion factors, Nbp1 and Ndc1, to the new SPB. Thus, we provide insight into a role for a SWI/SNF family complex in SPB duplication and ploidy maintenance. © 2018 Sing et al.

Automated placement of interfaces in conformational kinetics calculations using machine learning.

PubMed

Grazioli, Gianmarc; Butts, Carter T; Andricioaei, Ioan

2017-10-21

Several recent implementations of algorithms for sampling reaction pathways employ a strategy for placing interfaces or milestones across the reaction coordinate manifold. Interfaces can be introduced such that the full feature space describing the dynamics of a macromolecule is divided into Voronoi (or other) cells, and the global kinetics of the molecular motions can be calculated from the set of fluxes through the interfaces between the cells. Although some methods of this type are exact for an arbitrary set of cells, in practice, the calculations will converge fastest when the interfaces are placed in regions where they can best capture transitions between configurations corresponding to local minima. The aim of this paper is to introduce a fully automated machine-learning algorithm for defining a set of cells for use in kinetic sampling methodologies based on subdividing the dynamical feature space; the algorithm requires no intuition about the system or input from the user and scales to high-dimensional systems.

ROCK1 and 2 differentially regulate actomyosin organization to drive cell and synaptic polarity

PubMed Central

Badoual, Mathilde; Asmussen, Hannelore; Patel, Heather; Whitmore, Leanna; Horwitz, Alan Rick

2015-01-01

RhoGTPases organize the actin cytoskeleton to generate diverse polarities, from front–back polarity in migrating cells to dendritic spine morphology in neurons. For example, RhoA through its effector kinase, RhoA kinase (ROCK), activates myosin II to form actomyosin filament bundles and large adhesions that locally inhibit and thereby polarize Rac1-driven actin polymerization to the protrusions of migratory fibroblasts and the head of dendritic spines. We have found that the two ROCK isoforms, ROCK1 and ROCK2, differentially regulate distinct molecular pathways downstream of RhoA, and their coordinated activities drive polarity in both cell migration and synapse formation. In particular, ROCK1 forms the stable actomyosin filament bundles that initiate front–back and dendritic spine polarity. In contrast, ROCK2 regulates contractile force and Rac1 activity at the leading edge of migratory cells and the spine head of neurons; it also specifically regulates cofilin-mediated actin remodeling that underlies the maturation of adhesions and the postsynaptic density of dendritic spines. PMID:26169356

Cell identity regulators link development and stress responses in the Arabidopsis root.

PubMed

Iyer-Pascuzzi, Anjali S; Jackson, Terry; Cui, Hongchang; Petricka, Jalean J; Busch, Wolfgang; Tsukagoshi, Hironaka; Benfey, Philip N

2011-10-18

Stress responses in plants are tightly coordinated with developmental processes, but interaction of these pathways is poorly understood. We used genome-wide assays at high spatiotemporal resolution to understand the processes that link development and stress in the Arabidopsis root. Our meta-analysis finds little evidence for a universal stress response. However, common stress responses appear to exist with many showing cell type specificity. Common stress responses may be mediated by cell identity regulators because mutations in these genes resulted in altered responses to stress. Evidence for a direct role for cell identity regulators came from genome-wide binding profiling of the key regulator SCARECROW, which showed binding to regulatory regions of stress-responsive genes. Coexpression in response to stress was used to identify genes involved in specific developmental processes. These results reveal surprising linkages between stress and development at cellular resolution, and show the power of multiple genome-wide data sets to elucidate biological processes. Copyright © 2011 Elsevier Inc. All rights reserved.

Connexin 43 and ATP-sensitive potassium channels crosstalk: a missing link in hypoxia/ischemia stress.

PubMed

Ahmad Waza, Ajaz; Ahmad Bhat, Shabir; Ul Hussain, Mahboob; Ganai, Bashir A

2018-02-01

Connexin 43 (Cx43) is a gap junction protein expressed in various tissues and organs of vertebrates. Besides functioning as a gap junction, Cx43 also regulates diverse cellular processes like cell growth and differentiation, cell migration, cell survival, etc. Cx43 is critical for normal cardiac functioning and is therefore abundantly expressed in cardiomyocytes. On the other hand, ATP-sensitive potassium (K ATP ) channels are metabolic sensors converting metabolic changes into electrical activity. These channels are important in maintaining the neurotransmitter release, smooth muscle relaxation, cardiac action potential repolarization, normal physiology of cellular repolarization, insulin secretion and immune function. Cx43 and K ATP channels are part of the same signaling pathway, regulating cell survival during stress conditions and ischemia/hypoxia preconditioning. However, the underlying molecular mechanism for their combined role in ischemia/hypoxia preconditioning is largely unknown. The current review focuses on understanding the molecular mechanism responsible for the coordinated role of Cx43 and K ATP channel protein in protecting cardiomyocytes against ischemia/hypoxia stress.

Genetic epistasis between heparan sulfate and FGF-Ras signaling controls lens development

PubMed Central

Qu, Xiuxia; Hertzler, Kristina; Pan, Yi; Grobe, Kay; Robinson, Michael L.; Zhang, Xin

2011-01-01

Vertebrate lens development depends on a complex network of signaling molecules to coordinate cell proliferation, migration and differentiation. In this study, we have studied the role of heparan sulfate in lens specific signaling by generating a conditional ablation of heparan sulfate modification genes, Ndst1 and Ndst2. In this mutant, N-sulfation of heparan sulfate was disrupted after the lens induction stage, resulting in reduced lens cell proliferation, increased cell death and defective lens fiber differentiation in later lens development. The loss of Ndst function also prevented the assembly of Fgf/Fgfr complexes on the lens cell surface and disrupted ERK signaling within the lens. We further demonstrated that Ndst mutation completely inhibited the FGF1 and Fgf3 overexpression phenotypes, but Kras reactivation was sufficient to reverse the Ndst deficient lens differentiation defect. The epistatic relationship between Ndst and FGF-Ras signaling demonstrates that FGF signaling is the predominant signaling pathway controlled by Ndst in lens development. PMID:21536023

Coordinated transcriptional regulation of two key genes in the lignin branch pathway - CAD and CCR - is mediated through MYB- binding sites

PubMed Central

2010-01-01

Background Cinnamoyl CoA reductase (CCR) and cinnamyl alcohol dehydrogenase (CAD) catalyze the final steps in the biosynthesis of monolignols, the monomeric units of the phenolic lignin polymers which confer rigidity, imperviousness and resistance to biodegradation to cell walls. We have previously shown that the Eucalyptus gunnii CCR and CAD2 promoters direct similar expression patterns in vascular tissues suggesting that monolignol production is controlled, at least in part, by the coordinated transcriptional regulation of these two genes. Although consensus motifs for MYB transcription factors occur in most gene promoters of the whole phenylpropanoid pathway, functional evidence for their contribution to promoter activity has only been demonstrated for a few of them. Here, in the lignin-specific branch, we studied the functional role of MYB elements as well as other cis-elements identified in the regulatory regions of EgCAD2 and EgCCR promoters, in the transcriptional activity of these gene promoters. Results By using promoter deletion analysis and in vivo footprinting, we identified an 80 bp regulatory region in the Eucalyptus gunnii EgCAD2 promoter that contains two MYB elements, each arranged in a distinct module with newly identified cis-elements. A directed mutagenesis approach was used to introduce block mutations in all putative cis-elements of the EgCAD2 promoter and in those of the 50 bp regulatory region previously delineated in the EgCCR promoter. We showed that the conserved MYB elements in EgCAD2 and EgCCR promoters are crucial both for the formation of DNA-protein complexes in EMSA experiments and for the transcriptional activation of EgCAD2 and EgCCR promoters in vascular tissues in planta. In addition, a new regulatory cis-element that modulates the balance between two DNA-protein complexes in vitro was found to be important for EgCAD2 expression in the cambial zone. Conclusions Our assignment of functional roles to the identified cis-elements clearly demonstrates the importance of MYB cis-elements in the transcriptional regulation of two genes of the lignin-specific pathway and support the hypothesis that MYB elements serve as a common means for the coordinated regulation of genes in the entire lignin biosynthetic pathway. PMID:20584286

G protein-coupled receptor kinase 2 positively regulates epithelial cell migration

PubMed Central

Penela, Petronila; Ribas, Catalina; Aymerich, Ivette; Eijkelkamp, Niels; Barreiro, Olga; Heijnen, Cobi J; Kavelaars, Annemieke; Sánchez-Madrid, Francisco; Mayor, Federico

2008-01-01

Cell migration requires integration of signals arising from both the extracellular matrix and messengers acting through G protein-coupled receptors (GPCRs). We find that increased levels of G protein-coupled receptor kinase 2 (GRK2), a key player in GPCR regulation, potentiate migration of epithelial cells towards fibronectin, whereas such process is decreased in embryonic fibroblasts from hemizygous GRK2 mice or upon knockdown of GRK2 expression. Interestingly, the GRK2 effect on fibronectin-mediated cell migration involves the paracrine/autocrine activation of a sphingosine-1-phosphate (S1P) Gi-coupled GPCR. GRK2 positively modulates the activity of the Rac/PAK/MEK/ERK pathway in response to adhesion and S1P by a mechanism involving the phosphorylation-dependent, dynamic interaction of GRK2 with GIT1, a key scaffolding protein in cell migration processes. Furthermore, decreased GRK2 levels in hemizygous mice result in delayed wound healing rate in vivo, consistent with a physiological role of GRK2 as a regulator of coordinated integrin and GPCR-directed epithelial cell migration. PMID:18369319

Genome-wide screen identifies novel machineries required for both ciliogenesis and cell cycle arrest upon serum starvation

PubMed Central

Kim, Ji Hyun; Ki, Soo Mi; Joung, Je-Gun; Scott, Eric; Heynen-Genel, Susanne; Aza-Blanc, Pedro; Kwon, Chang Hyuk; Kim, Joon; Gleeson, Joseph G.; Lee, Ji Eun

2016-01-01

Biogenesis of the primary cilium, a cellular organelle mediating various signaling pathways, is generally coordinated with cell cycle exit/re-entry. Although the dynamic cell cycle-associated profile of the primary cilium has been largely accepted, the mechanism governing the link between ciliogenesis and cell cycle progression has been poorly understood. Using a human genome-wide RNAi screen, we identify genes encoding subunits of the spliceosome and proteasome as novel regulators of ciliogenesis. We demonstrate that 1) the mRNA processing-related hits are essential for RNA expression of molecules acting in cilia disassembly, such as AURKA and PLK1, and 2) the ubiquitin-proteasome systems (UPS)-involved hits are necessary for proteolysis of molecules acting in cilia assembly, such as IFT88 and CPAP. In particular, we show that these screen hit-associated mechanisms are crucial for both cilia assembly and cell cycle arrest in response to serum withdrawal. Finally, our data suggest that the mRNA processing mechanism may modulate the UPS-dependent decay of cilia assembly regulators to control ciliary resorption-coupled cell cycle re-entry. PMID:27033521

Common developmental genome deprogramming in schizophrenia - Role of Integrative Nuclear FGFR1 Signaling (INFS).

PubMed

Narla, S T; Lee, Y-W; Benson, C A; Sarder, P; Brennand, K J; Stachowiak, E K; Stachowiak, M K

2017-07-01

The watershed-hypothesis of schizophrenia asserts that over 200 different mutations dysregulate distinct pathways that converge on an unspecified common mechanism(s) that controls disease ontogeny. Consistent with this hypothesis, our RNA-sequencing of neuron committed cells (NCCs) differentiated from established iPSCs of 4 schizophrenia patients and 4 control subjects uncovered a dysregulated transcriptome of 1349 mRNAs common to all patients. Data reveals a global dysregulation of developmental genome, deconstruction of coordinated mRNA networks, and the formation of aberrant, new coordinated mRNA networks indicating a concerted action of the responsible factor(s). Sequencing of miRNA transcriptomes demonstrated an overexpression of 16 miRNAs and deconstruction of interactive miRNA-mRNA networks in schizophrenia NCCs. ChiPseq revealed that the nuclear (n) form of FGFR1, a pan-ontogenic regulator, is overexpressed in schizophrenia NCCs and overtargets dysregulated mRNA and miRNA genes. The nFGFR1 targeted 54% of all human gene promoters and 84.4% of schizophrenia dysregulated genes. The upregulated genes reside within major developmental pathways that control neurogenesis and neuron formation, whereas downregulated genes are involved in oligodendrogenesis. Our results indicate (i) an early (preneuronal) genomic etiology of schizophrenia, (ii) dysregulated genes and new coordinated gene networks are common to unrelated cases of schizophrenia, (iii) gene dysregulations are accompanied by increased nFGFR1-genome interactions, and (iv) modeling of increased nFGFR1 by an overexpression of a nFGFR1 lead to up or downregulation of selected genes as observed in schizophrenia NCCs. Together our results designate nFGFR1 signaling as a potential common dysregulated mechanism in investigated patients and potential therapeutic target in schizophrenia. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Cadmium, cobalt and lead cause stress response, cell cycle deregulation and increased steroid as well as xenobiotic metabolism in primary normal human bronchial epithelial cells which is coordinated by at least nine transcription factors.

PubMed

Glahn, Felix; Schmidt-Heck, Wolfgang; Zellmer, Sebastian; Guthke, Reinhard; Wiese, Jan; Golka, Klaus; Hergenröder, Roland; Degen, Gisela H; Lehmann, Thomas; Hermes, Matthias; Schormann, Wiebke; Brulport, Marc; Bauer, Alexander; Bedawy, Essam; Gebhardt, Rolf; Hengstler, Jan G; Foth, Heidi

2008-08-01

Workers occupationally exposed to cadmium, cobalt and lead have been reported to have increased levels of DNA damage. To analyze whether in vivo relevant concentrations of heavy metals cause systematic alterations in RNA expression patterns, we performed a gene array study using primary normal human bronchial epithelial cells. Cells were incubated with 15 microg/l Cd(II), 25 microg/l Co(II) and 550 microg/l Pb(II) either with individual substances or in combination. Differentially expressed genes were filtered out and used to identify enriched GO categories as well as KEGG pathways and to identify transcription factors whose binding sites are enriched in a given set of promoters. Interestingly, combined exposure to Cd(II), Co(II) and Pb(II) caused a coordinated response of at least seven stress response-related transcription factors, namely Oct-1, HIC1, TGIF, CREB, ATF4, SRF and YY1. A stress response was further corroborated by up regulation of genes involved in glutathione metabolism. A second major response to heavy metal exposure was deregulation of the cell cycle as evidenced by down regulation of the transcription factors ELK-1 and the Ets transcription factor GABP, as well as deregulation of genes involved in purine and pyrimidine metabolism. A third and surprising response was up regulation of genes involved in steroid metabolism, whereby promoter analysis identified up regulation of SRY that is known to play a role in sex determination. A forth response was up regulation of xenobiotic metabolising enzymes, particularly of dihydrodiol dehydrogenases 1 and 2 (AKR1C1, AKR1C2). Incubations with individual heavy metals showed that the response of AKR1C1 and AKR1C2 was predominantly caused by lead. In conclusion, we have shown that in vivo relevant concentrations of Cd(II), Co(II) and Pb(II) cause a complex and coordinated response in normal human bronchial epithelial cells. This study gives an overview of the most responsive genes.

c-Myb Coordinates Survival and the Expression of Genes That Are Critical for the Pre-BCR Checkpoint.

PubMed

Fahl, Shawn P; Daamen, Andrea R; Crittenden, Rowena B; Bender, Timothy P

2018-05-15

The c-Myb transcription factor is required for adult hematopoiesis, yet little is known about c-Myb function during lineage-specific differentiation due to the embryonic lethality of Myb -null mutations. We previously used tissue-specific inactivation of the murine Myb locus to demonstrate that c-Myb is required for differentiation to the pro-B cell stage, survival during the pro-B cell stage, and the pro-B to pre-B cell transition during B lymphopoiesis. However, few downstream mediators of c-Myb-regulated function have been identified. We demonstrate that c-Myb regulates the intrinsic survival of CD19 + pro-B cells in the absence of IL-7 by repressing expression of the proapoptotic proteins Bmf and Bim and that levels of Bmf and Bim mRNA are further repressed by IL-7 signaling in pro-B cells. c-Myb regulates two crucial components of the IL-7 signaling pathway: the IL-7Rα-chain and the negative regulator SOCS3 in CD19 + pro-B cells. Bypassing IL-7R signaling through constitutive activation of Stat5b largely rescues survival of c-Myb-deficient pro-B cells, whereas constitutively active Akt is much less effective. However, rescue of pro-B cell survival is not sufficient to rescue proliferation of pro-B cells or the pro-B to small pre-B cell transition, and we further demonstrate that c-Myb-deficient large pre-B cells are hypoproliferative. Analysis of genes crucial for the pre-BCR checkpoint demonstrates that, in addition to IL-7Rα, the genes encoding λ5, cyclin D3, and CXCR4 are downregulated in the absence of c-Myb, and λ5 is a direct c-Myb target. Thus, c-Myb coordinates survival with the expression of genes that are required during the pre-BCR checkpoint. Copyright © 2018 by The American Association of Immunologists, Inc.

A protein interaction map for cell polarity development

PubMed Central

Drees, Becky L.; Sundin, Bryan; Brazeau, Elizabeth; Caviston, Juliane P.; Chen, Guang-Chao; Guo, Wei; Kozminski, Keith G.; Lau, Michelle W.; Moskow, John J.; Tong, Amy; Schenkman, Laura R.; McKenzie, Amos; Brennwald, Patrick; Longtine, Mark; Bi, Erfei; Chan, Clarence; Novick, Peter; Boone, Charles; Pringle, John R.; Davis, Trisha N.; Fields, Stanley; Drubin, David G.

2001-01-01

Many genes required for cell polarity development in budding yeast have been identified and arranged into a functional hierarchy. Core elements of the hierarchy are widely conserved, underlying cell polarity development in diverse eukaryotes. To enumerate more fully the protein–protein interactions that mediate cell polarity development, and to uncover novel mechanisms that coordinate the numerous events involved, we carried out a large-scale two-hybrid experiment. 68 Gal4 DNA binding domain fusions of yeast proteins associated with the actin cytoskeleton, septins, the secretory apparatus, and Rho-type GTPases were used to screen an array of yeast transformants that express ∼90% of the predicted Saccharomyces cerevisiae open reading frames as Gal4 activation domain fusions. 191 protein–protein interactions were detected, of which 128 had not been described previously. 44 interactions implicated 20 previously uncharacterized proteins in cell polarity development. Further insights into possible roles of 13 of these proteins were revealed by their multiple two-hybrid interactions and by subcellular localization. Included in the interaction network were associations of Cdc42 and Rho1 pathways with proteins involved in exocytosis, septin organization, actin assembly, microtubule organization, autophagy, cytokinesis, and cell wall synthesis. Other interactions suggested direct connections between Rho1- and Cdc42-regulated pathways; the secretory apparatus and regulators of polarity establishment; actin assembly and the morphogenesis checkpoint; and the exocytic and endocytic machinery. In total, a network of interactions that provide an integrated response of signaling proteins, the cytoskeleton, and organelles to the spatial cues that direct polarity development was revealed. PMID:11489916

Cartilage-specific RBPjκ-dependent and -independent Notch signals regulate cartilage and bone development

PubMed Central

Kohn, Anat; Dong, Yufeng; Mirando, Anthony J.; Jesse, Alana M.; Honjo, Tasuku; Zuscik, Michael J.; O’Keefe, Regis J.; Hilton, Matthew J.

2012-01-01

The Notch signaling pathway has emerged as an important regulator of endochondral bone formation. Although recent studies have examined the role of Notch in mesenchymal and chondro-osteo progenitor cell populations, there has yet to be a true examination of Notch signaling specifically within developing and committed chondrocytes, or a determination of whether cartilage and bone formation are regulated via RBPjκ-dependent or -independent Notch signaling mechanisms. To develop a complete understanding of Notch signaling during cartilage and bone development we generated and compared general Notch gain-of-function (Rosa-NICDf/+), RBPjκ-deficient (Rbpjκf/f), and RBPjκ-deficient Notch gain-of-function (Rosa-NICDf/+;Rbpjκf/f) conditional mutant mice, where activation or deletion of floxed alleles were specifically targeted to mesenchymal progenitors (Prx1Cre) or committed chondrocytes (inducible Col2CreERT2). These data demonstrate, for the first time, that Notch regulation of chondrocyte maturation is solely mediated via the RBPjκ-dependent pathway, and that the perichodrium or osteogenic lineage probably influences chondrocyte terminal maturation and turnover of the cartilage matrix. Our study further identifies the cartilage-specific RBPjκ-independent pathway as crucial for the proper regulation of chondrocyte proliferation, survival and columnar chondrocyte organization. Unexpectedly, the RBPjκ-independent Notch pathway was also identified as an important long-range cell non-autonomous regulator of perichondral bone formation and an important cartilage-derived signal required for coordinating chondrocyte and osteoblast differentiation during endochondral bone development. Finally, cartilage-specific RBPjκ-independent Notch signaling likely regulates Ihh responsiveness during cartilage and bone development. PMID:22354840

PPARδ INDUCES CELL PROLIFERATION BY A CYCLIN E1-DEPENDENT MECHANISM AND IS UPREGULATED IN THYROID TUMORS

PubMed Central

Zeng, Lingchun; Geng, Yan; Tretiakova, Maria; Yu, Xuemei; Sicinski, Peter; Kroll, Todd G.

2008-01-01

Peroxisome proliferator-activated receptors (PPARs) are lipid sensing nuclear receptors that have been implicated in multiple physiologic processes including cancer. Here, we determine that PPARδ induces cell proliferation through a novel cyclin E1-dependent mechanism and is upregulated in many human thyroid tumors. The expression of PPARδ was induced coordinately with proliferation in primary human thyroid cells by activation of serum, TSH/cAMP/pKa or EGF/MEK/ERK mitogenic signaling pathways. Engineered overexpression of PPARδ increased thyroid cell number, the incorporation of BrdU and the phosphorylation of Rb 40–45% in just 2 days, one usual cell population doubling. The synthetic PPARδ agonist GW501516 augmented these PPARδ proliferation effects in a dose-dependent manner. Overexpression of PPARδ increased cyclin E1 protein 9-fold, whereas knock down of PPARδ by siRNA reduced both cyclin E1 protein and cell proliferation 2-fold. Induction of proliferation by PPARδ wasabrogated by knockdown of cyclin E1 by siRNA in primary thyroid cells and by knockout of cyclin E1 in mouse embryo fibroblasts, confirming a cyclin E1 dependence for this PPARδ pathway. In addition, the mean expression of native PPARδ was increased 2- to 5-fold (p<0.0001) and correlated with that of the in situ proliferation marker Ki67 (R=0.8571; p=0.02381) in six different classes of benign and malignant human thyroid tumors. Our experiments identify a PPARδ mechanism that induces cell proliferation through cyclin E1 and is regulated by growth factor and lipid signals. The data argue for systematic investigation of PPARδ antagonists as anti-neoplastic agents and implicate altered PPARδ-cyclin E1 signaling in thyroid and other carcinomas. PMID:18701481

Temporally distinct transcriptional regulation of myocyte dedifferentiation and Myofiber growth during muscle regeneration.

PubMed

Louie, Ke'ale W; Saera-Vila, Alfonso; Kish, Phillip E; Colacino, Justin A; Kahana, Alon

2017-11-09

Tissue regeneration requires a series of steps, beginning with generation of the necessary cell mass, followed by cell migration into damaged area, and ending with differentiation and integration with surrounding tissues. Temporal regulation of these steps lies at the heart of the regenerative process, yet its basis is not well understood. The ability of zebrafish to dedifferentiate mature "post-mitotic" myocytes into proliferating myoblasts that in turn regenerate lost muscle tissue provides an opportunity to probe the molecular mechanisms of regeneration. Following subtotal excision of adult zebrafish lateral rectus muscle, dedifferentiating residual myocytes were collected at two time points prior to cell cycle reentry and compared to uninjured muscles using RNA-seq. Functional annotation (GAGE or K-means clustering followed by GO enrichment) revealed a coordinated response encompassing epigenetic regulation of transcription, RNA processing, and DNA replication and repair, along with protein degradation and translation that would rewire the cellular proteome and metabolome. Selected candidate genes were phenotypically validated in vivo by morpholino knockdown. Rapidly induced gene products, such as the Polycomb group factors Ezh2 and Suz12a, were necessary for both efficient dedifferentiation (i.e. cell reprogramming leading to cell cycle reentry) and complete anatomic regeneration. In contrast, the late activated gene fibronectin was important for efficient anatomic muscle regeneration but not for the early step of myocyte cell cycle reentry. Reprogramming of a "post-mitotic" myocyte into a dedifferentiated myoblast requires a complex coordinated effort that reshapes the cellular proteome and rewires metabolic pathways mediated by heritable yet nuanced epigenetic alterations and molecular switches, including transcription factors and non-coding RNAs. Our studies show that temporal regulation of gene expression is programmatically linked to distinct steps in the regeneration process, with immediate early expression driving dedifferentiation and reprogramming, and later expression facilitating anatomical regeneration.

Immune dysregulation in patients with PTEN hamartoma tumor syndrome: Analysis of FOXP3 regulatory T cells.

PubMed

Chen, Hannah H; Händel, Norman; Ngeow, Joanne; Muller, James; Hühn, Michael; Yang, Huei-Ting; Heindl, Mario; Berbers, Roos-Marijn; Hegazy, Ahmed N; Kionke, Janina; Yehia, Lamis; Sack, Ulrich; Bläser, Frank; Rensing-Ehl, Anne; Reifenberger, Julia; Keith, Julia; Travis, Simon; Merkenschlager, Andreas; Kiess, Wieland; Wittekind, Christian; Walker, Lisa; Ehl, Stephan; Aretz, Stefan; Dustin, Michael L; Eng, Charis; Powrie, Fiona; Uhlig, Holm H

2017-02-01

Patients with heterozygous germline mutations in phosphatase and tensin homolog deleted on chromosome 10 (PTEN) experience autoimmunity and lymphoid hyperplasia. Because regulation of the phosphoinositide 3-kinase (PI3K) pathway is critical for maintaining regulatory T (Treg) cell functions, we investigate Treg cells in patients with heterozygous germline PTEN mutations (PTEN hamartoma tumor syndrome [PHTS]). Patients with PHTS were assessed for immunologic conditions, lymphocyte subsets, forkhead box P3 (FOXP3) + Treg cell levels, and phenotype. To determine the functional importance of phosphatases that control the PI3K pathway, we assessed Treg cell induction in vitro, mitochondrial depolarization, and recruitment of PTEN to the immunologic synapse. Autoimmunity and peripheral lymphoid hyperplasia were found in 43% of 79 patients with PHTS. Immune dysregulation in patients with PHTS included lymphopenia, CD4 + T-cell reduction, and changes in T- and B-cell subsets. Although total CD4 + FOXP3 + Treg cell numbers are reduced, frequencies are maintained in the blood and intestine. Despite pathogenic PTEN mutations, the FOXP3 + T cells are phenotypically normal. We show that the phosphatase PH domain leucine-rich repeat protein phosphatase (PHLPP) downstream of PTEN is highly expressed in normal human Treg cells and provides complementary phosphatase activity. PHLPP is indispensable for the differentiation of induced Treg cells in vitro and Treg cell mitochondrial fitness. PTEN and PHLPP form a phosphatase network that is polarized at the immunologic synapse. Heterozygous loss of function of PTEN in human subjects has a significant effect on T- and B-cell immunity. Assembly of the PTEN-PHLPP phosphatase network allows coordinated phosphatase activities at the site of T-cell receptor activation, which is important for limiting PI3K hyperactivation in Treg cells despite PTEN haploinsufficiency. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

A convex optimization approach for identification of human tissue-specific interactomes.

PubMed

Mohammadi, Shahin; Grama, Ananth

2016-06-15

Analysis of organism-specific interactomes has yielded novel insights into cellular function and coordination, understanding of pathology, and identification of markers and drug targets. Genes, however, can exhibit varying levels of cell type specificity in their expression, and their coordinated expression manifests in tissue-specific function and pathology. Tissue-specific/tissue-selective interaction mechanisms have significant applications in drug discovery, as they are more likely to reveal drug targets. Furthermore, tissue-specific transcription factors (tsTFs) are significantly implicated in human disease, including cancers. Finally, disease genes and protein complexes have the tendency to be differentially expressed in tissues in which defects cause pathology. These observations motivate the construction of refined tissue-specific interactomes from organism-specific interactomes. We present a novel technique for constructing human tissue-specific interactomes. Using a variety of validation tests (Edge Set Enrichment Analysis, Gene Ontology Enrichment, Disease-Gene Subnetwork Compactness), we show that our proposed approach significantly outperforms state-of-the-art techniques. Finally, using case studies of Alzheimer's and Parkinson's diseases, we show that tissue-specific interactomes derived from our study can be used to construct pathways implicated in pathology and demonstrate the use of these pathways in identifying novel targets. http://www.cs.purdue.edu/homes/mohammas/projects/ActPro.html mohammadi@purdue.edu. © The Author 2016. Published by Oxford University Press.

Coordination effect-regulated CO2 capture with an alkali metal onium salts/crown ether system

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

Yang, Zhen-Zhen; Jiang, Deen; Zhu, Xiang

2014-01-01

A coordination effect was employed to realize equimolar CO2 absorption, adopting easily synthesized amino group containing absorbents (alkali metal onium salts). The essence of our strategy was to increase the steric hindrance of cations so as to enhance a carbamic acid pathway for CO2 capture. Our easily synthesized alkali metal amino acid salts or phenolates were coordinated with crown ethers, in which highly sterically hindered cations were obtained through a strong coordination effect of crown ethers with alkali metal cations. For example, a CO2 capacity of 0.99 was attained by potassium prolinate/18-crown-6, being characterized by NMR, FT-IR, and quantum chemistrymore » calculations to go through a carbamic acid formation pathway. The captured CO2 can be stripped under very mild conditions (50 degrees C, N-2). Thus, this protocol offers an alternative for the development of technological innovation towards efficient and low energy processes for carbon capture and sequestration.« less

Biochemical evidence for Ku-independent backup pathways of NHEJ.

PubMed

Wang, Huichen; Perrault, Ange Ronel; Takeda, Yoshihiko; Qin, Wei; Wang, Hongyan; Iliakis, George

2003-09-15

Cells of higher eukaryotes process within minutes double strand breaks (DSBs) in their genome using a non-homologous end joining (NHEJ) apparatus that engages DNA-PKcs, Ku, DNA ligase IV, XRCC4 and other as of yet unidentified factors. Although chemical inhibition, or mutation, in any of these factors delays processing, cells ultimately remove the majority of DNA DSBs using an alternative pathway operating with an order of magnitude slower kinetics. This alternative pathway is active in mutants deficient in genes of the RAD52 epistasis group and frequently joins incorrect ends. We proposed, therefore, that it reflects an alternative form of NHEJ that operates as a backup (B-NHEJ) to the DNA-PK-dependent (D-NHEJ) pathway, rather than homology directed repair of DSBs. The present study investigates the role of Ku in the coordination of these pathways using as a model end joining of restriction endonuclease linearized plasmid DNA in whole cell extracts. Efficient, error-free, end joining observed in such in vitro reactions is strongly inhibited by anti-Ku antibodies. The inhibition requires DNA-PKcs, despite the fact that Ku efficiently binds DNA ends in the presence of antibodies, or in the absence of DNA-PKcs. Strong inhibition of DNA end joining is also mediated by wortmannin, an inhibitor of DNA-PKcs, in the presence but not in the absence of Ku, and this inhibition can be rescued by pre-incubating the reaction with double stranded oligonucleotides. The results are compatible with a role of Ku in directing end joining to a DNA-PK dependent pathway, mediated by efficient end binding and productive interactions with DNA-PKcs. On the other hand, efficient end joining is observed in extracts of cells lacking DNA-PKcs, as well as in Ku-depleted extracts in line with the operation of alternative pathways. Extracts depleted of Ku and DNA-PKcs rejoin blunt ends, as well as homologous ends with 3' or 5' protruding single strands with similar efficiency, but addition of Ku suppresses joining of blunt ends and homologous ends with 3' overhangs. We propose that the affinity of Ku for DNA ends, particularly when cooperating with DNA-PKcs, suppresses B-NHEJ by quickly and efficiently binding DNA ends and directing them to D-NHEJ for rapid joining. A chromatin-based model of DNA DSB rejoining accommodating biochemical and genetic results is presented and deviations between in vitro and in vivo results discussed.

Biochemical evidence for Ku-independent backup pathways of NHEJ

PubMed Central

Wang, Huichen; Perrault, Ange Ronel; Takeda, Yoshihiko; Qin, Wei; Wang, Hongyan; Iliakis, George

2003-01-01

Cells of higher eukaryotes process within minutes double strand breaks (DSBs) in their genome using a non-homologous end joining (NHEJ) apparatus that engages DNA-PKcs, Ku, DNA ligase IV, XRCC4 and other as of yet unidentified factors. Although chemical inhibition, or mutation, in any of these factors delays processing, cells ultimately remove the majority of DNA DSBs using an alternative pathway operating with an order of magnitude slower kinetics. This alternative pathway is active in mutants deficient in genes of the RAD52 epistasis group and frequently joins incorrect ends. We proposed, therefore, that it reflects an alternative form of NHEJ that operates as a backup (B-NHEJ) to the DNA-PK-dependent (D-NHEJ) pathway, rather than homology directed repair of DSBs. The present study investigates the role of Ku in the coordination of these pathways using as a model end joining of restriction endonuclease linearized plasmid DNA in whole cell extracts. Efficient, error-free, end joining observed in such in vitro reactions is strongly inhibited by anti-Ku antibodies. The inhibition requires DNA-PKcs, despite the fact that Ku efficiently binds DNA ends in the presence of antibodies, or in the absence of DNA-PKcs. Strong inhibition of DNA end joining is also mediated by wortmannin, an inhibitor of DNA-PKcs, in the presence but not in the absence of Ku, and this inhibition can be rescued by pre-incubating the reaction with double stranded oligonucleotides. The results are compatible with a role of Ku in directing end joining to a DNA-PK dependent pathway, mediated by efficient end binding and productive interactions with DNA-PKcs. On the other hand, efficient end joining is observed in extracts of cells lacking DNA-PKcs, as well as in Ku-depleted extracts in line with the operation of alternative pathways. Extracts depleted of Ku and DNA-PKcs rejoin blunt ends, as well as homologous ends with 3′ or 5′ protruding single strands with similar efficiency, but addition of Ku suppresses joining of blunt ends and homologous ends with 3′ overhangs. We propose that the affinity of Ku for DNA ends, particularly when cooperating with DNA-PKcs, suppresses B-NHEJ by quickly and efficiently binding DNA ends and directing them to D-NHEJ for rapid joining. A chromatin-based model of DNA DSB rejoining accommodating biochemical and genetic results is presented and deviations between in vitro and in vivo results discussed. PMID:12954774

Signal transduction networks and the biology of plant cells.

PubMed

Chrispeels, M J; Holuigue, L; Latorre, R; Luan, S; Orellana, A; Peña-Cortes, H; Raikhel, N V; Ronald, P C; Trewavas, A

1999-01-01

The development of plant transformation in the mid-1980s and of many new tools for cell biology, molecular genetics, and biochemistry has resulted in enormous progress in plant biology in the past decade. With the completion of the genome sequence of Arabidopsis thaliana just around the corner, we can expect even faster progress in the next decade. The interface between cell biology and signal transduction is emerging as a new and important field of research. In the past we thought of cell biology strictly in terms of organelles and their biogenesis and function, and researchers focused on questions such as, how do proteins enter chloroplasts? or, what is the structure of the macromolecules of the cell wall and how are these molecules secreted? Signal transduction dealt primarily with the perception of light (photomorphogenesis) or hormones and with the effect such signals have on enhancing the activity of specific genes. Now we see that the fields of cell biology and signal transduction are merging because signals pass between organelles and a single signal transduction pathway usually involves multiple organelles or cellular structures. Here are some examples to illustrate this new paradigm. How does abscisic acid (ABA) regulate stomatal closure? This pathway involves not only ABA receptors whose location is not yet known, but cation and anion channels in the plasma membrane, changes in the cytoskeleton, movement of water through water channels in the tonoplast and the plasma membrane, proteins with a farnesyl tail that can be located either in the cytosol or attached to a membrane, and probably unidentified ion channels in the tonoplast. In addition there are highly localized calcium oscillations in the cytoplasm resulting from the release of calcium stored in various compartments. The activities of all these cellular structures need to be coordinated during ABA-induced stomatal closure. For another example of the interplay between the proteins of signal transduction pathways and cytoplasmic structures, consider how plants mount defense responses against pathogens. Elicitors produced by pathogens bind to receptors on the plant plasma membrane or in the cytosol and eventually activate a large number of genes. This results in the coordination of activities at the plasma membrane (production of reactive oxygen species), in the cytoskeleton, localized calcium oscillations, and the modulation of protein kinases and protein phosphatases whose locations remain to be determined. The movement of transcription factors into the nucleus to activate the defense genes requires their release from cytosolic anchors and passage through the nuclear pore complexes of the nuclear envelope. This review does not cover all the recent progress in plant signal transduction and cell biology; it is confined to the topics that were discussed at a recent (November 1998) workshop held in Santiago at which lecturers from Chile, the USA and the UK presented recent results from their laboratories.

Identification of genes involved in Ca2+ ionophore A23187-mediated apoptosis and demonstration of a high susceptibility for transcriptional repression of cell cycle genes in B lymphoblasts from a patient with Scott syndrome

PubMed Central

Kozian, Detlef; Proulle, Valérie; Nitsche, Almut; Galitzine, Marie; Martinez, Marie-Carmen; Schumann, Beatrice; Meyer, Dominique; Herrmann, Matthias; Freyssinet, Jean-Marie; Kerbiriou-Nabias, Danièle

2005-01-01

Background In contrast to other agents able to induce apoptosis of cultured cells, Ca2+ ionophore A23187 was shown to elicit direct activation of intracellular signal(s). The phenotype of the cells derived from patients having the hemorrhagic disease Scott syndrome, is associated with an abnormally high proportion of apoptotic cells, both in basal culture medium and upon addition of low ionophore concentrations in long-term cultures. These features are presumably related to the mutation also responsible for the defective procoagulant plasma membrane remodeling. We analyzed the specific transcriptional re-programming induced by A23187 to get insights into the effect of this agent on gene expression and a defective gene regulation in Scott cells. Results The changes in gene expression upon 48 hours treatment with 200 nM A23187 were measured in Scott B lymphoblasts compared to B lymphoblasts derived from the patient's daughter or unrelated individuals using Affymetrix microarrays. In a similar manner in all of the B cell lines, results showed up-regulation of 55 genes, out of 12,000 represented sequences, involved in various pathways of the cell metabolism. In contrast, a group of 54 down-regulated genes, coding for histones and proteins involved in the cell cycle progression, was more significantly repressed in Scott B lymphoblasts than in the other cell lines. These data correlated with the alterations of the cell cycle phases in treated cells and suggested that the potent effect of A23187 in Scott B lymphoblasts may be the consequence of the underlying molecular defect. Conclusion The data illustrate that the ionophore A23187 exerts its pro-apoptotic effect by promoting a complex pattern of genetic changes. These results also suggest that a subset of genes participating in various steps of the cell cycle progress can be transcriptionally regulated in a coordinated fashion. Furthermore, this research brings a new insight into the defect in cultured Scott B lymphoblasts, leading to hypothesize that a mutated gene plays a role not only in membrane remodeling but also in signal transduction pathway(s) leading to altered transcriptional regulation of cell cycle genes. PMID:16242039

Transcriptional and post-transcriptional regulation of the ionizing radiation response by ATM and p53

PubMed Central

Venkata Narayanan, Ishwarya; Paulsen, Michelle T.; Bedi, Karan; Berg, Nathan; Ljungman, Emily A.; Francia, Sofia; Veloso, Artur; Magnuson, Brian; di Fagagna, Fabrizio d’Adda; Wilson, Thomas E.; Ljungman, Mats

2017-01-01

In response to ionizing radiation (IR), cells activate a DNA damage response (DDR) pathway to re-program gene expression. Previous studies using total cellular RNA analyses have shown that the stress kinase ATM and the transcription factor p53 are integral components required for induction of IR-induced gene expression. These studies did not distinguish between changes in RNA synthesis and RNA turnover and did not address the role of enhancer elements in DDR-mediated transcriptional regulation. To determine the contribution of synthesis and degradation of RNA and monitor the activity of enhancer elements following exposure to IR, we used the recently developed Bru-seq, BruChase-seq and BruUV-seq techniques. Our results show that ATM and p53 regulate both RNA synthesis and stability as well as enhancer element activity following exposure to IR. Importantly, many genes in the p53-signaling pathway were coordinately up-regulated by both increased synthesis and RNA stability while down-regulated genes were suppressed either by reduced synthesis or stability. Our study is the first of its kind that independently assessed the effects of ionizing radiation on transcription and post-transcriptional regulation in normal human cells. PMID:28256581

Proximity-Directed Labeling Reveals a New Rapamycin-Induced Heterodimer of FKBP25 and FRB in Live Cells

PubMed Central

2016-01-01

Mammalian target of rapamycin (mTOR) signaling is a core pathway in cellular metabolism, and control of the mTOR pathway by rapamycin shows potential for the treatment of metabolic diseases. In this study, we employed a new proximity biotin-labeling method using promiscuous biotin ligase (pBirA) to identify unknown elements in the rapamycin-induced interactome on the FK506-rapamycin binding (FRB) domain in living cells. FKBP25 showed the strongest biotin labeling by FRB–pBirA in the presence of rapamycin. Immunoprecipitation and immunofluorescence experiments confirmed that endogenous FKBP25 has a rapamycin-induced physical interaction with the FRB domain. Furthermore, the crystal structure of the ternary complex of FRB–rapamycin–FKBP25 was determined at 1.67-Å resolution. In this crystal structure we found that the conformational changes of FRB generate a hole where there is a methionine-rich space, and covalent metalloid coordination was observed at C2085 of FRB located at the bottom of the hole. Our results imply that FKBP25 might have a unique physiological role related to metallomics in mTOR signaling. PMID:27610411

Mediator can regulate mitotic entry and direct periodic transcription in fission yeast.

PubMed

Banyai, Gabor; Lopez, Marcela Davila; Szilagyi, Zsolt; Gustafsson, Claes M

2014-11-01

Cdk8 is required for correct timing of mitotic progression in fission yeast. How the activity of Cdk8 is regulated is unclear, since the kinase is not activated by T-loop phosphorylation and its partner, CycC, does not oscillate. Cdk8 is, however, a component of the multiprotein Mediator complex, a conserved coregulator of eukaryotic transcription that is connected to a number of intracellular signaling pathways. We demonstrate here that other Mediator components regulate the activity of Cdk8 in vivo and thereby direct the timing of mitotic entry. Deletion of Mediator components Med12 and Med13 leads to higher cellular Cdk8 protein levels, premature phosphorylation of the Cdk8 target Fkh2, and earlier entry into mitosis. We also demonstrate that Mediator is recruited to clusters of mitotic genes in a periodic fashion and that the complex is required for the transcription of these genes. We suggest that Mediator functions as a hub for coordinated regulation of mitotic progression and cell cycle-dependent transcription. The many signaling pathways and activator proteins shown to function via Mediator may influence the timing of these cell cycle events. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Drosophila melanogaster Activating Transcription Factor 4 Regulates Glycolysis During Endoplasmic Reticulum Stress

PubMed Central

Lee, Ji Eun; Oney, McKenna; Frizzell, Kimberly; Phadnis, Nitin; Hollien, Julie

2015-01-01

Endoplasmic reticulum (ER) stress results from an imbalance between the load of proteins entering the secretory pathway and the ability of the ER to fold and process them. The response to ER stress is mediated by a collection of signaling pathways termed the unfolded protein response, which plays important roles in development and disease. Here we show that in Drosophila melanogaster S2 cells, ER stress induces a coordinated change in the expression of genes involved in carbon metabolism. Genes encoding enzymes that carry out glycolysis were up-regulated, whereas genes encoding proteins in the tricarboxylic acid cycle and respiratory chain complexes were down-regulated. The unfolded protein response transcription factor Atf4 was necessary for the up-regulation of glycolytic enzymes and Lactate dehydrogenase (Ldh). Furthermore, Atf4 binding motifs in promoters for these genes could partially account for their regulation during ER stress. Finally, flies up-regulated Ldh and produced more lactate when subjected to ER stress. Together, these results suggest that Atf4 mediates a shift from a metabolism based on oxidative phosphorylation to one more heavily reliant on glycolysis, reminiscent of aerobic glycolysis or the Warburg effect observed in cancer and other proliferative cells. PMID:25681259